EP4157460A1 - Methods of treating aging-related disorders - Google Patents

Methods of treating aging-related disorders

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Publication number
EP4157460A1
EP4157460A1 EP21733685.8A EP21733685A EP4157460A1 EP 4157460 A1 EP4157460 A1 EP 4157460A1 EP 21733685 A EP21733685 A EP 21733685A EP 4157460 A1 EP4157460 A1 EP 4157460A1
Authority
EP
European Patent Office
Prior art keywords
target
soluble
binding domain
receptor
tgf
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
EP21733685.8A
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German (de)
English (en)
French (fr)
Inventor
Hing C. Wong
Xiaoyun Zhu
Bai LIU
Pallavi CHATURVEDI
Varghese George
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Immunitybio Inc
Original Assignee
HCW Biologics Inc
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Filing date
Publication date
Priority claimed from PCT/US2020/035598 external-priority patent/WO2021247003A1/en
Application filed by HCW Biologics Inc filed Critical HCW Biologics Inc
Publication of EP4157460A1 publication Critical patent/EP4157460A1/en
Pending legal-status Critical Current

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/177—Receptors; Cell surface antigens; Cell surface determinants
    • A61K38/179—Receptors; Cell surface antigens; Cell surface determinants for growth factors; for growth regulators
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/177—Receptors; Cell surface antigens; Cell surface determinants
    • A61K38/1793—Receptors; Cell surface antigens; Cell surface determinants for cytokines; for lymphokines; for interferons
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/19—Cytokines; Lymphokines; Interferons
    • A61K38/20—Interleukins [IL]
    • A61K38/2086—IL-13 to IL-16
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/36—Blood coagulation or fibrinolysis factors
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P39/00—General protective or antinoxious agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2300/00—Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00

Definitions

  • Senescence was first described in cultured human fibroblast cells that lost their ability to proliferate, reaching permanent arrest after about 50 population doublings (referred to as the Hayflick limit). Senescence is considered a stress response that can be induced by a wide range of intrinsic and extrinsic insults, including oxidative and genotoxic stress, DNA damage, telomere attrition, oncogenic activation, mitochondrial dysfunction, or chemotherapeutic agents. Senescent cells remain metabolically active and can influence tissue hemostasis, disease, and aging through their secretory phenotype. Senescence is considered as a physiologic process and is important in promoting wound healing, tissue homeostasis, regeneration, and regulation of fibrosis.
  • transient induction of senescent cells is observed during would healing and contributes to wound resolution.
  • Senescence also plays a role in tumor suppression.
  • the accumulation of senescent cells also drives aging and aging-related diseases and conditions.
  • the senescent phenotype also can trigger chronic inflammatory responses and consequently augment chronic inflammatory conditions to promote tumor growth.
  • the connection between senescence and aging was initially based on the observation that senescent cells accumulate in aged tissue.
  • the use of transgenic models has enabled the detection of senescent cells systematically in many aging-related disorders. Strategies to selectively eliminate senescent cells have demonstrated that senescent cells play a causal role in aging-related disorders.
  • immune cells are the effector cells to remove senescent cells naturally after the fulfillment of senescent-cell physiological roles.
  • the weakening of the immune system during the aging process allows the accumulation of senescent cells.
  • the present invention is based on the discovery that subcutaneous administration of an agent that results in a decrease in the activation of a TGF- ⁇ receptor or a common gamma-chain family cytokine receptor activating agent (e.g., complexes of gamma-chain cytokines and their cognate receptors) to a mammal promotes and activates immune cells to regain their capabilities of reducing senescent cells in vivo effectively, selectively, and safely.
  • a TGF- ⁇ receptor or a common gamma-chain family cytokine receptor activating agent e.g., complexes of gamma-chain cytokines and their cognate receptors
  • methods of killing or reducing the number of naturally-occurring and/or treatment-induced senescent cells in a subject that include administering to the subject a therapeutically effective amount of one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor.
  • methods of decreasing the accumulation of naturally-occurring and/or treatment-induced senescent cells in a subject that include administering to the subject a therapeutically effective amount of one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor.
  • SASP senescence associated secretory phenotype
  • common gamma-chain family cytokine receptor activating agent(s) e.g., complexes of gamma- chain cytokines and their cognate receptors.
  • the present invention is also based on the discovery that administration of NK cell activating agents to a mammal having a cancer resulted in a tumor inhibition and administration of NK cell activating agents to a diabetic animal model demonstrated improved skin and hair appearance and texture, and decreased blood glucose levels.
  • NK natural killer
  • s natural killer cell activating agent
  • methods of treating an aging-related disease or condition in a subject in need thereof that include administering to a subject identified as having an aging-related disease or condition a therapeutically effective amount of one or more natural killer (NK) cell activating agent (s) and/or a therapeutically effective number of activated NK cells.
  • methods of killing or reducing the number of senescent cells in a subject in need thereof that include administering to the subject a therapeutically effective amount of one or more NK cell activating agent(s) and/or or a therapeutically effective number of activated NK cells.
  • NK natural killer
  • methods of killing or reducing the number of naturally- occurring and/or treatment-induced senescent cells in a subject that include administering to the subject a therapeutically effective amount of one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor.
  • methods of decreasing the accumulation of naturally- occurring and/or treatment-induced senescent cells in a subject that include administering to the subject a therapeutically effective amount of one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor.
  • the subject has been previously diagnosed or identified as having an aging-related disease or an inflammatory disease.
  • the aging-related disease is inflamm-aging related.
  • the aging-related disease is selected from the group of: Alzheimer’s disease, aneurysm, cystic fibrosis, fibrosis in pancreatitis, glaucoma, hypertension, inflammatory bowel disease, intervertebral disc degeneration, osteoarthritis, type 2 diabetes mellitus, adipose atrophy, lipodystrophy, atherosclerosis, cataracts, COPD, idiopathic pulmonary fibrosis, kidney transplant failure, liver fibrosis, loss of bone mass, myocardial infarction, sarcopenia, wound healing, alopecia, cardiomyocyte hypertrophy, osteoarthritis, Parkinson’s disease, age-associated loss of lung tissue elasticity, age-related macular degeneration, cachexia, glomerulosclerosis, liver cirrhosis, NAFLD, osteoporosis, amyotrophic lateral sclerosis, Huntington’s disease, spinocerebellar ataxi
  • the aging- related disease is a cancer selected from the group consisting of: solid tumor, hematological tumor, sarcoma, osteosarcoma, glioblastoma, neuroblastoma, melanoma, rhabdomyosarcoma, Ewing sarcoma, osteosarcoma, B-cell neoplasms, multiple myeloma, B-cell lymphoma, B-cell non-Hodgkin’s lymphoma, Hodgkin’s lymphoma, chronic lymphocytic leukemia (CLL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), myelodysplastic syndromes (MDS), cutaneous T-cell lymphoma, retinoblastoma, stomach cancer, urothelial carcinoma, lung cancer, renal cell carcinoma, gastric and esophageal cancer
  • CLL chronic lymphocytic leuk
  • the inflammatory disease is selected from the group of: rheumatoid arthritis, inflammatory bowel disease, lupus erythematosus, lupus nephritis, diabetic nephropathy, CNS injury, Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, Crohn’s disease, multiple sclerosis, Guillain-Barre syndrome, psoriasis, Grave’s disease, ulcerative colitis, nonalcoholic steatohepatitis, mood disorders and cancer treatment-related cognitive impairment.
  • the treatment- induced senescent cells are chemotherapy-induced senescent cells.
  • the administration of the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor results in a decrease in the number or activity of naturally-occurring senescent cells and/or treatment-induced senescent cells in a target tissue in the subject.
  • the target tissue is selected from the group of: adipose tissue, pancreatic tissue, liver tissue, kidney tissue, lung tissue, heart tissue, vasculature, bone tissue, central nervous system (CNS) tissue, eye tissue, skin tissue, muscle tissue, and secondary lympho-organ tissue.
  • the TGF ⁇ receptor is a TGF- ⁇ receptor II (TGF ⁇ RII). In some embodiments of any of the methods described herein, the TGF ⁇ receptor is a TGF ⁇ RIII. In some embodiments of any of the methods described herein, at least one of the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor is a soluble TGF- ⁇ receptor, an extracellular domain of TGF- ⁇ receptor, an antibody that binds specifically to TGF- ⁇ , an antagonistic antibody that binds to a TGF- ⁇ receptor, an agent that binds to a latency-associated peptide (“LAP”), or an agent that binds to a TGF- ⁇ /LAP complex.
  • LAP latency-associated peptide
  • the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor decrease(s) the activation of a TGF- ⁇ receptor through binding to a LAP, or to a TGF- ⁇ /LAP complex.
  • At least one of the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor is a multi-chain chimeric polypeptide including: (a) a first chimeric polypeptide comprising: (i) a first target-binding domain; (ii) a soluble tissue factor domain; and (iii) a first domain of a pair of affinity domains; (b) a second chimeric polypeptide comprising: (i) a second domain of a pair of affinity domains; and (ii) a second target-binding domain, where one or both of the first target-binding domain and the second target-binding domain binds specifically to a ligand of a TGF- ⁇ receptor; or one or both of the first target-binding domain and the second target-binding domain is an antagonistic antigen- binding domain that binds specifically to a TGF- ⁇ receptor.
  • the TGF- ⁇ receptor is TGF ⁇ RII. In some embodiments of any of the methods described herein, the TGF- ⁇ receptor is TGF ⁇ RIII. In some embodiments of any of the methods described herein, the first target- binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide. In some embodiments of any of the methods described herein, the first chimeric polypeptide further includes a linker sequence between the first target- binding domain and the soluble tissue factor domain in the first chimeric polypeptide. In some embodiments of any of the methods described herein, the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the first target- binding domain and the second target-binding domain bind specifically to the same antigen. In some embodiments of any of the methods described herein, the first target- binding domain and the second target-binding domain bind specifically to different antigens. In some embodiments of any of the methods described herein, the first chimeric polypeptide further comprises one or more additional target-binding domain(s). In some embodiments of any of the methods described herein, the second chimeric polypeptide further comprises one or more additional target-binding domain(s). In some embodiments of any of the methods described herein, the soluble tissue factor domain is a soluble human tissue factor domain.
  • the soluble human tissue factor domain comprises a sequence that is at least 80% identical to SEQ ID NO: 93.
  • the pair of affinity domains is a sushi domain from an alpha chain of human IL-15 receptor (IL15R ⁇ ) and a soluble IL-15.
  • the soluble IL-15 has a D8N or D8A amino acid substitution.
  • the soluble IL-15 comprises a mutation to reduce or eliminate IL-15 activity.
  • the pair of affinity domains is selected from the group of: barnase and barnstar, a PKA and an AKAP, adapter/docking tag modules based on mutated RNase I fragments, and SNARE modules based on interactions of the proteins syntaxin, synaptotagmin, synaptobrevin, and SNAP25.
  • the first domain or the second domain of a pair of affinity domains is a soluble common gamma-chain family cytokine or an antigen-binding domain that binds specifically to a common gamma-chain family cytokine receptor.
  • the first target- binding domain and/or the second target-binding domain include a soluble TGF- ⁇ receptor.
  • the soluble TGF- ⁇ receptor is a soluble TGF ⁇ RII.
  • the soluble TGF ⁇ RII includes a first sequence that is at least 80% identical to SEQ ID NO: 183, and a second sequence that is at least 80% identical to SEQ ID NO: 183, wherein the first and second sequence are separated by a linker.
  • the soluble TGF ⁇ RII comprises a first sequence that is at least 90% identical to SEQ ID NO: 183, and a second sequence that is at least 90% identical to SEQ ID NO: 183. In some embodiments of any of the methods described herein, the soluble TGF ⁇ RII includes a first sequence of SEQ ID NO: 183, and a second sequence of SEQ ID NO: 183. In some embodiments of any of the methods described herein, the linker includes a sequence of SEQ ID NO: 102. In some embodiments of any of the methods described herein, the soluble TGF ⁇ RII includes a sequence that is at least 80% identical to SEQ ID NO: 188.
  • the soluble TGF ⁇ RII includes a sequence that is at least 90% identical to SEQ ID NO: 188. In some embodiments of any of the methods described herein, the soluble TGF- ⁇ RII includes a sequence of SEQ ID NO: 188. In some embodiments of any of the methods described herein, the first chimeric polypeptide includes a sequence that is at least 80% identical to SEQ ID NO: 236. In some embodiments of any of the methods described herein, the first chimeric polypeptide includes a sequence that is at least 90% identical to SEQ ID NO: 236. In some embodiments of any of the methods described herein, the first chimeric polypeptide includes a sequence of SEQ ID NO: 236.
  • the second chimeric polypeptide includes a sequence that is at least 80% identical to SEQ ID NO: 193. In some embodiments of any of the methods described herein, the first chimeric polypeptide includes a sequence that is at least 80% identical to SEQ ID NO: 236. In some embodiments of any of the methods described herein, the second chimeric polypeptide includes a sequence that is at least 90% identical to SEQ ID NO: 193. In some embodiments of any of the methods described herein, the second chimeric polypeptide includes a sequence of SEQ ID NO: 193. In some embodiments of any of the methods described herein, the first chimeric polypeptide comprises a sequence of SEQ ID NO: 236.
  • At least one of the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor is a single-chain chimeric polypeptide including: (i) a first target-binding domain; (ii) a soluble tissue factor domain; and (iii) a second target-binding domain, wherein one or both of the first target-binding domain and the second target-binding domain binds specifically to a ligand of a TGF- ⁇ receptor; or one or both of the first target-binding domain and the second target-binding domain is an antagonistic antigen-binding domain that binds specifically to a TGF- ⁇ receptor.
  • the TGF- ⁇ receptor is TGF- ⁇ RII. In some embodiments of any of the methods described herein, the TGF- ⁇ receptor is TGF ⁇ RIII. In some embodiments of any of the methods described herein, the first target- binding domain and the soluble tissue factor domain directly abut each other. In some embodiments of any of the methods described herein, the single-chain chimeric polypeptide further includes a linker sequence between the first target-binding domain and the soluble tissue factor domain. In some embodiments of any of the methods described herein, the soluble tissue factor domain and the second target-binding domain directly abut each other.
  • the single-chain chimeric polypeptide further includes a linker sequence between the soluble tissue factor domain and the second target-binding domain.
  • the first target- binding domain and the second target-binding domain bind specifically to the same antigen.
  • the first target- binding domain and the second target-binding domain bind specifically to different antigens.
  • the soluble tissue factor domain is a soluble human tissue factor domain.
  • the soluble human tissue factor domain includes a sequence that is at least 80% identical to SEQ ID NO: 93.
  • the single-chain chimeric polypeptide further includes one or more additional target-binding domains at its N- and/or C-terminus.
  • the first target-binding domain and/or the second target-binding domain comprise a soluble TGF- ⁇ receptor.
  • the soluble TGF- ⁇ receptor is a soluble TGF- ⁇ RII.
  • the soluble TGF- ⁇ RII includes a first sequence that is at least 80% identical to SEQ ID NO: 183, and a second sequence that is at least 80% identical to SEQ ID NO: 183, wherein the first and second sequence are separated by a linker.
  • the soluble TGF ⁇ RII includes a first sequence that is at least 90% identical to SEQ ID NO: 183, and a second sequence that is at least 90% identical to SEQ ID NO: 183.
  • the soluble TGF ⁇ RII includes a first sequence of SEQ ID NO: 183, and a second sequence of SEQ ID NO: 183.
  • the linker includes a sequence of SEQ ID NO: 102. In some embodiments of any of the methods described herein, the soluble TGF ⁇ RII includes a sequence that is at least 80% identical to SEQ ID NO: 188. In some embodiments of any of the methods described herein, the soluble TGF ⁇ RII includes a sequence that is at least 90% identical to SEQ ID NO: 188. In some embodiments of any of the methods described herein, the soluble TGF ⁇ RII includes a sequence of SEQ ID NO: 188.
  • the method includes administering two or more doses of the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor to the subject.
  • any two consecutive doses of the two or more doses are administered about 1 week to about one year apart.
  • any two consecutive doses of the two or more doses are administered about 1 week to about 6 months apart.
  • any two consecutive doses of the two or more doses are administered about 1 week to about 2 months apart.
  • any two consecutive doses of the two or more doses are administered about 1 week to about 1 month apart. In some embodiments of any of the methods described herein, the two or more doses are administered by subcutaneous administration. In some embodiments of any of the methods described herein, the two or more doses are administered by intramuscular administration. In some embodiments of any of the methods described herein, the two or more doses are administered over a period of time of about 1 year to about 60 years. In some embodiments of any of the methods described herein, the two or more doses are administered over a period of time of about 1 year to about 50 years.
  • the two or more doses are administered over a period of time of about 1 year to about 40 years. In some embodiments of any of the methods described herein, the two or more doses are administered over a period of time of about 1 year to about 30 years. In some embodiments of any of the methods described herein, the two or more doses are administered over a period of time of about 1 year to about 20 years. In some embodiments of any of the methods described herein, the two or more doses are administered over a period of time of about 1 year to about 10 years.
  • a first dose of the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor begins when the subject reaches an age of at least 30 years. In some embodiments of any of the methods described herein, a first dose of the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor begins when the subject reaches an age of at least 40 years. In some embodiments of any of the methods described herein, a first dose of the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor begins when the subject reaches an age of at least 50 years.
  • a first dose of the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor begins when the subject reaches an age of at least 60 years.
  • each of the two or more doses are administered at a dosage of about 0.01 mg of each agent that results in a decrease in the activation of a TGF- ⁇ receptor/kg to about 10 mg of each agent that results in a decrease in the activation of a TGF- ⁇ receptor/kg.
  • each of the two or more doses are administered at a dosage of about 0.02 mg of each agent that results in a decrease in the activation of a TGF- ⁇ receptor/kg to about 5 mg of each agent that results in a decrease in the activation of a TGF- ⁇ receptor/kg.
  • the subject is not diagnosed or identified as having an aging-related disease or an inflammatory disease.
  • the subject has not been previously treated with a chemotherapeutic agent.
  • the subject has not been previously treated with a therapeutic agent that induces cellular senescence.
  • NK natural killer
  • the senescent cells are senescent cancer cells, senescent monocytes, senescent lymphocytes, senescent astrocytes, senescent microglia, senescent neurons, senescent tissue fibroblasts, senescent dermal fibroblasts, senescent keratinocytes, or other differentiated tissue-specific dividing functional cells.
  • the senescent cancer cells are chemotherapy-induced senescent cells or radiation-induced senescent cells.
  • the subject has been identified or diagnosed as having an aging-related disease or condition.
  • the aging-related disease or condition is selected from the group of: a cancer, an autoimmune disease, a metabolic disease, a neurodegenerative disease, a cardiovascular disease, a skin disease, a progeria disease, and a fragility disease.
  • the cancer is selected from the group of: solid tumor, hematological tumor, sarcoma, osteosarcoma, glioblastoma, neuroblastoma, melanoma, rhabdomyosarcoma, Ewing sarcoma, osteosarcoma, B-cell neoplasms, multiple myeloma, B-cell lymphoma, B-cell non-Hodgkin’s lymphoma, Hodgkin’s lymphoma, chronic lymphocytic leukemia (CLL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), myelodysplastic syndromes (MDS), cutaneous T-cell lymphoma, retinoblastoma, stomach cancer, urothelial carcinoma, lung cancer, renal cell carcinoma, gastric and esophageal cancer, pancreatic cancer, prostate cancer,
  • CLL chronic lymphocytic leuk
  • the autoimmune disease is type-1 diabetes.
  • the metabolic disease is selected from the group of: obesity, a lipodystrophy, and type-2 diabetes mellitus.
  • the neurodegenerative disease is selected from the group of: Alzheimer’s disease, Parkinson’s disease, and dementia.
  • the cardiovascular disease is selected from the group of: coronary artery disease, atherosclerosis, and pulmonary arterial hypertension.
  • the skin disease is selected from the group of: wound healing, alopecia, wrinkles, senile lentigo, skin thinning, xeroderma pigmentosum, and dyskeratosis congenita.
  • the progeria disease is selected from the group of: progeria and Hutchinson-Gilford Progeria Syndrome.
  • the fragility disease is selected from the group of: frailty, responsiveness to vaccination, osteoporosis, and sarcopenia.
  • the aging-related disease or condition is selected from the group of: osteoarthritis, adipose atrophy, idiopathic pulmonary fibrosis, kidney transplant failure, liver fibrosis, loss of bone mass, sarcopenia, age-associated loss of lung tissue elasticity, osteoporosis, age-associated renal dysfunction, and chemical-induced renal dysfunction.
  • the aging-related disease or condition is type-2 diabetes or atherosclerosis.
  • the administering results in a decrease in the number of senescent cells in a target tissue in the subject.
  • the target tissue is selected from the group of: adipose tissue, pancreatic tissue, liver tissue, lung tissue, vasculature, bone tissue, central nervous system (CNS) tissue, eye tissue, skin tissue, muscle tissue, and secondary lympho-organ tissue.
  • the administering results in an increase in the expression levels of CD25, CD69, mTORC1, SREBP1, IFN- ⁇ , and granzyme B in activated NK cells.
  • methods of treating an aging-related disease or condition in a subject in need thereof that include administering to a subject identified as having an aging-related disease or condition a therapeutically effective number of activated NK cells.
  • the senescent cells are senescent cancer cells, senescent monocytes, senescent lymphocytes, senescent astrocytes, senescent microglia, senescent neurons, senescent tissue fibroblasts, senescent dermal fibroblasts, senescent keratinocytes, or other differentiated tissue-specific dividing functional cells.
  • the senescent cancer cells are chemotherapy-induced senescent cells or radiation-induced senescent cells.
  • the subject has been identified or diagnosed as having an aging-related disease or condition.
  • the aging-related disease or condition is selected from the group of: a cancer, an autoimmune disease, a metabolic disease, a neurodegenerative disease, a cardiovascular disease, a skin disease, a progeria disease, and a fragility disease.
  • the cancer is selected from the group of: solid tumor, hematological tumor, sarcoma, osteosarcoma, glioblastoma, neuroblastoma, melanoma, rhabdomyosarcoma, Ewing sarcoma, osteosarcoma, B-cell neoplasms, multiple myeloma, B-cell lymphoma, B-cell non-Hodgkin’s lymphoma, Hodgkin’s lymphoma, chronic lymphocytic leukemia (CLL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), myelodysplastic syndromes (MDS), cutaneous T-cell lymphoma, retinoblastoma, stomach cancer, urothelial carcinoma, lung cancer, renal cell carcinoma, gastric and esophageal cancer, pancreatic cancer, prostate cancer,
  • CLL chronic lymphocytic leuk
  • the autoimmune disease is type-1 diabetes.
  • the metabolic disease is selected from the group of: obesity, a lipodystrophy, and type-2 diabetes mellitus.
  • the neurodegenerative disease is selected from the group of: Alzheimer’s disease, Parkinson’s disease, and dementia.
  • the cardiovascular disease is selected from the group of: coronary artery disease, atherosclerosis, and pulmonary arterial hypertension.
  • the skin disease is selected from the group of: wound healing, alopecia, wrinkles, senile lentigo, skin thinning, xeroderma pigmentosum, and dyskeratosis congenita.
  • the progeria disease is selected from the group of: progeria and Hutchinson-Gilford Progeria Syndrome.
  • the fragility disease is selected from the group of: frailty, responsiveness to vaccination, osteoporosis, and sarcopenia.
  • the aging-related disease or condition is selected from the group of: age-related macular degeneration, osteoarthritis, adipose atrophy, idiopathic pulmonary fibrosis, kidney transplant failure, liver fibrosis, loss of bone mass, sarcopenia, age-associated loss of lung tissue elasticity, osteoporosis, age-associated renal dysfunction, and chemical-induced renal dysfunction.
  • Some embodiments of any of the methods described herein further include: obtaining a resting NK cell; and contacting the resting NK cell in vitro in a liquid culture medium including one or more NK cell activating agent(s), where the contacting results in the generation of the activated NK cells that are subsequently administered to the subject.
  • the resting NK cell is an autologous NK cell obtained from the subject. In some embodiments of any of the methods described herein, the resting NK cell is an allogeneic resting NK cell. In some embodiments of any of the methods described herein, the resting NK cell is an artificial NK cell. In some embodiments of any of the methods described herein, the resting NK cell is a haploidentical resting NK cell. In some embodiments of any of the methods described herein, the resting NK cell is a genetically-engineered NK cell carrying a chimeric antigen receptor or recombinant T cell receptor.
  • Some embodiments of any of the methods described herein further include isolating the activated NK cells before the activated NK cells are administered to the subject. Some embodiments of any of the methods described herein further include introducing a nucleic acid that encodes a chimeric antigen receptor or a recombinant T cell receptor into the resting NK cell or the activated NK cell prior to administration to the subject. Also provided herein are methods of improving the texture and/or appearance of skin and/or hair in a subject in need thereof over a period of time that include administering to the subject a therapeutically effective amount of one or more natural killer (NK) cell activating agent(s).
  • NK natural killer
  • Some embodiments of any of the methods described herein further include: obtaining a resting NK cell; and contacting the resting NK cell in vitro in a liquid culture medium including one or more NK cell activating agent(s), where the contacting results in the generation of the activated NK cells that are subsequently administered to the subject.
  • the resting NK cell is an autologous NK cell obtained from the subject.
  • the resting NK cell is an allogeneic resting NK cell. In some embodiments of any of the methods described herein, the resting NK cell is an artificial NK cell. In some embodiments of any of the methods described herein, the resting NK cell is a haploidentical resting NK cell. In some embodiments of any of the methods described herein, the resting NK cell is a genetically-engineered NK cell carrying a chimeric antigen receptor or recombinant T cell receptor. Some embodiments of any of the methods described herein further include isolating the activated NK cells before the activated NK cells are administered to the subject.
  • the method provides for an improvement in the texture and/or appearance of skin of the subject over the period of time. In some embodiments of any of the methods described herein, the method results in a decrease in the rate of formation of wrinkles in the skin of the subject over the period of time. In some embodiments of any of the methods described herein, the method results in an improvement in the coloration of skin of the subject over the period of time. In some embodiments of any of the methods described herein, the method results in a reduction of age spots on skin of the subject over the period of time. In some embodiments of any of the methods described herein, the method results in an improvement in the texture of skin of the subject over the period of time.
  • the method provides for an improvement in the texture and/or appearance of hair of the subject over the period of time. In some embodiments of any of the methods described herein, the method results in a decrease in the rate of formation of gray hair in the subject over the period of time. In some embodiments of any of the methods described herein, the method results in a decrease in the number of gray hairs of the subject over the period of time. In some embodiments of any of the methods described herein, the method results in a decrease in the rate of hair loss in the subject over time. In some embodiments of any of the methods described herein, the method results in an improvement in the texture of hair of the subject over the period of time.
  • the period of time is between about one month and about 10 years. In some embodiments of any of the methods described herein, the method results in a decrease in the number of senescent dermal fibroblasts in the skin of the subject over the period of time. Also provided herein are methods of assisting in the treatment of obesity in a subject in need thereof over a period of time that include administering to the subject a therapeutically effective amount of one or more natural killer (NK) cell activating agent(s). Also provided herein are methods of assisting in the treatment of obesity in a subject in need thereof over a period of time that include administering to the subject a therapeutically effective number of activated NK cells.
  • NK natural killer
  • any of the methods described herein further include: obtaining a resting NK cell; and contacting the resting NK cell in vitro in a liquid culture medium including one or more NK cell activating agent(s), where the contacting results in the generation of the activated NK cells that are subsequently administered to the subject.
  • the resting NK cell is an autologous NK cell obtained from the subject.
  • the resting NK cell is an allogeneic resting NK cell.
  • the resting NK cell is an artificial NK cell.
  • the resting NK cell is a haploidentical resting NK cell. In some embodiments of any of the methods described herein, the resting NK cell is a genetically-engineered NK cell carrying a chimeric antigen receptor or recombinant T cell receptor. Some embodiments of any of the methods described herein further include isolating the activated NK cells before the activated NK cells are administered to the subject. In some embodiments of any of the methods described herein, the method results in a decrease in the mass of the subject over the period of time. In some embodiments of any of the methods described herein, the method results in a decrease in the body mass index (BMI) of the subject over the period of time.
  • BMI body mass index
  • the method results in a decrease in the rate of progression from pre-diabetes to type-2 diabetes in the subject. In some embodiments of any of the methods described herein, the method results in a decrease in fasting serum glucose level in the subject. In some embodiments of any of the methods described herein, the method results in an increase in insulin sensitivity in the subject. In some embodiments of any of the methods described herein, the method results in a decrease in the severity of atherosclerosis in the subject. In some embodiments of any of the methods described herein, the period of time is between about two weeks and about 10 years.
  • At least one of the one or more NK cell activating agent(s) results in activation of one or more of: a receptor for IL-2, a receptor for IL-7, a receptor for IL-12, a receptor for IL-15, a receptor for IL- 18, a receptor for IL-21, a receptor for IL-33, CD16, CD69, CD25, CD59, CD352, NKp80, DNAM-1, 2B4, NKp30, NKp44, NKp46, NKG2D, KIR2DS1, KIR2Ds2/3, KIR2DL4, KIR2DS4, KIR2DS5, and KIR3DS1.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL-2 is a soluble IL-2 or an agonistic antibody that binds specifically to an IL-2 receptor. In some embodiments of any of the methods described herein, the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL-7 is a soluble IL-7 or an agonistic antibody that binds specifically to an IL-7 receptor.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL- 12 is a soluble IL-12 or an agonistic antibody that binds specifically to an IL-12 receptor.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL- 15 is a soluble IL-15 or an agonistic antibody that binds specifically to an IL-15 receptor.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL- 21 is a soluble IL-21 or an agonistic antibody that binds specifically to an IL-21 receptor.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL- 33 is a soluble IL-33 or an agonistic antibody that binds specifically to an IL-33 receptor.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for CD16 is an agonistic antibody that binds specifically to a CD16.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for CD69 is an agonistic antibody that binds specifically to a CD69.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for CD25 or CD59 is an agonistic antibody that binds specifically to CD25 or CD59.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for CD352 is an agonistic antibody that binds specifically to a CD352.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for NKp80 is an agonistic antibody that binds specifically to an NKp80.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for DNAM-1 is an agonistic antibody that binds specifically to a DNAM-1.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for 2B4 is an agonistic antibody that binds specifically to a 2B4.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for NKp30 is an agonistic antibody that binds specifically to an NKp30.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for NKp44 is an agonistic antibody that binds specifically to an NKp44.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for NKp46 is an agonistic antibody that binds specifically to an NKp46. In some embodiments of any of the methods described herein, the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for NKG2D is an agonistic antibody that binds specifically to an NKG2D. In some embodiments of any of the methods described herein, the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for KIR2DS1 is an agonistic antibody that binds specifically to a KIR2DS1.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for KIR2DS2/3 is an agonistic antibody that binds specifically to a KIR2DS2/3.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for KIR2DL4 is an agonistic antibody that binds specifically to a KIR2DL4.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for KIR2DS4 is an agonistic antibody that binds specifically to a KIR2DS4. In some embodiments of any of the methods described herein, the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for KIR2DS5 is an agonistic antibody that binds specifically to a KIR2DS5.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for KIR3DS1 is an agonistic antibody that binds specifically to a KIR3DS1.
  • At least one of the one or more NK cell activating agent(s) results in a decrease in the activation of one or more of: PD-1, a TGF- ⁇ receptor, TIGIT, CD1, TIM-3, Siglec-7, IRP60, Tactile, IL1R8, NKG2A/KLRD1, KIR2DL1, KIR2DL2/3, KIR2DL5, KIR3DL1, KIR3DL2, ILT2/LIR-1, and LAG-2.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of PD-1 is an antagonistic antibody that binds specifically to PD-1, a soluble PD-1, a soluble PD-L1, or an antibody that binds specifically to PD-L1.
  • at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of a TGF- ⁇ receptor is a soluble TGF- ⁇ receptor, an antibody that binds specifically to TGF- ⁇ , or an antagonistic antibody that binds specifically to a TGF- ⁇ receptor.
  • At least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of TIGIT is an antagonistic antibody that binds specifically to TIGIT, a soluble TIGIT, or an antibody that binds specifically to a ligand of TIGIT.
  • at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of CD1 is an antagonistic antibody that binds specifically to CD1, a soluble CD1, or an antibody that binds specifically to a ligand of CD1.
  • At least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of TIM-3 is an antagonistic antibody that binds specifically to TIM-3, a soluble TIM-3, or an antibody that binds specifically to a ligand of TIM-3.
  • at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of Siglec-7 is an antagonistic antibody that binds specifically to Siglec-7 or an antibody that binds specifically to a ligand of Siglec-7.
  • At least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of IRP60 is an antagonistic antibody that binds specifically to IRP60 or an antibody that binds specifically to a ligand of IRP60. In some embodiments of any of the methods described herein, at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of Tactile is an antagonistic antibody that binds specifically to Tactile or an antibody that binds specifically to a ligand of Tactile.
  • At least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of IL1R8 is an antagonistic antibody that binds specifically to IL1R8 or an antibody that binds specifically to a ligand of IL1R8.
  • at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of NKG2A/KLRD1 is an antagonistic antibody that binds specifically to NKG2A/KLRD1 or an antibody that binds specifically to a ligand of NKG2A/KLRD1.
  • At least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of KIR2DL1 is an antagonistic antibody that binds specifically to KIR2DL1 or an antibody that binds specifically to a ligand of KIR2DL1.
  • at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of KIR2DL2/3 is an antagonistic antibody that binds specifically to KIR2DL2/3 or an antibody that binds specifically to a ligand of KIR2DL2/3.
  • At least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of KIR2DL5 is an antagonistic antibody that binds specifically to KIR2DL5 or an antibody that binds specifically to a ligand of KIR2DL5.
  • at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of KIR3DL1 is an antagonistic antibody that binds specifically to KIR3DL1 or an antibody that binds specifically to a ligand of KIR3DL1.
  • At least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of KIR3DL2 is an antagonistic antibody that binds specifically to KIR3DL2 or an antibody that binds specifically to a ligand of KIR3DL2.
  • at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of ILT2/LIR-1 is an antagonistic antibody that binds specifically to ILT2/LIR-1 or an antibody that binds specifically to a ligand of ILT2/LIR-1.
  • At least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of LAG-2 is an antagonistic antibody that binds specifically to LAG-2 or an antibody that binds specifically to a ligand of LAG-2.
  • at least one of the one or more NK cell activating agent(s) is a single-chain chimeric polypeptide that includes: (i) a first target-binding domain; (ii) a soluble tissue factor domain; and (iii) a second target-binding domain.
  • the first target-binding domain and the soluble tissue factor domain directly abut each other.
  • the single- chain chimeric polypeptide further includes a linker sequence between the first target- binding domain and the soluble tissue factor domain.
  • the soluble tissue factor domain and the second target-binding domain directly abut each other.
  • the single-chain chimeric polypeptide further includes a linker sequence between the soluble tissue factor domain and the second target-binding domain.
  • the first target-binding domain and the second target-binding domain directly abut each other.
  • the single-chain chimeric polypeptide further includes a linker sequence between the first target-binding domain and the second target-binding domain.
  • the second target- binding domain and the soluble tissue factor domain directly abut each other.
  • the single-chain chimeric polypeptide further includes a linker sequence between the second target-binding domain and the soluble tissue factor domain.
  • the first target- binding domain and the second target-binding domain bind specifically to the same antigen. In some embodiments of any of the methods described herein, the first target- binding domain and the second target-binding domain bind specifically to the same epitope. In some embodiments of any of the methods described herein, the first target- binding domain and the second target-binding domain include the same amino acid sequence. In some embodiments of any of the methods described herein, the first target- binding domain and the second target-binding domain bind specifically to different antigens. In some embodiments of any of the methods described herein, one or both of the first target-binding domain and the second target-binding domain is an antigen-binding domain.
  • the first target- binding domain and the second target-binding domain are each an antigen-binding domain.
  • the antigen- binding domain includes a scFv or a single domain antibody.
  • one or both of the first target-binding domain and the second target-binding domain bind to a target selected from the group of: CD16a, CD33, CD20, CD19, CD22, CD123, PDL-1, TIGIT, PD-1, TIM3, CTLA4, MICA, MICB, IL-6, IL-8, TNF ⁇ , CD26, CD36, ULBP2, CD30, CD200, IGF-1R, MUC4AC, MUC5AC, Trop-2, CMET, EGFR, HER1, HER2, HER3, PSMA, CEA, B7H3, EPCAM, BCMA, P-cadherin, CEACAM5, a UL16-binding protein, HLA- DR, DLL4, TYRO3, AXL, MER, CD122, CD155, PDGF-DD, a ligand of TGF- ⁇ receptor II (TGF- ⁇ RII), a ligand of TGF- ⁇ RIII, a
  • a target selected from the group of: CD16
  • one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin or cytokine protein.
  • the soluble interleukin or cytokine protein is selected from the group of: IL-1, IL-2, IL-3, IL- 7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin or cytokine receptor.
  • the soluble interleukin or cytokine receptor is a soluble TGF- ⁇ receptor II (TGF- ⁇ RII) a soluble TGF- ⁇ RIII, a soluble receptor for TNF ⁇ , a soluble receptor for IL-4, or a soluble receptor for IL-10.
  • TGF- ⁇ RII soluble TGF- ⁇ receptor II
  • soluble TGF- ⁇ RIII soluble TGF- ⁇ RIII
  • soluble receptor for TNF ⁇ a soluble receptor for TNF ⁇
  • a soluble receptor for IL-4 or a soluble receptor for IL-10.
  • the soluble tissue factor domain is a soluble human tissue factor domain.
  • the soluble human tissue factor domain includes a sequence that is at least 80% identical to SEQ ID NO: 93.
  • the soluble human tissue factor domain includes a sequence that is at least 90% identical to SEQ ID NO: 93. In some embodiments of any of the methods described herein, the soluble human tissue factor domain includes a sequence that is at least 95% identical to SEQ ID NO: 93.
  • the soluble human tissue factor domain does not include one or more of: a lysine at an amino acid position that corresponds to amino acid position 20 of mature wildtype human tissue factor protein; an isoleucine at an amino acid position that corresponds to amino acid position 22 of mature wildtype human tissue factor protein; a tryptophan at an amino acid position that corresponds to amino acid position 45 of mature wildtype human tissue factor protein; an aspartic acid at an amino acid position that corresponds to amino acid position 58 of mature wildtype human tissue factor protein; a tyrosine at an amino acid position that corresponds to amino acid position 94 of mature wildtype human tissue factor protein; an arginine at an amino acid position that corresponds to amino acid position 135 of mature wildtype human tissue factor protein; and a phenylalanine at an amino acid position that corresponds to amino acid position 140 of mature wildtype human tissue factor protein.
  • the soluble human tissue factor domain does not include any of: a lysine at an amino acid position that corresponds to amino acid position 20 of mature wildtype human tissue factor protein; an isoleucine at an amino acid position that corresponds to amino acid position 22 of mature wildtype human tissue factor protein; a tryptophan at an amino acid position that corresponds to amino acid position 45 of mature wildtype human tissue factor protein; an aspartic acid at an amino acid position that corresponds to amino acid position 58 of mature wildtype human tissue factor protein; a tyrosine at an amino acid position that corresponds to amino acid position 94 of mature wildtype human tissue factor protein; an arginine at an amino acid position that corresponds to amino acid position 135 of mature wildtype human tissue factor protein; and a phenylalanine at an amino acid position that corresponds to amino acid position 140 of mature wildtype human tissue factor protein.
  • the soluble tissue factor domain is not capable of binding Factor VIIa. In some embodiments of any of the methods described herein, the soluble tissue factor domain does not convert inactive Factor X into Factor Xa. In some embodiments of any of the methods described herein, the single-chain chimeric polypeptide does not blood stimulate coagulation in a mammal. In some embodiments of any of the methods described herein, the single-chain chimeric polypeptide further includes one or more additional target-binding domains at its N- and/or C-terminus. In some embodiments of any of the methods described herein, the single-chain chimeric polypeptide includes one or more additional target-binding domains at its N- terminus.
  • one or more additional target-binding domains directly abuts the first target-binding domain, the second target-binding domain, or the soluble tissue factor domain.
  • the single-chain chimeric polypeptide further includes a linker sequence between one of the at least one additional target-binding domains and the first target-binding domain, the second target-binding domain, or the soluble tissue factor domain.
  • the single-chain chimeric polypeptide includes one or more additional target-binding domains at its C- terminus.
  • one of the one or more additional target-binding domains directly abuts the first target-binding domain, the second target-binding domain, or the soluble tissue factor domain.
  • the single-chain chimeric polypeptide further includes a linker sequence between one of the at least one additional target-binding domains and the first target-binding domain, the second target-binding domain, or the soluble tissue factor domain.
  • the single-chain chimeric polypeptide includes one or more additional target binding domains at its N- terminus and the C-terminus.
  • one of the one or more additional antigen binding domains at the N-terminus directly abuts the first target-binding domain, the second target-binding domain, or the soluble tissue factor domain.
  • the single-chain chimeric polypeptide further includes a linker sequence between one of the one or more additional antigen-binding domains at the N-terminus and the first target-binding domain, the second target-binding domain, or the soluble tissue factor domain.
  • one of the one or more additional antigen binding domains at the C-terminus directly abuts the first target- binding domain, the second target-binding domain, or the soluble tissue factor domain.
  • the single-chain chimeric polypeptide further includes a linker sequence between one of the one or more additional antigen-binding domains at the C-terminus and the first target-binding domain, the second target-binding domain, or the soluble tissue factor domain.
  • two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to the same antigen.
  • two or more of the first target- binding domain, the second target-binding domain, and the one or more additional target- binding domains bind specifically to the same epitope. In some embodiments of any of the methods described herein, two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains include the same amino acid sequence. In some embodiments of any of the methods described herein, the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains each bind specifically to the same antigen.
  • the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains each bind specifically to the same epitope. In some embodiments of any of the methods described herein, the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains each include the same amino acid sequence. In some embodiments of any of the methods described herein, the first target- binding domain, the second target-binding domain, and the one or more additional target- binding domains bind specifically to different antigens.
  • one or more of the first target-binding domain, the second target-binding domain, and the one or more target-binding domains is an antigen-binding domain.
  • the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains are each an antigen- binding domain.
  • the antigen-binding domain includes a scFv or a single domain antibody.
  • one or more of the first target-binding domain, the second target-binding domain, and the one or more target-binding domains bind specifically to a target selected from the group of: CD16a, CD33, CD20, CD19, CD22, CD123, PDL-1, TIGIT, PD-1, TIM3, CTLA4, MICA, MICB, IL-6, IL-8, TNF ⁇ , CD26, CD36, ULBP2, CD30, CD200, IGF-1R, MUC4AC, MUC5AC, Trop-2, CMET, EGFR, HER1, HER2, HER3, PSMA, CEA, B7H3, EPCAM, BCMA, P-cadherin, CEACAM5, a UL16-binding protein, HLA-DR, DLL4, TYRO3, AXL, MER, CD122, CD155, PDGF-DD, a ligand of TGF- ⁇ receptor II (TGF- ⁇ RII), a lig
  • one or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains is a soluble interleukin or cytokine protein.
  • the soluble interleukin or cytokine protein is selected from the group of: IL-1, IL-2, IL-3, IL-7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains is a soluble interleukin or cytokine receptor.
  • the soluble receptor is a soluble TGF- ⁇ receptor II (TGF- ⁇ RII) a soluble TGF- ⁇ RIII, a soluble receptor for TNF ⁇ , a soluble receptor for IL-4, or a soluble receptor for IL-10.
  • At least one of the one or more NK cell activating agent(s) is a multi-chain chimeric polypeptide that includes: (a) a first chimeric polypeptide including: (i) a first target-binding domain; (ii) a soluble tissue factor domain; and (iii) a first domain of a pair of affinity domains; and (b) a second chimeric polypeptide including: (i) a second domain of a pair of affinity domains; and (ii) a second target-binding domain, where the first chimeric polypeptide and the second chimeric polypeptide associate through the binding of the first domain and the second domain of the pair of affinity domains.
  • the first target- binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence between the first target- binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the first target- binding domain and the second target-binding domain bind specifically to the same antigen. In some embodiments of any of the methods described herein, the first target- binding domain and the second target-binding domain bind specifically to the same epitope. In some embodiments of any of the methods described herein, the first target- binding domain and the second target-binding domain include the same amino acid sequence. In some embodiments of any of the methods described herein, the first target- binding domain and the second target-binding domain bind specifically to different antigens. In some embodiments of any of the methods described herein, one or both of the first target-binding domain and the second target-binding domain is an antigen-binding domain.
  • the first target- binding domain and the second target-binding domain are each antigen-binding domains.
  • the antigen-binding domain includes a scFv or a single domain antibody.
  • one or both of the first target-binding domain and the second target-binding domain bind specifically to a target selected from the group of: CD16a, CD33, CD20, CD19, CD22, CD123, PDL-1, TIGIT, PD-1, TIM3, CTLA4, MICA, MICB, IL-6, IL-8, TNF ⁇ , CD26, CD36, ULBP2, CD30, CD200, IGF-1R, MUC4AC, MUC5AC, Trop-2, CMET, EGFR, HER1, HER2, HER3, PSMA, CEA, B7H3, EPCAM, BCMA, P-cadherin, CEACAM5, a UL16-binding protein, HLA-DR, DLL4, TYRO3, AXL, MER, CD122, CD155, PDGF-DD, a ligand of TGF- ⁇ receptor II (TGF- ⁇ RII), a ligand of TGF- ⁇ RIII, a
  • a target selected from the group of: CD16
  • one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin or cytokine protein.
  • the soluble interleukin or cytokine protein is selected from the group of: IL-1, IL-2, IL-3, IL- 7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin or cytokine receptor.
  • the soluble receptor is a soluble TGF- ⁇ receptor II (TGF- ⁇ RII) a soluble TGF- ⁇ RIII, a soluble receptor for TNF ⁇ , a soluble receptor for IL-4, or a soluble receptor for IL-10.
  • the first chimeric polypeptide further includes one or more additional target-binding domain(s), where at least one of the one or more additional antigen-binding domain(s) is positioned between the soluble tissue factor domain and the first domain of the pair of affinity domains.
  • the first chimeric polypeptide further includes a linker sequence between the soluble tissue factor domain and the at least one of the one or more additional antigen-binding domain(s), and/or a linker sequence between the at least one of the one or more additional antigen-binding domain(s) and the first domain of the pair of affinity domains.
  • the first chimeric polypeptide further includes one or more additional target-binding domains at the N- terminal and/or C-terminal end of the first chimeric polypeptide.
  • At least one of the one or more additional target- binding domains directly abuts the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence between the at least one of the one or more additional target-binding domains and the first domain of the pair of affinity domains.
  • the at least one of the one or more additional target-binding domains directly abuts the first target-binding domain in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence between the at least one of the one or more additional target-binding domains and the first target-binding domain.
  • at least one of the one or more additional target-binding domains is disposed at the N- and/or C-terminus of the first chimeric polypeptide, and at least one of the one or more additional target- binding domains is positioned between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the at least one additional target- binding domain of the one or more additional target-binding domains disposed at the N- terminus directly abuts the first target-binding domain or the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence disposed between the at least one additional target-binding domain and the first target-binding domain or the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the at least one additional target-binding domain of the one or more additional target-binding domains disposed at the C-terminus directly abuts the first target-binding domain or the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence disposed between the at least one additional target- binding domain and the first target-binding domain or the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the at least one of the one or more additional target-binding domains positioned between the soluble tissue factor domain and the first domain of the pair of affinity domains directly abuts the soluble tissue factor domain and/or the first domain of the pair of affinity domains.
  • the first chimeric polypeptide further includes a linker sequence disposed (i) between the soluble tissue factor domain and the at least one of the one or more additional target-binding domains positioned between the soluble tissue factor domain and the first domain of the pair of affinity domains, and/or (ii) between the first domain of the pair of affinity domains and the at least one of the one or more additional target-binding domains positioned between the soluble tissue factor domain and the first domain of the pair of affinity domains.
  • the second chimeric polypeptide further includes one or more additional target-binding domains at the N-terminal end or the C-terminal end of the second chimeric polypeptide.
  • At least one of the one or more additional target-binding domains directly abuts the second domain of the pair of affinity domains in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence between at least one of the one or more additional target-binding domains and the second domain of the pair of affinity domains in the second chimeric polypeptide.
  • at least one of the one or more additional target-binding domains directly abuts the second target-binding domain in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence between at least one of the one or more additional target-binding domains and the second target-binding domain in the second chimeric polypeptide.
  • two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to the same antigen.
  • two or more of the first target- binding domain, the second target-binding domain, and the one or more additional target- binding domains bind specifically to the same epitope.
  • two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains include the same amino acid sequence. In some embodiments of any of the methods described herein, the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains each bind specifically to the same antigen. In some embodiments of any of the methods described herein, the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains each bind specifically to the same epitope.
  • the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains each include the same amino acid sequence. In some embodiments of any of the methods described herein, the first target- binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to different antigens. In some embodiments of any of the methods described herein, one or more of the first target-binding domain, the second target-binding domain, and the one or more target-binding domains is an antigen-binding domain.
  • the first target- binding domain, the second target-binding domain, and the one or more additional target- binding domains are each an antigen-binding domain.
  • the antigen-binding domain includes a scFv.
  • one or more of the first target-binding domain, the second target-binding domain, and the one or more target-binding domains bind specifically to a target selected from the group of: CD16a, CD33, CD20, CD19, CD22, CD123, PDL-1, TIGIT, PD-1, TIM3, CTLA4, MICA, MICB, IL-6, IL-8, TNF ⁇ , CD26, CD36, ULBP2, CD30, CD200, IGF-1R, MUC4AC, MUC5AC, Trop-2, CMET, EGFR, HER1, HER2, HER3, PSMA, CEA, B7H3, EPCAM, BCMA, P-cadherin, CEACAM5, a UL16-binding protein, HLA-DR, DLL4, TYRO3, AXL, MER, CD122, CD155, PDGF-DD, a ligand of TGF- ⁇ receptor II (TGF- ⁇ RII), a lig
  • one or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains is a soluble interleukin or cytokine protein.
  • the soluble interleukin or cytokine protein is selected from the group of: IL-1, IL-2, IL-3, IL-7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains is a soluble interleukin or cytokine receptor.
  • the soluble receptor a soluble TGF- ⁇ receptor II (TGF- ⁇ RII) a soluble TGF- ⁇ RIII, a soluble receptor for TNF ⁇ , a soluble receptor for IL-4, or a soluble receptor for IL-10.
  • the soluble tissue factor domain is a soluble human tissue factor domain.
  • the soluble human tissue factor domain includes a sequence that is at least 80% identical to SEQ ID NO: 93. In some embodiments of any of the methods described herein, the soluble human tissue factor domain includes a sequence that is at least 90% identical to SEQ ID NO: 93. In some embodiments of any of the methods described herein, the soluble human tissue factor domain includes a sequence that is at least 95% identical to SEQ ID NO: 93.
  • the soluble human tissue factor domain does not include one or more of: a lysine at an amino acid position that corresponds to amino acid position 20 of mature wildtype human tissue factor protein; an isoleucine at an amino acid position that corresponds to amino acid position 22 of mature wildtype human tissue factor protein; a tryptophan at an amino acid position that corresponds to amino acid position 45 of mature wildtype human tissue factor protein; an aspartic acid at an amino acid position that corresponds to amino acid position 58 of mature wildtype human tissue factor protein; a tyrosine at an amino acid position that corresponds to amino acid position 94 of mature wildtype human tissue factor protein; an arginine at an amino acid position that corresponds to amino acid position 135 of mature wildtype human tissue factor protein; and a phenylalanine at an amino acid position that corresponds to amino acid position 140 of mature wildtype human tissue factor protein.
  • the soluble human tissue factor domain does not include any of: a lysine at an amino acid position that corresponds to amino acid position 20 of mature wildtype human tissue factor protein; an isoleucine at an amino acid position that corresponds to amino acid position 22 of mature wildtype human tissue factor protein; a tryptophan at an amino acid position that corresponds to amino acid position 45 of mature wildtype human tissue factor protein; an aspartic acid at an amino acid position that corresponds to amino acid position 58 of mature wildtype human tissue factor protein; a tyrosine at an amino acid position that corresponds to amino acid position 94 of mature wildtype human tissue factor protein; an arginine at an amino acid position that corresponds to amino acid position 135 of mature wildtype human tissue factor protein; and a phenylalanine at an amino acid position that corresponds to amino acid position 140 of mature wildtype human tissue factor protein.
  • the soluble tissue factor domain is not capable of binding to Factor VIIa. In some embodiments of any of the methods described herein, the soluble tissue factor domain does not convert inactive Factor X into Factor Xa. In some embodiments of any of the methods described herein, the multi-chain chimeric polypeptide does not stimulate blood coagulation in a mammal. In some embodiments of any of the methods described herein, the pair of affinity domains is a sushi domain from an alpha chain of human IL-15 receptor (IL-15R ⁇ ) and a soluble IL-15. In some embodiments of any of the methods described herein, the soluble IL-15 has a D8N or D8A amino acid substitution.
  • the human IL-15R ⁇ is a mature full-length IL-15R ⁇ .
  • the pair of affinity domains is selected from the group of: barnase and barnstar, a PKA and an AKAP, adapter/docking tag modules based on mutated RNase I fragments, and SNARE modules based on interactions of the proteins syntaxin, synaptotagmin, synaptobrevin, and SNAP25.
  • At least one of the one or more NK cell activating agent(s) is a multi-chain chimeric polypeptide that includes: (a) a first and second chimeric polypeptides, where each includes: (i) a first target-binding domain; (ii) a Fc domain; and (iii) a first domain of a pair of affinity domains; and (b) a third and fourth chimeric polypeptide, where each includes: (i) a second domain of a pair of affinity domains; and (ii) a second target-binding domain, where the first and second chimeric polypeptides and the third and fourth chimeric polypeptides associate through the binding of the first domain and the second domain of the pair of affinity domains, and the first and second chimeric polypeptides associate through their Fc domains.
  • the first target- binding domain and the Fc domain directly abut each other in the first and second chimeric polypeptides.
  • the first and second chimeric polypeptides further include a linker sequence between the first target-binding domain and the Fc domain in the first and second chimeric polypeptides.
  • the Fc domain and the first domain of the pair of affinity domains directly abut each other in the first and second chimeric polypeptides.
  • the first chimeric polypeptide further includes a linker sequence between the Fc domain and the first domain of the pair of affinity domains in the first and second chimeric polypeptides.
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the third and fourth chimeric polypeptides.
  • the third and fourth chimeric polypeptides further include a linker sequence between the second domain of the pair of affinity domains and the second target-binding domain in the third and fourth chimeric polypeptides.
  • the first target- binding domain and the second target-binding domain bind specifically to the same antigen. In some embodiments of any of the methods described herein, the first target- binding domain and the second target-binding domain bind specifically to the same epitope. In some embodiments of any of the methods described herein, the first target- binding domain and the second target-binding domain include the same amino acid sequence. In some embodiments of any of the methods described herein, the first target- binding domain and the second target-binding domain bind specifically to different antigens. In some embodiments of any of the methods described herein, one or both of the first target-binding domain and the second target-binding domain is an antigen- binding domain.
  • the first target-binding domain and the second target-binding domain are each antigen-binding domains.
  • the antigen- binding domain includes a scFv or a single domain antibody.
  • one or both of the first target-binding domain and the second target-binding domain bind specifically to a target selected from the group of: CD16a, CD33, CD20, CD19, CD22, CD123, PDL-1, TIGIT, PD-1, TIM3, CTLA4, MICA, MICB, IL-6, IL-8, TNF ⁇ , CD26, CD36, ULBP2, CD30, CD200, IGF-1R, MUC4AC, MUC5AC, Trop-2, CMET, EGFR, HER1, HER2, HER3, PSMA, CEA, B7H3, EPCAM, BCMA, P-cadherin, CEACAM5, a UL16-binding protein, HLA-DR, DLL4, TYRO3, AXL, MER, CD122, CD155, PDGF-DD, a ligand of TGF- ⁇ receptor II (TGF- ⁇ RII), a ligand of TGF- ⁇ RIII, a
  • a target selected from the group of: CD16
  • one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin or cytokine protein.
  • the soluble interleukin or cytokine protein is selected from the group of: IL-1, IL-2, IL-3, IL- 7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin or cytokine receptor.
  • the soluble receptor is a soluble TGF- ⁇ receptor II (TGF- ⁇ RII) a soluble TGF- ⁇ RIII, a soluble receptor for TNF ⁇ , a soluble receptor for IL-4, or a soluble receptor for IL-10.
  • TGF- ⁇ RII soluble TGF- ⁇ receptor II
  • soluble TGF- ⁇ RIII soluble TGF- ⁇ RIII
  • soluble receptor for TNF ⁇ a soluble receptor for IL-4
  • a soluble receptor for IL-10 soluble tissue factor domain
  • the soluble tissue factor domain is a soluble human tissue factor domain that does not stimulate blood coagulation.
  • the soluble tissue factor domain comprises or consists of a sequence from a wildtype soluble human tissue factor.
  • methods of killing or reducing the number of naturally- occurring and/or treatment-induced senescent cells in a subject that include administering to the subject a therapeutically effective amount of one or more common gamma-chain family cytokine receptor activating agent(s).
  • methods of decreasing the accumulation of naturally- occurring and/or treatment-induced senescent cells in a subject that include administering to the subject a therapeutically effective amount of one or more common gamma-chain family cytokine receptor activating agent(s).
  • the subject has been previously diagnosed or identified as having an aging-related disease or an inflammatory disease.
  • the aging-related disease is inflamm-aging related.
  • the aging-related disease is selected from the group of: Alzheimer’s disease, aneurysm, cystic fibrosis, fibrosis in pancreatitis, glaucoma, hypertension, idiopathic pulmonary fibrosis, inflammatory bowel disease, intervertebral disc degeneration, osteoarthritis, type 2 diabetes mellitus, adipose atrophy, lipodystrophy, atherosclerosis, cataracts, COPD, kidney transplant failure, liver fibrosis, loss of bone mass, myocardial infarction, sarcopenia, wound healing, alopecia, cardiomyocyte hypertrophy, osteoarthritis, Parkinson’s disease, age-associated loss of lung tissue elasticity, age-related macular degeneration, cachexia, glomerulosclerosis, liver cirrhosis, NAFLD, osteoporosis, amyotrophic lateral sclerosis, Huntington’s disease, spinocerebellar ataxia, multiple sclerosis, neurodeficide
  • the aging-related disease is a cancer selected from the group of: solid tumor, hematological tumor, sarcoma, osteosarcoma, glioblastoma, neuroblastoma, melanoma, rhabdomyosarcoma, Ewing sarcoma, osteosarcoma, B-cell neoplasms, multiple myeloma, B-cell lymphoma, B-cell non-Hodgkin’s lymphoma, Hodgkin’s lymphoma, chronic lymphocytic leukemia (CLL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), myelodysplastic syndromes (MDS), cutaneous T-cell lymphoma, retinoblastoma, stomach cancer, urothelial carcinoma, lung cancer, renal cell carcinoma, gastric and esophageal cancer, pancreatic cancer, prostate cancer, breast
  • the inflammatory disease is selected from the group of: rheumatoid arthritis, inflammatory bowel disease, lupus erythematosus, lupus nephritis, diabetic nephropathy, CNS injury, Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, Crohn’s disease, multiple sclerosis, Guillain-Barre syndrome, psoriasis, Grave’s disease, ulcerative colitis, nonalcoholic steatohepatitis, mood disorders and cancer treatment-related cognitive impairment.
  • the treatment-induced senescent cells are chemotherapy- induced senescent cells.
  • the administration of the one or more common gamma-chain family cytokine receptor activating agent(s) results in a decrease in the number of naturally-occurring senescent cells and/or treatment-induced senescent cells in a target tissue in the subject.
  • the target tissue is selected from the group of: adipose tissue, pancreatic tissue, liver tissue, kidney tissue, lung tissue, vasculature, bone tissue, central nervous system (CNS) tissue, eye tissue, skin tissue, muscle tissue, and secondary lympho-organ tissue.
  • At least one of the one or more common gamma-chain family cytokine receptor activating agent(s) is a complex of a common gamma-chain family cytokine or a functional fragment thereof and an antibody or antibody fragment that binds specifically to the common gamma-chain family cytokine or the functional fragment thereof.
  • At least one of the one or more common gamma-chain family cytokine receptor activating agent(s) is a single-chain chimeric polypeptide comprising: (i) a first target-binding domain; (ii) a soluble tissue factor domain; and (iii) a second target-binding domain, wherein one or both of the first target-binding domain and the second target-binding domain is a soluble common gamma-chain family cytokine, an agonistic antigen-binding domain that binds specifically to a common gamma-chain family cytokine receptor, a soluble common gamma-chain family cytokine receptor, or an antigen-binding domain that binds specifically to a common gamma-chain family cytokine.
  • one or both of the first target-binding domain and the second target-binding domain comprises a soluble common gamma-chain family cytokine.
  • the soluble common gamma-chain family cytokine is selected from the group consisting of: soluble IL-2, soluble IL-4, soluble IL-7, soluble IL-9, soluble IL-15, and soluble IL-21.
  • one or both of the first target-binding domain and the second target-binding domain comprises an agonistic antigen-binding domain that binds specifically to a common gamma-chain family cytokine receptor.
  • the common gamma-chain family cytokine receptor is a receptor for one or more of IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21.
  • the agonistic antigen-binding domain is an scFv, a VHH, or a VNAR.
  • the first target-binding domain and the soluble tissue factor domain directly abut each other.
  • the single-chain chimeric polypeptide further comprises a linker sequence between the first target-binding domain and the soluble tissue factor domain.
  • the soluble tissue factor domain and the second target-binding domain directly abut each other.
  • the single-chain chimeric polypeptide further comprises a linker sequence between the soluble tissue factor domain and the second target-binding domain.
  • the first target-binding domain and the second target-binding domain bind specifically to the same antigen.
  • the first target-binding domain and the second target-binding domain bind specifically to the same epitope.
  • the first target-binding domain and the second target-binding domain comprise the same amino acid sequence.
  • the first target-binding domain and the second target-binding domain bind specifically to different antigens.
  • the soluble tissue factor domain is a soluble human tissue factor domain.
  • the soluble human tissue factor domain comprises a sequence that is at least 80% identical to SEQ ID NO: 93.
  • the single-chain chimeric polypeptide further comprises one or more additional target- binding domains at its N- and/or C-terminus.
  • At least one of the one or more common gamma-chain family cytokine receptor activating agent(s) is a multi-chain chimeric polypeptide comprising: (a) a first chimeric polypeptide comprising: (i) a first target-binding domain; (ii) a soluble tissue factor domain; and (iii) a first domain of a pair of affinity domains; (b) a second chimeric polypeptide comprising: (i) a second domain of a pair of affinity domains; and (ii) a second target-binding domain, wherein one or both of the first target- binding domain and the second target-binding domain is a soluble common gamma-chain family cytokine, an agonistic antigen-binding domain that binds specifically to a common gamma-chain family cytokine receptor, a soluble common gamma-chain family cytokine receptor, or an antigen-binding domain that binds specifically to a common
  • one or both of the first target-binding domain and the second target-binding domain comprises a soluble common gamma-chain family cytokine.
  • the soluble common gamma-chain family cytokine is selected from the group of: soluble IL-2, soluble IL-4, soluble IL-7, soluble IL-9, soluble IL-15, and soluble IL-21.
  • one or both of the first target-binding domain and the second target-binding domain comprises an agonistic antigen-binding domain that binds specifically to a common gamma-chain family cytokine receptor.
  • the common gamma-chain family cytokine receptor is a receptor for one or more of IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21.
  • the agonistic antigen-binding domain is an scFv, a VHH, or a VNAR.
  • the first target-binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further comprises a linker sequence between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the first target-binding domain and the second target- binding domain bind specifically to the same antigen. In some embodiments, the first target-binding domain and the second target-binding domain bind specifically to different antigens. In some embodiments, the first chimeric polypeptide further comprises one or more additional target-binding domain(s). In some embodiments, the second chimeric polypeptide further comprises one or more additional target-binding domains. In some embodiments, the soluble tissue factor domain is a soluble human tissue factor domain. In some embodiments, the soluble human tissue factor domain comprises a sequence that is at least 80% identical to SEQ ID NO: 93.
  • the pair of affinity domains is a sushi domain from an alpha chain of human IL-15 receptor (IL15R ⁇ ) and a soluble IL-15.
  • the pair of affinity domains is selected from the group of: barnase and barnstar, a PKA and an AKAP, adapter/docking tag modules based on mutated RNase I fragments, and SNARE modules based on interactions of the proteins syntaxin, synaptotagmin, synaptobrevin, and SNAP25.
  • the first domain or the second domain of a pair of affinity domains is a soluble common gamma-chain family cytokine or an antigen-binding domain that binds specifically to a common gamma-chain family cytokine receptor.
  • at least one of the one or more common gamma-chain family cytokine receptor activating agent(s) is soluble IL-15 or an IL-15 agonist.
  • the soluble IL-15 is at least 90% identical to SEQ ID NO: 82.
  • the IL-15 agonist comprises a complex of IL-15 and all or a portion of a soluble IL-15 receptor (IL-15R).
  • the portion of the soluble IL- 15R is a portion of IL-15R ⁇ . In some embodiments, the portion of the soluble IL-15R ⁇ is a sushi domain of IL-15R ⁇ . In some embodiments, the IL-15 agonist further comprises an Fc domain. In some embodiments, the IL-15 agonist comprises a fusion protein comprising IL-15 and a sushi domain from an IL-15R ⁇ . In some embodiments, one of the one or more common gamma-chain family cytokine receptor activating agent(s) is a soluble IL-2 or an IL-2 agonist.
  • one of the one or more common gamma-chain family cytokine receptor activating agent(s) is an antibody or an antigen- binding antibody fragment that binds specifically to a common gamma-chain family cytokine.
  • the method comprises administering one, two or more doses of the one or more common gamma-chain family cytokine receptor activating agent(s) to the subject. In some embodiments, any two consecutive doses of the two or more doses are administered about 1 week to about one year apart. In some embodiments, any two consecutive doses of the two or more doses are administered about 1 week to about 6 months apart. In some embodiments, any two consecutive doses of the two or more doses are administered about 1 week to about 2 months apart.
  • any two consecutive doses of the two or more doses are administered about 1 week to about 1 month apart.
  • the one, two or more doses are administered by subcutaneous administration.
  • the two or more doses are administered by intramuscular administration.
  • the two or more doses are administered over a period of time of about 1 year to about 60 years.
  • the two or more doses are administered over a period of time of about 1 year to about 50 years.
  • the two or more doses are administered over a period of time of about 1 year to about 40 years.
  • the two or more doses are administered over a period of time of about 1 year to about 30 years.
  • the two or more doses are administered over a period of time of about 1 year to about 20 years. In some embodiments, the two or more doses are administered over a period of time of about 1 year to about 10 years. In some embodiments, each of the two or more doses are administered at a dosage of about 0.01 mg of each common gamma-chain family cytokine receptor activating agent/kg to about 10 mg of each common gamma-chain family cytokine receptor activating agent/kg.
  • each of the two or more doses are administered at a dosage of about 0.02 mg of each common gamma-chain family cytokine receptor activating agent/kg to about 5 mg of each common gamma-chain family cytokine receptor activating agent/kg.
  • a first dose of the one or more common gamma-chain family cytokine receptor activating agent(s) begins when the subject reaches an age of at least 30 years. In some embodiments, a first dose of the one or more common gamma- chain family cytokine receptor activating agent(s) begins when the subject reaches an age of at least 40 years.
  • a first dose of the one or more common gamma-chain family cytokine receptor activating agent(s) begins when the subject reaches an age of at least 50 years. In some embodiments, a first dose of the one or more common gamma-chain family cytokine receptor activating agent(s) begins when the subject reaches an age of at least 60 years. In some embodiments, the subject is not diagnosed or identified as having an aging-related disease or an inflammatory disease. In some embodiments, the subject has not been previously treated with a chemotherapeutic agent. In some embodiments, the subject has not been previously treated with a therapeutic agent that induces cellular senescence.
  • the method further comprises administering to the subject at least one or more agent(s) that results in a decrease in the activation of a TGF- ⁇ receptor.
  • the agent that results in a decrease in the activation of a TGF- ⁇ receptor is a soluble TGF- ⁇ receptor, an extracellular domain of TGF- ⁇ receptor, an antibody that binds specifically to TGF- ⁇ , an antagonistic antibody that binds to a TGF- ⁇ receptor, an agent that binds to a LAP, or an agent that binds to a TGF- ⁇ /LAP complex.
  • the one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ decrease(s) the activation of a TGF- ⁇ receptor through binding to a LAP, or to a TGF- ⁇ /LAP complex.
  • the soluble human tissue factor domain does not initiate blood coagulation.
  • the method further comprises administering an additional therapeutic agent selected from the group of: combinations of agents, such as checkpoint inhibitors, chemotherapy drugs, and therapeutic antibodies.
  • the single-chain chimeric polypeptide is stable in human serum for at least 10 days at 37 °C.
  • the multi-chain chimeric polypeptide is stable in human serum for at least 10 days at 37 °C. In some embodiments of any of the methods described herein, the single-chain chimeric polypeptide does not have significant clotting activity. In some embodiments of any of the methods described herein, the multi-chain chimeric polypeptide does not have significant clotting activity. In some embodiments of any of the methods described herein, the method results in rejuvenation of aged immune cells in the subject. In some embodiments of any of the methods described herein, the rejuvenation of the aged immune cells results in a reduction of number of diseased cells or infectious agents in the subject.
  • the aged immune cells include one or more of aged NK cells, aged NKT cells, aged T cells, aged B cells, aged monocytes, aged macrophages, aged neutrophils, aged basophils, aged eosinophils, aged Kupffer cells, and aged microgial cells.
  • the diseased cells include cancer cells, virally-infected cells, and intracellularly- bacterially-infected cells.
  • the infectious agents include virus, bacterium, fungus, and parasite.
  • chimeric refers to a polypeptide that includes amino acid sequences (e.g., domains) originally derived from two different sources (e.g., two different naturally-occurring proteins, e.g., from the same or different species).
  • a chimeric polypeptide can include domains from at least two different naturally occurring human proteins.
  • a chimeric polypeptide can include a domain that is a synthetic sequence (e.g., a scFv) and a domain that is derived from a naturally-occurring protein (e.g., a naturally-occurring human protein).
  • a chimeric polypeptide can include at least two different domains that are synthetic sequences (e.g., two different scFvs).
  • An “activated NK cell” is a NK cell demonstrating increased expression levels of two or more (e.g., three, four, five, or six) of CD25, CD69, MTOR-C1, SREBP, IFN- ⁇ , and a granzyme (e.g., granzyme B), e.g., as compared to a resting NK cell. Exemplary methods for identifying the expression levels of CD25, CD69, MTOR-C1, SREBP, IFN- ⁇ , and a granzyme (e.g., granzyme B) are described herein.
  • a “resting NK cell” is a NK cell that has a reduced expression of two or more (e.g., three, four, five, or six) of CD25, CD69, MTOR-C1, SREBP, IFN- ⁇ , and a granzyme (e.g., granzyme B), e.g., as compared to an activated NK cell.
  • An “NK cell activating agent” is an agent that induces or promotes (alone or in combination with additional NK cell activating agents) a resting NK cell to develop into an activated NK cell. Non-limiting examples and aspects of NK cell activating agents are described herein.
  • an “antigen-binding domain” is one or more protein domain(s) (e.g., formed from amino acids from a single polypeptide or formed from amino acids from two or more polypeptides (e.g., the same or different polypeptides) that is capable of specifically binding to one or more different antigen(s).
  • an antigen-binding domain can bind to an antigen or epitope with specificity and affinity similar to that of naturally-occurring antibodies.
  • the antigen-binding domain can be an antibody or a fragment thereof.
  • an antigen-binding domain can include an alternative scaffold. Non-limiting examples of antigen-binding domains are described herein. Additional examples of antigen-binding domains are known in the art.
  • a “soluble tissue factor domain” refers to a polypeptide having at least 70% identity (e.g., at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity, at least 99% identity, or 100% identical) to a segment of a wildtype mammalian tissue factor protein (e.g., a wildtype human tissue factor protein) that lacks the transmembrane domain and the intracellular domain.
  • soluble tissue factor domains are described herein.
  • the term “soluble interleukin protein” is used herein to refer to a mature and secreted interleukin protein or a biologically active fragment thereof.
  • a soluble interleukin protein can include a sequence that is at least 70% identical, at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical to a wildtype mature and secreted mammalian interleukin protein (e.g., a wildtype human interleukin protein) and retains its biological activity.
  • a wildtype mature and secreted mammalian interleukin protein e.g., a wildtype human interleukin protein
  • soluble interleukin proteins are described herein.
  • the term “soluble cytokine protein” is used herein to refer to a mature and secreted cytokine protein or a biologically active fragment thereof.
  • a soluble cytokine protein can include a sequence that is at least 70% identical, at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical to a wildtype mature and secreted mammalian interleukin protein (e.g., a wildtype human interleukin protein) and retains its biological activity.
  • a wildtype mature and secreted mammalian interleukin protein e.g., a wildtype human interleukin protein
  • soluble interleukin receptor is used herein in the broadest sense to refer to a polypeptide that lacks a transmembrane domain (and optionally an intracellular domain) that is capable of binding one or more of its natural ligands (e.g., under physiological conditions, e.g., in phosphate buffered saline at room temperature).
  • a soluble interleukin receptor can include a sequence that is at least 70% identical (e.g., at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical) to an extracellular domain of wildtype interleukin receptor and retains its ability to specifically bind to one or more of its natural ligands, but lacks its transmembrane domain (and optionally, further lacks its intracellular domain).
  • soluble interleukin receptors are described herein.
  • soluble cytokine receptor is used herein in the broadest sense to refer to a polypeptide that lacks a transmembrane domain (and optionally an intracellular domain) that is capable of binding one or more of its natural ligands (e.g., under physiological conditions, e.g., in phosphate buffered saline at room temperature).
  • a soluble cytokine receptor can include a sequence that is at least 70% identical (e.g., at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical) to an extracellular domain of wildtype cytokine receptor and retains its ability to specifically bind to one or more of its natural ligands, but lacks its transmembrane domain (and optionally, further lacks its intracellular domain).
  • soluble cytokine receptors are described herein.
  • antibody is used herein in its broadest sense and includes certain types of immunoglobulin molecules that include one or more antigen-binding domains that specifically bind to an antigen or epitope.
  • An antibody specifically includes, e.g., intact antibodies (e.g., intact immunoglobulins), antibody fragments, and multi-specific antibodies.
  • an antigen-binding domain is an antigen-binding domain formed by a VH -VL dimer. Additional examples of an antibody are described herein. Additional examples of an antibody are known in the art.
  • “Affinity” refers to the strength of the sum total of non-covalent interactions between an antigen-binding site and its binding partner (e.g., an antigen or epitope).
  • affinity refers to intrinsic binding affinity, which reflects a 1:1 interaction between members of an antigen-binding domain and an antigen or epitope.
  • the affinity of a molecule X for its partner Y can be represented by the dissociation equilibrium constant (KD).
  • KD dissociation equilibrium constant
  • the kinetic components that contribute to the dissociation equilibrium constant are described in more detail below.
  • Affinity can be measured by common methods known in the art, including those described herein. Affinity can be determined, for example, using surface plasmon resonance (SPR) technology (e.g., BIACORE®) or biolayer interferometry (e.g., FORTEBIO®).
  • SPR surface plasmon resonance
  • FORTEBIO® biolayer interferometry
  • pair of affinity domains is two different protein domain(s) that bind specifically to each other with a KD of less than of less than 1 x 10 -7 M (e.g., less than 1 x 10 -8 M, less than 1 x 10 -9 M, less than 1 x 10 -10 M, or less than 1 x 10 -11 M).
  • a pair of affinity domains can be a pair of naturally-occurring proteins.
  • a pair of affinity domains can be a pair of synthetic proteins. Non- limiting examples of pairs of affinity domains are described herein.
  • epipe means a portion of an antigen that specifically binds to an antigen-binding domain.
  • Epitopes can, e.g., consist of surface-accessible amino acid residues and/or sugar side chains and may have specific three-dimensional structural characteristics, as well as specific charge characteristics. Conformational and non- conformational epitopes are distinguished in that the binding to the former but not the latter may be lost in the presence of denaturing solvents. An epitope may comprise amino acid residues that are directly involved in the binding, and other amino acid residues, which are not directly involved in the binding. Methods for identifying an epitope to which an antigen-binding domain binds are known in the art. The term “treatment” means to ameliorate at least one symptom of a disorder.
  • the disorder being treated is cancer and to ameliorate at least one symptom of cancer includes reducing aberrant proliferation, gene expression, signaling, translation, and/or secretion of factors.
  • the methods of treatment include administering a therapeutically effective amount of a composition that reduces at least one symptom of a disorder to a subject who is in need of, or who has been determined to be in need of such treatment.
  • all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Methods and materials are described herein for use in the present invention; other, suitable methods and materials known in the art can also be used. The materials, methods, and examples are illustrative only and not intended to be limiting.
  • Figures 1A-1B show the results of immunostimulation of an exemplary multi- chain polypeptide in C57BL/6 mice.
  • Figure 1A shows the spleen weight of mice treated with increasing dosage of the exemplary multi-chain polypeptide as compared to mice treated with the control solution.
  • Figure 1B shows the percentages of immune cell types present in the spleen of mice treated with increasing dosage of the exemplary multi-chain polypeptide as compared to mice treated with the control solution.
  • Figures 2A-2B show the duration of immunostimulation of an exemplary multi- chain polypeptide in C57BL/6 mice.
  • Figure 2A shows the spleen weight over a period of 92 hours in mice treated with 3 mg/kg of the exemplary multi-chain polypeptide.
  • Figure 2B shows the percentages of immune cell types present in the spleen over a period of 92 hours in mice treated with 3 mg/kg of the exemplary multi-chain polypeptide.
  • Figures 3A-3B show the expression of Ki67 and Granzyme B in immune cells induced by the exemplary multi-chain polypeptide.
  • Figure 3A shows the expression of Ki67 in CD4 + T cells, CD8 + T cells, natural killer (NK) cells, and CD19 + B cells at various time points post-treatment with the multi-chain polypeptide.
  • Figure 3B shows the expression of Granzyme B in CD4 + T cells, CD8 + T cells, natural killer (NK) cells, and CD19 + B cells at various time points post-treatment with the multi-chain polypeptide.
  • Figure 4 shows the effect of tumor inhibition by splenocytes prepared from mice treated with an exemplary multi-chain polypeptide at various time points after treatment.
  • Figures 5A-5B show the percentages and the proliferation rate of CD4 + T cells, CD8 + T cells, Natural Killer (NK) cells, and CD19 + B cells in the blood of B6.129P2- ApoE tm1Unc /J mice (purchased from The Jackson Laboratory) fed a control diet, a high fat diet and untreated, and mice fed a high fat diet and treated with TGFRt15-TGFRs, 2t2, or 21t15-TGFRs.
  • Figure 5A shows the percentages of the different cell types in each control and experimental group.
  • Figure 5B shows the proliferation rate of the of the different cell types in each control and experimental group.
  • Figures 6A-6E show exemplary physical appearance of mice fed either a control or high fat diet and were either untreated or treated with TGFRt15-TGFRs, 2t2, or 21t15- TGFRs.
  • Figure 7 shows the fasting body weight of mice fed either a control or a high fat diet and were either untreated or treated with TGFRt15-TGFRs, 2t2, or 21t15-TGFRs.
  • Figure 8 shows the fasting blood glucose levels of mice fed either a control or a high fat diet and were either untreated or treated with TGFRt15-TGFRs, 2t2, or 21t15- TGFRs.
  • Figures 9A-9F show chemotherapy-induced senescent B16F10 cells and expression of senescent genes.
  • Figure 9A shows chemotherapy induction of senescent B16F10 cells visualized using SA ⁇ -gal staining.
  • Figures 9B-9F show expression of p21 CIP1 , IL6, DPP4, RATE1E, and ULBP1 over time in the chemotherapy-induced senescent B16F10 cells.
  • Figures 10A-10F show colony formation and expression of stem cell markers by chemotherapy-induced senescent B16F10 cells.
  • Figure 10A shows colony formation by chemotherapy-induced senescent B16F10 cells.
  • Figures 10B and 10C show expression of Oct4 mRNA and Notch4 mRNA by chemotherapy-induced senescent B16F10 cells as compared to control B16F10 cells.
  • Figures 10D-10F show percentage of chemotherapy- induced senescent B16F10 cells double-positive for two out of the three stem cell markers including CD44, CD24, and CD133.
  • Figures 11A-11C show migratory and invasive properties of chemotherapy- induced senescent B16F10 cells.
  • Figure 11A shows the results of a migration assay comparing chemotherapy-induced senescent cells with stem cell properties (B16F10- SNC-CSC) with control B16F10 cells.
  • Figures 11B and 11C show the results of an invasion assay comparing chemotherapy-induced senescent cells with stem cell properties (B16F10-SNC-CSC) with control B16F10 cells.
  • Figures 12A and 12B show in vitro expanded NK cells and their cytotoxicity against chemotherapy-induced senescent cells with stem cell properties (B16F10-SNC- CSC) or control B16F10 cells.
  • Figure 12A shows an exemplary schematic of a process of obtaining in vitro expanded NK cells.
  • Figure 12 B shows cytotoxicity of the expanded NK cells against chemotherapy-induced senescent cells with stem cell properties (B16F10-SNC-CSC) or control B16F10 cells.
  • Figures 13A-13C show results of combination treatment using a mouse melanoma model.
  • Figure 13A shows an exemplary schematic for treating melanoma in a mouse model.
  • Figures 13B and 13C show the change in tumor volume over time with combination treatments including TGFRt15-TGFRs as compared to chemotherapy or TA99 treatment alone.
  • Figure 14 shows induction of senescence in the human pancreatic tumor cell line SW1990 and expression of CD44 and CD24 in senescent SW1990 cells as compared to control SW1990 cells.
  • Figure 15 shows expression of senescent markers by chemotherapy-induced senescent SW1990 cells.
  • Figure 16 shows the cytotoxicity of in vitro activated human NK cells against chemotherapy-induced senescent SW1990 cells or control SW1990 cells.
  • Figure 17 shows a schematic diagram of an exemplary IL-12/IL-15R ⁇ Su DNA construct.
  • Figure 18 shows a schematic diagram of an exemplary IL-18/TF/IL-15 DNA construct.
  • Figure 19 shows a schematic diagram of the interaction between the exemplary IL-12/IL-15R ⁇ Su and IL-18/TF/IL-15 DNA constructs.
  • Figure 20 shows a schematic diagram of the interaction between the exemplary IL-12/IL-15R ⁇ Su and IL-18/TF/IL-15 fusion proteins resulting in IL-18/TF/IL-15:IL- 12/IL-15R ⁇ Su complex (18t15-12s).
  • Figure 21 shows a chromatograph of 18t15-12s purification elution from an anti- TF antibody affinity column.
  • Figure 22 shows an exemplary chromatographic profile of anti-TF Ab /SEC- purified 18t15-12s protein following elution on an analytical size exclusion column, demonstrating separation of monomeric multiprotein 18t15-12s complexes from protein aggregates.
  • Figure 23 shows an example of a 4-12% SDS-PAGE of the 18t15-12s complex following disulfide bond reduction. Lane 1: SeeBlue Plus2 marker; Lane 2: anti-TF Ab- purified 18t15-12s (0.5 ⁇ g); Lane 3: anti-TF Ab-purified 18t15-12s (1 ⁇ g).
  • Figure 24 shows SDS PAGE analysis of deglycosylated and non-deglycosylated 18t15-12s.
  • Lane 1 anti-TF Ab-purified 18t15-12s (0.5 ⁇ g), non-deglycosylated; Lane 2: anti-TF Ab -purified 18t15-12s (1 ⁇ g), non-deglycosylated; Lane 3: 18t15-12s (1 ⁇ g), deglycosylated, Lane 4: Mark12 unstained maker.
  • Figure 25 shows a sandwich ELISA for the 18t15-12s complex, comprising an anti-human tissue factor antibody capture and a biotinylated anti-human IL-12 detection antibody (BAF 219).
  • Figure 26 shows a sandwich ELISA for the 18t15-12s complex, comprising an anti-human tissue factor antibody capture and a biotinylated anti-human IL-15 detection antibody (BAM 247).
  • Figure 27 shows a sandwich ELISA for the 18t15-12s complex, comprising an anti-human tissue factor antibody capture and a biotinylated anti-human IL-18 detection antibody (D045-6).
  • Figure 28 shows a sandwich ELISA for the 18t15-12s complex, comprising an anti-human tissue factor (I43) capture antibody and an anti-human tissue factor detection antibody.
  • Figure 29 shows proliferation of IL-15-dependent 32D ⁇ cells mediated by the 18t15-12s complex (open squares) and recombinant IL-15 (black squares).
  • Figure 30 shows biological activity of IL-18 within the 18t15-12s complex (open squares), where recombinant IL-18 (black squares) and recombinant IL-12 (black circles) serve as positive and negative controls, respectively.
  • Figure 31 shows biological activity of IL-12 within the 18t15-12s complex (open squares), where recombinant IL-12 (black circles) and recombinant IL-18 (open squares) serve as positive and negative controls, respectively.
  • Figures 32A and 32B show cell-surface expression of CD25 on NK cells induced by the 18t15-12s complex and cell-surface CD69 expression of NK cells induced by the 18t15-12s complex.
  • Figure 33 shows a flow cytometry graph of intracellular IFN- ⁇ expression of NK cells induced by the 18t15-12s complex.
  • Figure 34 shows cytotoxicity of 18t15-12s induced human NK cells against K562 cells.
  • Figure 35 shows a schematic diagram of an exemplary IL-12/IL-15R ⁇ Su/ ⁇ CD16 DNA construct.
  • Figure 36 shows a schematic diagram of an exemplary IL-18/TF/IL-15 DNA construct.
  • Figure 37 shows a schematic diagram of the interaction between the exemplary IL-12/IL-15R ⁇ Su/ ⁇ CD16scFv and IL-18/TF/IL-15 DNA constructs.
  • Figure 38 shows a schematic diagram of an exemplary 18t15-12s/ ⁇ CD16 protein complex.
  • Figure 39 shows a sandwich ELISA for the 18t15-12s16 complex, comprising an anti-human tissue factor antibody capture antibody and a biotinylated anti-human IL-12 (BAF 219) (dark line) or an anti-human tissue factor detection antibody (light line).
  • Figure 40 shows a schematic diagram of an exemplary TGF ⁇ RII/IL-15R ⁇ Su DNA construct.
  • Figure 41 shows a schematic diagram of an exemplary IL-21/TF/IL-15 construct.
  • Figure 42 shows a schematic diagram of the interaction between the exemplary IL- IL-21/TF/IL-15 and TGF ⁇ RII/IL-15R ⁇ Su constructs.
  • Figure 43 shows a schematic diagram of the interaction between the exemplary TGF ⁇ RII/IL-15R ⁇ Su and IL-21/TF/IL-15 fusion proteins, resulting in an IL-21/TF/IL- 15/TGF ⁇ RII/IL-15R ⁇ Su complex (21t15-TGFRs).
  • Figure 44 shows a chromatograph of 21t15-TGFRs purification elution from an anti-TF antibody affinity column.
  • Figure 45 shows an exemplary 21t15-TGFRs size exclusion chromatograph showing a main protein peak and a high molecular weight peak
  • Figure 46 shows an example of a 4-12% SDS-PAGE of the 21t15-TGFRs complex following disulfide bond reduction.
  • Lane 1 Mark12 unstained marker (numbers on the left side indicate molecular weights in kDa); Lane 2: 21t15-TGFRs (0.5 ⁇ g); Lane 3: 21t15-TGFRs (1 ⁇ g); Lane 4: 21t15-TGFRs, deglycosylated (1 ⁇ g), wherein the MW was the expected size of 53kDa and 39.08 kDa.
  • Figure 47 shows a sandwich ELISA for the 21t15-TGFRs complex, comprising an anti-human tissue factor capture and a biotinylated anti-human IL-21 detection antibody (13-7218-81, BioLegend).
  • Figure 48 shows a sandwich ELISA for the 21t15-TGFRs complex, comprising an anti-human tissue factor antibody capture and a biotinylated anti-human IL-15 detection antibody (BAM 247, R&D Systems).
  • Figure 49 shows a sandwich ELISA for the 21t15-TGFRs complex, comprising an anti-human tissue factor antibody capture and a biotinylated anti-human TGF ⁇ RII detection antibody (BAF241, R&D Systems).
  • Figure 50 shows a sandwich ELISA for the 21t15-TGFRs complex, comprising an anti-human tissue factor (I43) capture antibody and an anti-human tissue factor detection antibody.
  • I43 anti-human tissue factor
  • Figure 51 shows IL-15-dependent proliferation of 32D ⁇ cells mediated by the 21t15-TGFRs complex (open squares) compared to IL-15 (black squares).
  • Figure 52 shows biological activity of the TGF ⁇ RII domain within the 21t15- TGFRs complex (open squares). TGF ⁇ RII/Fc (black squares) served as a positive control.
  • Figure 53 shows a flow cytometry graph of cell-surface CD25 expression of NK cells induced by the 21t15-TGFRs complex.
  • Figure 54 shows a flow cytometry graph of cell-surface CD69 expression of NK cells induced by the 21t15-TGFRs complex.
  • Figure 55 shows a flow cytometry graph of intracellular IFN- ⁇ expression of NK cells induced by the 21t15-TGFRs complex.
  • Figure 56 shows cytotoxicity of 21t15-TGFRs-induced human NK cells against K562 cells.
  • Figure 57 are schematic diagrams of an exemplary ⁇ CD3scFv/TF/ ⁇ CD28scFv single-chain chimeric polypeptide.
  • Figure 58 is a chromatograph showing the elution of an exemplary ⁇ CD3scFv/TF/ ⁇ CD28scFv single-chain chimeric polypeptide from an anti-tissue factor affinity column.
  • Figure 59 is a chromatograph showing the elution of a Superdex 200 Increase 10/300 GL gel filtration column loaded with an exemplary ⁇ CD3scFv/TF/ ⁇ CD28scFv single-chain chimeric polypeptide.
  • Figure 60 is a sodium dodecyl sulfate polyacrylamide gel (4-12% NuPage Bis- Tris gel) of an exemplary ⁇ CD3scFv/TF/ ⁇ CD28scFv single-chain chimeric polypeptide purified using an anti-tissue factor affinity column.
  • Figure 61 is a graph showing the ELISA quantitation of an exemplary ⁇ CD3scFv/TF/ ⁇ CD28scFv single-chain chimeric polypeptide performed using the methods described in Example 1. Purified tissue factor was used as the control.
  • Figure 62 is a graph showing the ability of an exemplary ⁇ CD3scFv/TF/ ⁇ CD28scFv single-chain chimeric polypeptide to stimulate CD25 expression in CD4 + T-cells isolated from blood from two donors. The experiments were performed as described in Example 2.
  • Figure 63 is a graph showing the ability of an exemplary ⁇ CD3scFv/TF/ ⁇ CD28scFv single-chain chimeric polypeptide to stimulate CD25 expression in CD8 + T-cells isolated from blood from two donors. The experiments were performed as described in Example 2.
  • Figure 64 is a graph showing the ability of an exemplary ⁇ CD3scFv/TF/ ⁇ CD28scFv single-chain chimeric polypeptide to stimulate CD69 expression in CD4 + T-cells isolated from blood from two donors. The experiments were performed as described in Example 2.
  • Figure 65 shows a schematic diagram of an exemplary IL-7/IL-15R ⁇ Su DNA construct.
  • Figure 66 shows a schematic diagram of an exemplary IL-21/TF/IL-15 DNA construct.
  • Figure 67 shows a schematic diagram of the interaction between the exemplary IL-7/IL-15R ⁇ Su and IL-21/TF/IL-15 DNA constructs.
  • Figure 68 shows a schematic diagram of the interaction between the exemplary IL-7/IL-15R ⁇ Su and IL-21/TF/IL-15 fusion proteins resulting in an IL-21/TF/IL-15:IL- 7/IL-15R ⁇ Su complex (21t15-7s).
  • Figure 69 shows a schematic diagram of an exemplary IL-21/IL-15R ⁇ Su DNA construct.
  • Figure 70 shows a schematic diagram of an exemplary IL-7/TF/IL-15 DNA construct.
  • Figure 71 shows a schematic diagram of the interaction between the exemplary IL-21/IL-15R ⁇ Su and IL-7/TF/IL-15 DNA constructs.
  • Figure 72 shows a schematic diagram of the interaction between the exemplary IL-21/IL-15R ⁇ Su and IL-7/TF/IL-15 fusion proteins resulting in an IL-7/TF/IL-15:IL- 21/IL-15R ⁇ SU complex (7t15-21s).
  • Figure 73 shows the oxygen consumption rate (OCR) in pmoles/min for human NK cells isolated from blood (2 x 10 6 cells/mL) of two different donors.
  • Figure 74 shows the extracellular acidification rate (ECAR) in mpH/minute for human NK cells isolated from blood (2 x 10 6 cells/mL) of two different donors.
  • Figure 75 shows a schematic of the 7t15-16s21 construct.
  • Figure 76 shows an additional schematic of the 7t15-16s21 construct.
  • Figures 77A and 77B show binding of 7t15-16s21 to CHO cells expressing human CD16b as compared to a control protein.
  • Figures 78A-78C are results from ELISA experiments using antibodies against IL-15, IL-21, and IL-7 in detecting 7t15-16s21.
  • Figure 79 shows results of the 32D ⁇ cell proliferation assay with 7t15-16s21 or recombinant IL-15.
  • Figure 80 shows the chromatographic profile of 7t15-16s21 protein containing cell culture supernatant following binding and elution on anti-TF antibody resin.
  • Figure 81 shows the analytical SEC Profile of 7t15-16s21.
  • Figure 82 shows a schematic of the TGFRt15-16s21 construct.
  • Figure 83 shows an additional schematic of the TGFRt15-16s21 construct.
  • Figures 84A and 84B show binding affinity of TGFRT15-16S21 and 7t15-21s with CHO cells expressing human CD16b.
  • Figure 84A shows binding affinity of TGFRT15-16S21 with CHO cells expressing human CD16b.
  • Figure 84B shows binding affinity of 7t15-21s with CHO cells expressing human CD16b.
  • Figure 85 shows results of TGF ⁇ 1 inhibition by TGFRt15-16s21 and TGFR-Fc.
  • Figure 86 shows results of 32D ⁇ cell proliferation assay with TGFRt15-16s21 or recombinant IL-15.
  • Figures 87A-87C show results of detecting IL-15, IL-21, and TGF ⁇ RII in TGFRt15-16s21 with corresponding antibodies using ELISA.
  • Figure 88 shows the chromatographic profile of TGFRt15-16s21 protein containing cell culture supernatant following binding and elution on anti-TF antibody resin.
  • Figure 89 shows results of a reduced SDS-PAGE analysis of TGFRt15-16s21.
  • Figure 90 shows a schematic of the 7t15-7s construct.
  • Figure 91 shows an additional schematic of the 7t15-7s construct.
  • Figure 92 shows the chromatographic profile of 7t15-7s protein containing cell culture supernatant following binding and elution on anti-TF antibody resin.
  • Figure 93 shows detection of TF, IL-15 and IL-7 in 7t15-7s using ELISA.
  • Figures 94A and 94B show spleen weight and the percentages of immune cell types in 7t15-7s -treated and control-treated mice.
  • Figure 94A shows spleen weight in mice treated with 7t15-7s as compared to PBS control.
  • Figure 94B shows the percentage of CD4 + T cells, CD8 + T cells, and NK cells in mice treated with 7t15-7s as compared to PBS control.
  • Figure 95 shows a schematic of the TGFRt15-TGFRs construct.
  • Figure 96 shows an additional schematic of the TGFRt15-TGFRs construct.
  • Figure 97 shows results of TGF ⁇ 1 inhibition by TGFRt15-TGFRs and TGFR-Fc.
  • Figure 98 shows results of 32D ⁇ cell proliferation assay with TGFRt15-TGFRs or recombinant IL-15
  • Figures 99A and 99B show results of detecting IL-15 and TGF ⁇ RII in TGFRt15- TGFRs with corresponding antibodies using ELISA.
  • Figure 100 is a line graph showing the chromatographic profile of TGFRt15- TGFRs protein containing cell culture supernatant following binding and elution on anti- TF antibody resin.
  • Figure 101 shows the analytical SEC profile of TGFRt15-TGFRs.
  • Figure 102 shows TGFRt15-TGFRs before and after deglycosylation as analyzed by reduced SDS-PAGE.
  • Figures 103A and 103B show spleen weight and the percentages of immune cell types in TGFRt15-TGFRs-treated and control-treated mice.
  • Figure 103A shows spleen weight in mice treated with TGFRt15-TGFRs as compared to PBS control.
  • Figure 103B shows the percentage of CD4 + T cells, CD8 + T cells, and NK cells in mice treated with TGFRt15-TGFRs as compared to PBS control.
  • Figure 104A and 104B show the spleen weight and immunostimulation over 92 hours in mice treated with TGFRt15-TGFRs.
  • Figure 104A shows spleen weight of mice treated with TGFRt15-TGFRs at 16, 24, 48, 72, and 92 hours after treatment.
  • Figure 104B shows the percentages of immune cells in mice treated with TGFRt15-TGFRs at 16, 24, 48, 72, and 92 hours after treatment.
  • Figure 105A and 105B show Ki67 and Granzyme B expression in mice treated with TGFRt15-TGFRs over time.
  • Figure 106 shows enhancement of cytotoxicity of splenocytes by TGFRt15- TGFRs in C57BL/6 Mice.
  • Figure 107 shows changes in tumor size in response to PBS treatment, chemotherapy alone, TGFRt15-TGFRs alone, or chemotherapy and TGFRt15-TGFRs combination, in a pancreatic cancer mouse model.
  • Figure 108 shows the cytotoxicity of NK cells isolated from mice treated with TGFRt15-TGFRs.
  • Figure 109 shows a schematic of the 7t15-21s137L (long version) construct.
  • Figure 110 shows an additional schematic of the 7t15-21s137L (long version) construct.
  • Figure 111 is a line graph showing the chromatographic profile of 7t15-21s137L (long version) protein containing cell culture supernatant following binding and elution on anti-TF antibody resin.
  • Figure 112 shows the analytical SEC profile of 7t15-21s137L (long version).
  • Figure 113 shows binding of 7t15-21s137L (short version) to CD137L (4.1BBL)
  • Figures 114A-114C show detection of IL-15, IL21, and IL7 in 7t15-21s137L (short version) with ELISA.
  • Figure 114A shows detection of IL-15 in 7t15-21s137L (short version) with ELISA.
  • Figure 114B shows detection of IL21 in 7t15-21s137L (short version) with ELISA.
  • Figure 114C shows detection of IL7 in 7t15-21s137L (short version) with ELISA.
  • Figure 115 shows results from a CTLL-2 cell proliferation assay.
  • Figure 116 shows the activity of 7t15-1s137L (short version) in promoting IL21R containing B9 cell proliferation.
  • Figure 117 shows a schematic of the 7t15-TGFRs construct.
  • Figure 118 shows an additional schematic of the 7t15-TGFRs construct.
  • Figure 119 shows results of TGF ⁇ 1 inhibition by 7t15-TGFRs and TGFR-Fc.
  • Figures 120A-120C show detection of IL-15, TGF ⁇ RII, and IL-7 in 7t15-TGFRs with ELISA.
  • Figure 121 shows results of a 32D ⁇ cell proliferation assay with 7t15-TGFRs or recombinant IL-15.
  • Figure 122 is a line graph showing the chromatographic profile of 7t15-TGFRs protein containing cell culture supernatant following binding and elution on anti-TF antibody resin.
  • Figure 123 shows 7t15-TGFRs before and after deglycosylation as analyzed using reduced SDS-PAGE.
  • Figure 124 shows ELISA detection of IL-7, IL-15 and TGF ⁇ RII in the 7t15- TGFRs protein.
  • Figures 125A and 125B show spleen weight and the percentages of immune cell types in 7t15-TGFRs-treated and control-treated mice.
  • Figure 125A shows spleen weight in mice treated with 7t15-TGFRs at various dosages, as compared to PBS control.
  • Figure 125B shows the percentage of CD4 + T cells, CD8 + T cells, and NK cells in mice treated with 7t15-TGFRs at various dosages, as compared to PBS control.
  • F igures 126A and 126B show upregulation of CD44 expression of CD4 + and CD8 + T cells by 7t15-TGFRs in C57BL/6 mice.
  • Figures 127A and 127B show upregulation of Ki67 expression and Granzyme B expression of CD8 + T cells and NK Cells by 7t15-TGFRs in C57BL/6 mice.
  • Figure 128 shows enhancement of cytotoxicity of splenocytes by 7t15-TGFRs in C57BL/6 mice.
  • Figure 129 shows a schematic of the TGFRt15-21s137L construct.
  • Figure 130 shows an additional schematic of the TGFRt15-21s137L construct.
  • Figure 131 is a line graph showing the chromatographic profile of TGFRt15-21s137L protein containing cell culture supernatant following binding and elution on anti-TF antibody resin.
  • Figure 132 shows a schematic of the TGFRt15-TGFRs21 construct.
  • Figure 133 shows an additional schematic of the TGFRt15-TGFRs21 construct.
  • Figure 134 is a line graph showing the chromatographic profile of TGFRt15- TGFRs21 protein containing cell culture supernatant following binding and elution on anti-TF antibody resin.
  • Figure 135 shows TGFRt15-TGFRs21 before and after deglycosylation as analyzed by reduced SDS-PAGE.
  • Figures 136A and 136B show detection of components of TGFRt15-TGFRs21 using ELISA.
  • Figures 137A and 137B show the percentages and proliferation of CD4 + T cells, CD8 + T cells, and natural killer (NK) cells present in the spleen of control-treated and TGFRt15-TGFRs21-treated mice.
  • Figure 138 shows upregulation of Granzyme B expression of splenocytes in mice treated with TGFRt15-TGFRs21.
  • Figure 139 shows enhancement of cytotoxicity of splenocytes by TGFRt15-TGFRs21 in C57BL/6 Mice.
  • Figure 140 shows a schematic of the TGFRt15-TGFRs16 construct.
  • Figure 141 shows an additional schematic of the TGFRt15-TGFRs16 construct.
  • Figure 142 shows a schematic of the TGFRt15-TGFRs137L construct.
  • Figure 143 shows an additional schematic of the TGFRt15-TGFRs137L construct.
  • Figure 144 are schematic diagrams of an exemplary 2t2 single-chain chimeric polypeptide.
  • Figure 145 shows IL-2 activity in 2t2 as compared to recombinant IL-2 using a 32D ⁇ cell proliferation assay.
  • Figure 146 shows IL-2 activity in 2t2 as compared to recombinant IL-2 using a CTLL-2 cell proliferation assay.
  • Figure 147 shows the fasting blood glucose levels in ApoE -/- mice fed with standard chow or a high fat diet and treated with a PBS control (untreated) or with 2t2.
  • Figure 148 shows the ratio of CD4 + CD25 + FoxP3 + T regulatory cells in blood lymphocytes from ApoE -/- mice fed with standard chow or a high fat diet and treated with a PBS control (untreated) or with 2t2.
  • Figure 149 is a line graph showing the chromatographic profile of 2t2 protein containing cell culture supernatant following binding and elution on anti-TF antibody resin.
  • Figure 150 shows an analytical SEC profile of 2t2.
  • Figures 151A and 151B show reduced SDS-PAGE analysis of 2t2 before and after deglycosylation.
  • Figure 151A shows reduced SDS-PAGE analysis of 2t2 before deglycosylation.
  • Figure 151B shows reduced SDS-PAGE analysis of 2t2 after deglycosylation.
  • Figures 152A and152B show results of immunostimulation in C57BL/6 mice using 2t2.
  • Figure 152A shows spleen weight following treatment with 2t2.
  • Figure 152B shows the percentages of immune cell types following 2t2 treatment.
  • Figure 153 shows upregulation of CD25 expression of CD4 + T cells in mice treated with 2t2.
  • Figure 154 shows the pharmacokinetics of 2t2 in C57BL/6 mice.
  • Figures 155A and 155B show effects of 2t2 in attenuating the formation of high fat-induced atherosclerotic plaques in ApoE -/- mice.
  • Figure 155A shows a representative view of atherosclerotic plaques from ApoE -/- mice fed with standard chow or a high fat diet and treated with either PBS control or 2t2.
  • Figure 155B shows the results of quantitative analysis of atherosclerotic plaques of each group.
  • Figure 156 shows fasting glucose levels in 2t2 treated-mice as compared to control-treated mice.
  • Figure 157 shows the percentage of CD4 + CD25 + FoxP3 + Tregs in blood lymphocytes from mice treated with 2t2 and control-treated mice.
  • Figure 158 are schematic diagrams of an exemplary 15t15 single-chain chimeric polypeptide.
  • Figure 159 shows the IL-15 activity of 15t15 as compared to recombinant IL-15 in a 32D ⁇ cell proliferation assay.
  • Figure 160 is a line graph showing the chromatographic profile of 15t15 protein containing cell culture supernatant following binding and elution on anti-TF antibody resin.
  • Figures 161A and 161B show reduced SDS-PAGE analysis of 15t15 before and after deglycosylation.
  • Figure 161A shows reduced SDS-PAGE analysis of 15t15 before deglycosylation.
  • Figure 161B shows reduced SDS-PAGE analysis of 15t15 after deglycosylation.
  • Figures 162A and 162B is a set of histograms (Figure 162A) and a set of graphs ( Figure 162B) showing the change in the surface phenotype of NK cells after stimulation with 18t15-12s, 18t15-12s16, and 7t15-21s + anti-TF antibody.
  • Figure 163 is a set of graphs showing changes in the surface phenotype of lymphocyte populations after stimulation with 18t15-12s, 18t15-12s16, and 7t15-21s.
  • Figure 164 is a set of graphs showing an increase in glycolysis in NK cells following treatment with 18t15-12s.
  • Figure 165 is a set of graphs showing an increase in phospho-STAT4 and phospho-STAT5 levels in NK cells after stimulation with 18t15-12s.
  • Figure 166 is a set of graphs showing that overnight stimulation of NK cells with 18t15-12s enhances cell metabolism.
  • Figure 167A-C is a set of graphs showing immunostimulation in C57BL/6 mice following treatment with 2t2.
  • Figure 168A-B is a set of graphs showing immunostimulation in C57BL/6 mice following treatment with TGFRt15-TGFRs.
  • Figure 169A-C is a set of graphs showing in vivo stimulation of Tregs, NK cells, and CD8 + T cells in ApoE -/- mice fed with a Western diet and treated with TGFRt15- TGFRs or 2t2.
  • Figure 170A-B is a set of graphs showing induction of splenocyte proliferation by 2t2 in C57BL/6 mice.
  • Figure 171A-C is a set of graphs showing immunostimulation in C57BL/6 mice following treatment with TGFRt15-TGFRs.
  • Figure 172A-B is a set of graphs showing in vivo induction of proliferation of NK cells and CD8 + T cells in ApoE -/- mice fed with a Western diet and treated with TGFRt15-TGFRs or 2t2.
  • Figure 173 is a schematic and a set of graphs showing the persistence of 7t15-21s and anti-TF antibody-expanded NK cells in NSG mice following treatment with 7t15-21, TGFRt15-TGFRs or 2t2.
  • Figure 174A-B is a set of graphs showing enhancement of cytotoxicity of NK cells following treatment of NK cells with TGFRt15-TGFRs.
  • Figure 175A-B is a set of graphs showing enhancement of ADCC activity of NK cells following treatment of NK cells with TGFRt15-TGFRs.
  • Figure 176 is a graph of in vitro killing of senescent B16F10 melanoma cells by TGFRt15-TGFRs/2t2-activated mouse NK cells.
  • Figure 177A-H is a set of graphs showing antitumor activity of TGFRt15-TGFRs plus anti-TRP1 antibody (TA99) in combination with chemotherapy in a melanoma mouse model.
  • Figure 178A-C is a set of graphs showing amelioration of the Western diet- induced hyperglycemia in ApoE -/- mice by 2t2.
  • Figure 179 is a set of graphs showing cell surface staining summarizing the differentiation of NK cells into cytokine-induced memory like NK cells (CIML-NK Cells) after stimulation with 18t15-12s and cultured in rhIL-15.
  • Figure 180 shows upregulation of CD44 memory T cells. The upper panel shows upregulation of CD44 memory T cells upon treatment with TGFRt15-TGFRs. The lower panel shows upregulation of CD44 memory T cells upon treatment with 2t2.
  • Figures 181A and 181B show improvement in hair regrowth following depilation in mice treated with 2t2 or IL-2.
  • Figure 181A shows skin pigmentation 10 days after depilation in PBS-, 2t2-, or IL-2-treated mice.
  • Figure 181B shows percent pigmentation in PBS-, 2t2-, or IL-2-treated mice as analyzed using the ImageJ software.
  • Figure 182 shows skin pigmentation 14 days after depilation in PBS-, 2t2-, or IL- 2-treated mice.
  • Figure 183 shows a graph of Factor X (FX) activation following treatment with single-chain or multi-chain chimeric polypeptides.
  • Figure 184 shows clotting time for a buffer with varying concentrations of Innovin in a prothrombin time (PT) test.
  • Figure 185 shows clotting time for multi-chain chimeric polypeptides in a PT Assay.
  • Figure 186 shows clotting time of the multi-chain chimeric polypeptides in a PT assay when mixed with 32DB cells.
  • Figure 187 shows clotting time of multi-chain chimeric polypeptides in a PT assay when mixed with human PBMC.
  • Figure 188 shows binding of 7t15-21s137L (long version) and 7t15-21s137L (short version) to CD137 (4.1BB).
  • Figure 189A-189D show detection of IL7, IL21, IL15, and 4.1BBL in 7t15- 21s137L (long version) by the respective antibodies using ELISA.
  • Figure 190 shows IL-15 activity of 7t15-21s137L (long version) and 7t15- 21s137L (short version) as evaluated by an IL2R ⁇ ⁇ ⁇ -containing CTLL2 cell proliferation assay.
  • Figures 191A-191C show human blood lymphocyte pStat5a responses in CD4 + CD25 hi T reg cells, CD4 + CD25-T con cells, or in CD8 + T con cells in response to 2t2 or IL2 treatment.
  • Figure 191A shows pSTAT5 responses in CD4 + CD25 hi Treg cells.
  • Figure C191B shows pSTAT5 responses in CD4 + CD25-T con cells.
  • Figure 191C shows pSTAT5 responses in CD8 + T con cells.
  • Figures 192A-192E is a set of imaging showing that treatment with an IL-2 based molecule (2t2) can induce formation of hair follicles following depilation in mouse model.
  • Figure 192A is an image from a control mouse - only depilation done after hair was shaved
  • Figure 192B is an image from a mouse where depilation was followed by low dose IL-2 (1 mg/kg) administration
  • Figures 192C-192E show images from mice where depilation was followed by 2t2 at 0.3 mg/kg, ( Figure 192C), 1 mg/kg ( Figure 192D), and ( Figure 192E) 3 mg/kg.
  • Black arrows indicate anagen-phase hair follicles that will later extend into dermis and facilitate hair growth.
  • Figure 193 shows the total number of anagen phase hair follicles counted per 10 fields for each treatment group.
  • Figure 194 is a graph showing the percentage different in DNA demethylation in NK cells (relative to unexposed NK cells) from two different donors following expansion with 7t15-21s+ anti-tissue factor (TF)-antibody (IgG1) (50 nM).
  • Figure 195 is a set of graphs showing the immune-phenotype from peripheral blood analysis after 4 days post single dose treatment with TGFRt15-TGFRs.
  • Figure 196 is a set of graphs showing the immune-phenotype from peripheral blood analysis after 4 days post single dose treatment with TGFRt15-TGFRs.
  • Figure 197 is a graph showing ⁇ -Gal staining analysis by FACS at seven days after the second administration with TGFRt15-TGFRs.
  • Figure 198 is a set of graphs showing the levels of senescence markers in liver tissue determined using qPCR at 7 days after the second administration with TGFRt15- TGFRs.
  • Figure 199 is a set of graphs showing the levels of senescence markers in kidney tissue determined using qPCR at 7 days after the second administration with TGFRt15- TGFRs.
  • Figure 200 is a set of graphs showing the levels of senescence markers in skin tissue determined using qPCR at 7 days after the second administration with TGFRt15- TGFRs.
  • Figure 201 is a set of graphs showing the levels of senescence markers in lung tissue determined using qPCR at 7 days after the second administration with TGFRt15- TGFRs.
  • Figure 202 is a set of histological images showing ⁇ -Gal staining on kidney tissue at 7 days post second treatment with TGFRt15-TGFRs.
  • Figures 203A-203C show chemotherapy induces p21 CIP1 p21 senescence- associated gene expression in C57BL/6 mice.
  • Figure 203A is an exemplary schematic showing the experimental treatment regimen.
  • Figures 203B and 203C are graphs showing expression of p21 CIP1 p21 in lung (B) and liver (C) tissues respectively.
  • Figure 204 is a set of graphs showing immune-phenotype and cell proliferation following treatment with IL-15-based agents at day 3 post treatment.
  • Figures 205A-205C are graphs showing TGFRt15-TGFRs treatment reduces senescence-associated gene expression in C57BL/6 mice. The graphs show expression of p21 CIP1 p21 and CD26 in lung (A and B) and p21 CIP1 p21 in liver (C) tissues respectively.
  • Figure 206 is a set of graphs showing CD4 + , CD8 + , and Treg cell percentages and proliferation.
  • Figure 207 is a set of graphs showing NK, CD19 + and monocyte cell percentages and proliferation.
  • Figures 208A-208C are graphs showing evaluation of senescence markers p21 CIP1 p21 and CD26 in lung and liver tissues.
  • Figures 208A and 208B show lung p21 CIP1 p21 (A) and lung CD26 (B) senescence markers.
  • Figure 208C shows liver p21 CIP1 p21 senescence marker.
  • Figure 209 shows a schematic diagram of the interaction between the exemplary TGF ⁇ RII/IL-15R ⁇ Su and TGF ⁇ RII/TF/IL-15Mut proteins resulting in TGFRt15*-TGFRs complex.
  • Figure 210 shows a schematic diagram of the interaction between the exemplary TGF ⁇ RII/IL-15R ⁇ Su and TGF ⁇ RII/TF/IL-15Mut proteins.
  • Figures 211A is a graph showing the binding activity of TGFRt15-TGFRs to TGF- ⁇ 1 and LAP.
  • Figure 211B is a graph showing the binding activity of TGFRII/Fc to TGF- ⁇ 1 and LAP.
  • Figure 211C is a graph showing the binding activity of TGFRt15-TGFRs to TGF- ⁇ 1 and LAP.
  • Figure 211D is a graph showing the binding activity of TGFRt15*-TGFRs to TGF- ⁇ 1 and LAP.
  • Figure 211E is a graph showing the binding activity of TGFRt15-TGFRs, TGFRt15*-TGFRs, and 7t15-21s to CTLL-2 cells.
  • Figure 212A is a graph of TGF- ⁇ 1 blocking activity of TGFRt15-TGFRs and TGFRt15*-TGFRs.
  • Figure 212B is a graph of the IL-15 biological activity of TGFRt15-TGFRs and TGFRt15*-TGFRs.
  • Figure 212C is a graph showing that TGF- ⁇ 1, TGF- ⁇ 2, and TGF- ⁇ 3 each similarly inhibit IL-4-induced CTLL-2 growth in the absence of TGFRt15*-TGFRs.
  • Figure 212D is a graph showing that TGFRt15*-TGFRs significantly reversed the inhibition of TGF- ⁇ 1 and TGF- ⁇ 3 of IL-4-induced CTLL-2 cell growth.
  • Figure 213A shows that there is no significant damage to the IL-15 domain of TGFRt15-TGFRs following 10-day incubation 4°C, 25 °C, or 37 °C.
  • Figure 213B shows that there is no significant damage to the TGF ⁇ -RII domain of TGFRt15-TGFRs following 10-day incubation 4°C, 25 °C, or 37 °C.
  • Figure 213C is a graph showing TGF- ⁇ 1 neutralizing activity of TGFRt15- TGFRs following incubation in human serum for 10 days at 4°C, 25 °C, or 37 °C.
  • Figure 213D is a graph showing IL-15 activity of TGFRt15-TGFRs following incubation in human serum for 10 days at 4 °C, 25 °C, or 37°C.
  • Figure 214A is a graph showing cell-mediated cell cytotoxicity in an assay using NK cells and the constructs shown.
  • Figure 214B is a graph showing cell-mediated cell cytotoxicity in an assay using PMBCs and the constructs shown.
  • Figure 214C is a graph showing intracellular granzyme B production in an assay using NK cells and the constructs shown.
  • Figure 214D is a graph showing intracellular granzyme B production in an assay using PBMCs and the constructs shown.
  • Figure 214E is a graph showing interferon-gamma production in an assay using NK cells and the constructs shown.
  • Figure 214F is a graph showing interferon-gamma production in an assay using PMBCs and the constructs shown.
  • Figure 215 is a graph showing the pharmacokinetics (half-life, t 1/2 ) of TGFRt15- TGFRs evaluated in female C57BL/6 mice.
  • Figure 216 is a graph showing toxicity of TGFRt15-TGFRs in C57BL/6 mice.
  • Figure 217 is a graph showing antitumor activity of TGFRt15-TGFRs in a C57BL/6 murine melanoma model.
  • Figure 218 shows activity of TGFRt15-TGFRs in nine-week old C57BL6/j male mice, wherein the mice were given 50 ⁇ l of bleomycin (2.5 mg/kg, single dose) through the oropharyngeal route and then were given TGFRt15-TGFRs subcutaneously (3 mg/kg) on day 17 following bleomycin treatment.
  • bleomycin 2.5 mg/kg, single dose
  • Figure 219 shows fasting plasma glucose levels in db/db mice 4 days post treatment with TGFRt15-TGFRs or TGFRt15*-TGFRs.
  • Figures 220A-220C show TGF ⁇ 1-3 levels in db/db mice 4 days post treatment with TGFRt15-TGFRs or TGFRt15*-TGFRs: TGF ⁇ 1 ( Figure 220A), TGF ⁇ 2 ( Figure 220B), and TGF ⁇ 3 ( Figure 220C).
  • Figures 221A-E show lymphocyte subsets in db/db mice 4 days post treatment with TGFRt15-TGFRs or TGFRt15*-TGFRs: blood NK cells ( Figure 221A), blood Ki67 + NK cells ( Figure 221B), blood granzyme B + (GzmB + ) ( Figure 221C), blood CD8 + ( Figure 221D), and blood CD8 + Ki67 + T cells (Figure 221E).
  • Figure 222A shows the interaction of TGFRt15*-TGFRs or TGFRt15-TGFRs with latent TGF ⁇ 1 (SLC) or with CD39 (control).
  • Figure 222B shows the interaction of TGFRt15*-TGFRs and TGFRII-Fc with latent TGF ⁇ 1.
  • Figure 223 is a graph showing the clotting time of Innovin in the PT assay.
  • Figure 224 is a graph showing the clotting time of TGFRt15-TGFRs in the PT assay.
  • Figure 225 are graphs showing gene expression of senescence markers PAI-1, IL- 1 ⁇ , IL6, and IL-1 ⁇ in kidney and comparing young vs PBS or TGFRt15-TGFRs treated aged mice with short term vs long term follow-up.
  • Figure 226 are graphs showing gene expression of senescence markers IL-1 ⁇ and IL6 in liver.
  • Figure 227 shows protein expression of senescence marker PAI-1 in kidney.
  • Figure 228 are graphs showing that IL15SA (positive control) or TGFRt15*- TGFRs + IL15SA mediated an increase in the percentages of CD3 + CD8 + , CD3-NK1.1 + , and CD3 + CD45 + immune cells in the blood, whereas treatment with TGFRt15*-TGFRs had little or no effect on the percentage of these cell populations.
  • Figure 229 are graphs showing that IL15SA (positive control) or TGFRt15*- TGFRs + IL15SA mediated an increase in the percentages of CD3 + CD8 + , CD3-NK1.1 + , and CD3 + CD45 + immune cells in the spleen, whereas treatment with TGFRt15*-TGFRs had little or no effect on the percentage of these cell populations.
  • Figure 230A shows gene expression of senescence marker p21, in kidney and liver tissues, post test article treatment.
  • Figure 230B shows gene expression of senescence marker PAI1, in kidney and liver tissues, post study treatment.
  • Figure 230C shows gene expression of senescence marker IL-1 ⁇ , in kidney and liver tissues, post study treatment.
  • Figure 230D shows gene expression of senescence marker IL-6, in kidney and liver tissues, post study treatment.
  • Figure 231A shows CD4 + , CD8 + , and Treg cell percentages and proliferation following treatment with the agents shown.
  • Figure 231B shows NK, CD19 + , and monocyte cell percentages and proliferation following treatment with the agents shown.
  • Figure 232A shows evaluation of gene expression of senescence markers p21 in lung tissue of mice following chemotherapy and treatment with the agents shown.
  • Figure 232B shows evaluation of gene expression of senescence marker CD26 in lung tissue of mice following chemotherapy and treatment with the agents shown.
  • Figure 232C shows evaluation of gene expression of senescence marker p21 in liver tissue of mice following chemotherapy and treatment with the agents shown.
  • Figures 233A-B are graphs showing TGFRt15-TGFRs treatment enhances the immune cell proliferation, expansion and activation in the peripheral blood of B16F10 tumor bearing mice.
  • Figure 234 are graphs showing TGFRt15-TGFRs treatment decreases levels of TGF ⁇ in the plasma of B16F10 tumor bearing mice.
  • Figure 235 are graphs showing TGFRt15-TGFRs treatment reduces levels of proinflammatory cytokines in the plasma of B16F10 tumor bearing mice.
  • Figure 236 shows TGFRt15-TGFRs treatment enhances NK and CD8 expansion in the spleen of B16F10 tumor bearing mice.
  • Figures 237A-B show TGFRt15-TGFRs treatment enhances glycolytic activity of splenocytes in B16F10 tumor bearing mice.
  • Figures 238A-B show TGFRt15-TGFRs treatment enhances mitochondrial respiration of splenocytes in B16F10 tumor bearing mice.
  • Figures 239A-B show TGFRt15-TGFRs treatment enhances NK and CD8 immune cell infiltration (TILs) into tumors of B16F10 tumor bearing mice.
  • Figure 240 shows histopathological analysis of tumors following TGFRt15- TGFRs treatment, wherein following TGFRt15-TGFRs+TA99 antibody treatment, tumors displayed less mitotic and necrotic activity.
  • FIG. 241 is a graph showing anti-PD-L1 antibody in combination with TGFRt15-TGFRs+TA99 antibody and chemotherapy in B16F10 melanoma mouse model.
  • Figure 242 is a graph showing that anti-tumor efficacy of TGFRt15-TGFRs in B16F10 melanoma mouse model is dependent on NK and CD8 T cells.
  • Figures 243A-B are graphs showing gene expression of senescence markers p21, IL-1 ⁇ and IL6 in liver and lung tissues of tumor bearing mice following chemotherapy.
  • Figure 244 is a graph showing induction of gene expression of senescence markers p21, IL6, H2AX, and NK cell ligands, Rae1e and ULBP1 by docetaxel treatment of B16F10 GFP cells.
  • Figure 245 shows tumor infiltrating lymphocytes/day after 4 days post treatment in tumor bearing mice.
  • Figures 246A-B show flow cytometry analysis on tumor cells indicating that mice which received immunotherapy treatment showed lower number of GFP positive senescent tumor cells post 4 days and 10 days of treatment as compared to the PBS control group ( Figure 246A), and tumor cells plated in 24 well plate evaluated by fluorescence microscopy (Figure 246B).
  • Figure 247 shows TGF ⁇ levels in kidney of mice after inducing kidney injury with cisplatin and treatment with TGFRt15-TGFRs.
  • Figures 248A-C show the toxicological effects of repeat dose subcutaneous administration of TGFRt15-TGFRs in C57BL/6 mice. Changes in body weights are shown through SD21 ( Figure 248A). Spleen weights (Figure 248B) and blood cells counts and differentials (Figure 248C) are indicated for mice at SD7 after one dose and SD21 after two doses of TGFRt15-TGFRs.
  • Figure 249 shows plasma levels of TGF- ⁇ isoforms in mice after in vivo treatment with PBS, TGFRt15-TGFRs (3 mg/kg) or TGFRt15*-TGFRs (3 mg/kg).
  • Figures 250A-B show the changes in rates of glycolytic capacity (ECAR) ( Figure 250A) and mitochondrial respiratory capacity (OCR) ( Figure 250B) in splenocytes of mice following in vivo treatment with PBS, TGFRt15-TGFRs, TGFRt15*-TGFRs or IL15SA.
  • ECAR glycolytic capacity
  • OCR mitochondrial respiratory capacity
  • Figures 251A-B show the changes in rates of glycolytic capacity (ECAR) ( Figure 251A) and mitochondrial respiratory capacity (OCR) ( Figure 251B) in mouse splenocytes following in vitro treatment with PBS, TGFRt15-TGFRs, or TGFRt15*-TGFRs.
  • Figures 252A-E show the changes in tumor growth and survival of B16F10 melanoma tumors in C57BL/6 mice following in vitro treatment with PBS, TGFRt15- TGFRs, or TGFRt15*-TGFRs.
  • Tumor volume (Figure 252A) and mouse survival (based on tumor volume ⁇ 4000 mm 3 ) ( Figure 252B) were assessed.
  • mice were intraperitoneally treated with anti-CD8, anti-NK, or anti-CD8 and anti-NK Abs for 1 week to deplete immune cells prior to injection with B16F10 melanoma tumor cells as in Figure 252A.
  • Tumor bearing mice were then treated with PBS or 20 mg/kg TGFRt15-TGFRs on day 1 and 4 post-tumor cell inoculation.
  • Tumor volume of animals Figure 252C
  • mouse survival Figure 252D
  • B16F10 tumor bearing mice were treated with PBS or 20 mg/kg of TGFRt15-TGFRs on day 1 and 7 post-tumor inoculation (Figure 252E).
  • Figures 253A-B show treatment effects on fasting plasma glucose (Figure 253A) and insulin (Figure 253B) levels in db/db mice receiving PBS (control) or TGFRt15- TGFRs.
  • Figure 254A shows the fold change in gene expression levels in pancreas of db/db mice receiving TGFRt15-TGFRs compared to PBS control.
  • Figures 254B-D show the average fold change in pancreatic expression levels for genes of the SASP, Aging and Beta cell indices, respectively, for db/db mice receiving TGFRt15-TGFRs compared to PBS control.
  • Figures 255A-B show multispectral imaging of pancreatic tissue sections from db/db mice treated with PBS (control) ( Figure 255A) or TGFRt15-TGFRs ( Figure 255B).
  • FIG. 255A A representative pancreatic islet is shown, insulin + islet beta cells as OPAL-520, insulin + p21 + beta cells as OPAL-570 (seen as white cells in gray-scale image) was reduced in TGRt15-TGFRs treated group (Figure 255B) compared to PBS treated group ( Figure 255A).
  • Figures 255C and 255D show levels of islet insulin + (Figure 255C) and islet insulin + p21 + ( Figure 255D) cells in pancreatic tissue sections from db/db mice treated with PBS (control) or TGFRt15-TGFRs.
  • Figures 256A-C show treatment effects on the percentage of blood immune cell subsets in db/db mice receiving PBS (control) or TGFRt15-TGFRs.
  • Figure 257 shows the percentage of Ki67 positive immune cells induced in the blood following subcutaneous treatment of Cynomolgus monkeys with TGFRt15-TGFRs compared to PBS (vehicle).
  • Figure 258 shows the extracellular acidification rate (ECAR) representing glycolytic function of splenocytes isolated from young (6-week-old) and aged (72-week- old) mice 4 days after in vivo treatment with PBS, TGFRt15-TGFRs (3 mg/kg) or TGFRt15*-TGFRs (3 mg/kg).
  • ECAR extracellular acidification rate
  • Figure 259 shows the oxygen consumption rate (OCR) representing mitochondrial respiration of splenocytes isolated from young (6-week-old) and aged (72-week-old) mice 4 days after in vivo treatment with PBS, TGFRt15-TGFRs (3 mg/kg) or TGFRt15*- TGFRs (3 mg/kg).
  • Figure 260 shows the percentages of immune cell subsets in the blood of young (6-week-old) and aged (72-week-old) mice 4 days after in vivo treatment with PBS, TGFRt15-TGFRs (3 mg/kg) or TGFRt15*-TGFRs (3 mg/kg).
  • Figure 261 shows the percentages of immune cell subsets in the spleen of young (6-week-old) and aged (72-week-old) mice 4 days after in vivo treatment with PBS, TGFRt15-TGFRs or TGFRt15*-TGFRs.
  • Figure 262 shows gene expression levels for IL1- ⁇ , IL1- ⁇ , IL-6, p21 and PAI-1 in liver of aged mice after one or two doses of TGFRt15-TGFRs treatment.
  • Figure 263 shows the inflammation score of liver tissues of aged mice after one or two doses of TGFRt15-TGFRs treatment.
  • Figure 264 shows expression levels of IL1- ⁇ , IL1- ⁇ , IL-6, IL-8, TGF- ⁇ , PAI-1, collagen and fibronectin protein in liver of aged mice after with one or two doses treatment of TGFRt15-TGFRs.
  • Figure 265 shows the levels of ⁇ -galactosidase in liver tissues of aged mice 4 days after in vivo treatment with PBS or TGFRt15-TGFRs.
  • Figure 266 shows the survival curves of 72-week-old C57BL/6 mice following subcutaneous treatment with PBS or one dose of TGFRt15-TGFRs (3 mg/kg).
  • Figure 267 shows protein levels of SASP factors in livers of B16F10 tumor- bearing mice following chemotherapy and TGFRt15-TGFRs + TA99 therapy.
  • Figures 268A-B show effects of CD8 + T cells (dpCD8) and NK cell (dpNK) antibody depletion on the levels of TIS B16F10-GFP cells ( Figure 268A) and NK and CD8 + T cells (Figure 268B) in the tumors of mice following chemotherapy and TGFRt15-TGFRs + TA99 therapy.
  • Figures 269A-E show the anti-tumor activity and mechanism of action of TGFRt15-TGFRs + TA99 in combination with immune checkpoint inhibitor in B16F10 tumor-bearing mice.
  • Figure 269A shows an exemplary schematic for treating B16F10 melanoma in a mouse model.
  • Figure 269B shows the change in tumor volume over time and at day 18 following combination treatments including TGFRt15-TGFRs+TA99+anti- PD-L1 antibody following doxetaxel as compared to PBS or chemotherapy treatment alone.
  • Figures 269C and 269D show treatment effects on the percentages of tumor infiltrating CD28 + CD8 + T cells and splenic IFN ⁇ + CD8 + T cells on day 18.
  • Figure 269E shows treatment effects on the levels (MFI) of NKG2D of tumor infiltrating CD8 + and CD8 + CD44 hi T cells on day 18.
  • Figures 270A-D show the changes in tumor growth and survival of SW1990 human pancreatic tumors in C57BL/6 scid mice following in vitro treatment with PBS, gemcitabine and nab-paclitaxel chemotherapy, TGFRt15-TGFRs, or TGFRt15- TGFRs+chemotherapy.
  • Figure 270A shows an exemplary schematic for treating SW1990 human pancreatic tumors in a xenograft mouse model.
  • Figure 270B and 270C show the change in tumor volume over time and at day 38, respectively, following combination treatments including TGFRt15-TGFRs + chemotherapy as compared to PBS or chemotherapy treatment alone.
  • Figure 270D shows treatment effects on survival of mice bearing SW1990 human pancreatic tumors.
  • DETAILED DESCRIPTION Provided herein are methods of killing or reducing the number of naturally- occurring and/or treatment-induced senescent cells in a subject that include administering to the subject a therapeutically effective amount of one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor. Also provided herein are methods of decreasing the accumulation of naturally-occurring and/or treatment-induced senescent cells in a subject that include administering to the subject a therapeutically effective amount of one or more agent(s) that result(s) in a decrease in the activation of a TGF- ⁇ receptor.
  • NK natural killer
  • methods of treating an aging-related disease or condition in a subject in need thereof that include administering to a subject identified as having an aging-related disease or condition a therapeutically effective amount of one or more natural killer (NK) cell activating agent(s) and/or a therapeutically effective number of activated NK cells.
  • methods of killing or reducing the number of senescent cells in a subject in need thereof that include administering to the subject a therapeutically effective amount of one or more NK cell activating agent(s) and/or a therapeutically effective number of activated NK cells.
  • NK natural killer
  • Activated NK Cells Some embodiments of any of the methods described herein can include administering to a subject (e.g., any of the exemplary subjects described herein) a therapeutically effective number of activated NK cells (e.g., human activated NK cells).
  • An activated NK cell is an NK cell (e.g., a human NK cell) that has increased expression levels of two or more (e.g., three, four, five, or six) of CD25, CD69, MTOR-C1, SREBP1, IFN- ⁇ , and a granzyme (e.g., granzyme B), e.g., as compared to a resting NK cell (e.g., a human resting NK cell).
  • a granzyme e.g., granzyme B
  • an activated NK cell can have at least a 10% increase (e.g., at least a 15% increase, at least a 20% increase, at least a 25% increase, at least a 30% increase, at least a 35% increase, at least a 40% increase, at least a 45% increase, at least a 50% increase, at least a 55% increase, at least a 60% increase, at least a 65% increase, at least a 70% increase, at least a 75% increase, at least a 80% increase, at least a 85% increase, at least a 90% increase, at least a 95% increase, at least a 100% increase, at least a 120% increase, at least a 140% increase, at least a 160% increase, at least a 180% increase, at least a 200% increase, at least a 220% increase, at least a 240% increase, at least a 260% increase, at least a 280% increase, or at least a 300% increase) in the expression levels of two of more (e.g., three, four,
  • an activated NK cell can optionally further have at least a 10% increase (e.g., at least a 15% increase, at least a 20% increase, at least a 25% increase, at least a 30% increase, at least a 35% increase, at least a 40% increase, at least a 45% increase, at least a 50% increase, at least a 55% increase, at least a 60% increase, at least a 65% increase, at least a 70% increase, at least a 75% increase, at least a 80% increase, at least a 85% increase, at least a 90% increase, at least a 95% increase, at least a 100% increase, at least a 120% increase, at least a 140% increase, at least a 160% increase, at least a 180% increase, at least a 200% increase, at least a 220% increase, at least a 240% increase, at least a 260% increase, at least a 280% increase, or at least a 300% increase) in the expression levels of two of more (e.g., at least
  • an activated NK cell (e.g., a human activated NK cell) can have about a 10% increase to about a 500% increase, about a 10% increase to about a 450% increase, about a 10% increase to about a 400% increase, about a 10% increase to about a 350% increase, about a 10% increase to about a 300% increase, about a 10% increase to about a 280% increase, about a 10% increase to about a 260% increase, about a 10% increase to about a 240% increase, about a 10% increase to about a 220% increase, about a 10% increase to about a 200% increase, about a 10% increase to about a 180% increase, about a 10% increase to about a 160% increase, about a 10% increase to about a 140% increase, about a 10% increase to about a 120% increase, about a 10% increase to about a 100% increase, about a 10% increase to about a 80% increase, about a 10% increase to about a 60% increase, about a 10% increase to about a 40%
  • an activated NK cell can further have about a 10% increase to about a 500% increase (e.g., or any of the subranges of this range described herein) in the expression levels of two of more (e.g., 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, or 29) of CD25, CD59, CD352, NKp80, DNAM-1, 2B4, NKp30, NKp44, NKp46, NKG2D, CD16, KIR2DS1, KIR2Ds2/3, KIR2DL4, KIR2DS4, KIR2DS5, KIR3DS1, NKG2C, CCR7, CXCR3, L- Selectin, CXCR1, CXCR2, CX3CR1, ChemR23, CXCR4, CCR5, S1P5, c-Kit, mTORC1, e.g., as compared to a resting NK cell (e.g., a human activated NK cell
  • Non-limiting examples of assays that can be used to determine the expression level of CD25, CD69, CD59, CD352, NKp80, DNAM-1, 2B4, NKp30, NKp44, NKp46, NKG2D, CD16, KIR2DS1, KIR2Ds2/3, KIR2DL4, KIR2DS4, KIR2DS5, KIR3DS1, NKG2C, CCR7, CXCR3, L-Selectin, CXCR1, CXCR2, CX3CR1, ChemR23, CXCR4, CCR5, S1P5, c-Kit, mTORC1, MYC, SREBP1, IFN- ⁇ , and a granzyme (e.g., granzyme B) include, e.g., immunoblotting, fluorescence-assisted cell sorting, enzyme-linked immunosorbent assays, and RT-PCR.
  • Non-limiting examples of commercial ELISA assays that can be used to determine the expression level of CD25 are available from Diaclone, Covalab Biotechnology, and Caltag Medsystems.
  • the protein and cDNA sequences for mature human CD25 are shown below.
  • Mature Human CD25 Protein (SEQ ID NO: 1) elcdddppe iphatfkama ykegtmlnce ckrgfrriks gslymlctgn sshsswdnqc qctssatrnt tkqvtpqpee qkerkttemq spmqpvdqas lpghcreppp weneateriy hfvvgqmvyy qcvqgyralh rgpaesvckm thgktrwtqp qlictgemet sqfpgeekpq aspegrpese tsclvtttdf
  • Mature Human CD16a Protein (SEQ ID NO: 23) maegtlwqil cvssdaqpqt fegvkgadpp tlppgsflpg pvlwwgslar lqteksdevs rkgnwwvtem gggagerlft ssclvglvpl glrislvtcp lqcgimwqll lptallllvs agmrtedlpk avvflepqwy rvlekdsvtl kcqgaysped nstqwfhnes lissqassyf idaatvddsg eyrcqtnlst lsdpvqlevh igwlllqapr wvfkeedpih lrchswknta lhkvtylqng kgrkyfh
  • Mature Human mTOR Protein (SEQ ID NO: 65) mlgtgpaaat taattssnvs vlqqfasglk srneetraka akelqhyvtm elremsqees trfydqlnhh ifelvsssda nerkggilai asligveggn atrigrfany lrnllpsndp vvmemaskai grlamagdtf taeyvefevk ralewlgadr negrrhaavl vlrelaisvp tfffqqvqpf fdnifvavwd pkqairegav aalraclilt tqrepkemqk pqwyrhtfee aekgfdetla kekgmnrddr ihgallilne lvrissmege
  • Mature Human SREBP1 Protein (SEQ ID NO: 67) MDEPPFSEAALEQALGEPCDLDAALLTDIEDMLQLINNQDSDFPGLFDPPYAGSG AGGTDPASPDTSSPGSLSPPPATLSSSLEAFLSGPQAAPSPLSPPQPAPTPLKMYPS MPAFSPGPGIKEESVPLSILQTPTPQPLPGALLPQSFPAPAPPQFSSTPVLGYPSPPG GFSTGSPPGNTQQPLPGLPLASPPGVPPVSLHTQVQSVVPQQLLTVTAAPTAAPV TTTVTSQIQQVPVLLQPHFIKADSLLLTAMKTDGATVKAAGLSPLVSGTTVQTGP LPTLVSGGTILATVPLVVDAEKLPINRLAAGSKAPASAQSRGEKRTAHNAIEKRY RSSINDKIIELKDLVVGTEAKLNKSAVLRKAIDYIRFLQHSNQKLKQENLSLRTAV HKS
  • Mature Human IFN- ⁇ (SEQ ID NO: 69) qdpyvke aenlkkyfna ghsdvadngt lflgilknwk eesdrkimqs qivsfyfklf knfkddqsiq ksvetikedmnvkffnsnkk krddfekltn ysvtdlnvqr kaiheliqvm aelspaaktg krkrsqmlfrg Human IFN- ⁇ cDNA (SEQ ID NO: 70) caggac ccatatgtaa aagaagcaga aaaccttaag aaatatttta atgcaggtca ttcagatgta gcggataatggaactctttt cttaggcatt t
  • Mature Human Granzyme B (SEQ ID NO: 71) iiggheakph srpymaylmi wdqkslkrcg gflirddfvl taahcwgssi nvtlgahnik eqeptqqfip vkrpiphpay npknfsndim llqlerkakr travqplrlp snkaqvkpgq tcsvagwgqt aplgkhshtl qevkmtvqed rkcesdlrhy ydstielcvg dpeikktsfk gdsggplvcn kvaqgivsyg rnngmpprac tkvssfvhwi kktmkry Human Granzyme B
  • Human Myc Protein (SEQ ID NO: 329) mdffrvvenq qppatmplnvsftnrnydld ydsvqpyfyc deeenfyqqq qqselqppap sediwkkfel lptpplspsrrsglcspsyv avtpfslrgd ndggggsfst adqlemvtel lggdmvnqsf icdpddetfikniiiqdcmw sgfsaaaklv seklasyqaa rkdsgspnpa rghsvcstss lylqdlsaaasecidpsvvf pyplndsssp kscasqdssa fspsdslls stesss
  • activated NK cells e.g., human activated NK cells
  • activated NK cells can show about a 10% increase to about a 500% increase (or any of the subranges of this range described herein) ability to kill senescent cells (e.g., any of the senescent cells described herein) in a subject (e.g., any of the subjects described herein) or in vivo as compared to resting NK cells (e.g., human resting NK cells).
  • activated NK cells can show increased (e.g., at least a 10% increase, at least a 20% increase, at least a 30% increase, at least a 40% increase, at least a 50% increase, at least a 60% increase, at least a 70% increase, at least 80% increase, at least a 90% increase, at least a 100% increase, at least a 120% increase, at least a 140% increase, at least a 160% increase, at least a 180% increase, at least a 200% increase, at least a 220% increase, at least a 240% increase, at least a 260% increase, at least a 280% increase, or at least a 300% increase) cytotoxic activity in a contact-cytotoxicity assay in the presence of an antibody that binds specifically to an antigen present on a senescent or target cell, e.g., as compared to a resting NK cell (e.g., human resting NK cells).
  • increased e.g., at least a 10% increase, at least a
  • activated NK cells e.g., human activated NK cells
  • activated NK cells can show increased (e.g., about a 10% increase to about a 500% increase, or any of the subranges of this range described herein) cytotoxic activity in a contact-cytotoxicity assay in the presence of an antibody that binds specifically to an antigen present on a senescent or target cell, e.g., as compared to a resting NK cell (e.g., human resting NK cells).
  • an activated NK cell can be produced by a method that includes obtaining a resting NK cell; and contacting the resting NK cell in vitro in a liquid culture medium including one or more NK cell activating agent(s), where the contacting results in the generation of the activated NK cells that are subsequently administered to the subject.
  • the resting NK cell is an autologous NK cell obtained from the subject.
  • the resting NK cell is an autologous NK cell obtained from the subject.
  • the resting NK cell is an haploidentical resting NK cells.
  • the resting NK cell is an allogeneic resting NK cell.
  • the resting NK cell is an artificial NK cell. In some examples of any of these methods, the resting NK cell is a genetically-engineered NK cell carrying a chimeric antigen receptor or recombinant T cell receptor.
  • the liquid culture medium is a serum-free liquid culture medium. In some embodiments of any of the methods described herein, the liquid culture medium is a chemically-defined liquid culture medium. Some examples of these methods further include isolating the activated NK cells (and optionally further administering a therapeutically effective amount of the activated NK cells to a subject, e.g., any of the subjects described herein).
  • the contacting step is performed for a period of about 2 hours to about 20 days (e.g., about 2 hours to about 18 days, about 2 hours to about 16 days, about 2 hours to about 14 days, about 2 hours to about 12 days, about 2 hours to about 10 days, about 2 hours to about 8 days, about 2 hours to about 7 days, about 2 hours to about 6 days, about 2 hours to about 5 days, about 2 hours to about 4 days, about 2 hours to about 3 days, about 2 hours to about 2 days, about 2 hours to about 1 day, about 6 hours to about 18 days, about 6 hours to about 16 days, about 6 hours to about 14 days, about 6 hours to about 12 days, about 6 hours to about 10 days, about 6 hours to about 8 days, about 6 hours to about 7 days, about 6 hours to about 6 days, about 6 hours to about 5 days, about 6 hours to about 4 days, about 6 hours to about 3 days, about 6 hours to about 2 days, about 6 hours to about 1 day, about 12 hours to about 18 days, about 6 hours to about 16 days, about 6 hours to about 14
  • an NK cell activating agent can be a protein.
  • an NK cell activating agent can be a single-chain chimeric polypeptide (e.g. any of the single-chain chimeric polypeptides described herein), a multi-chain chimeric polypeptide (e.g. any of the multi-chain chimeric polypeptides described herein, e.g., the exemplary type A and type B multi-chain chimeric polypeptides described herein), an antibody, a recombinant cytokine or an interleukin (e.g.
  • the NK cell activating agent can be a small molecule (e.g., a glycogen synthase kinase-3 (GSK3) inhibitor, e.g., CHIR99021 as described in Cichocki et al., Cancer Res.77:5664-5675, 2017) or an aptamer.
  • GSK3 glycogen synthase kinase-3
  • At least one of the one or more NK cell activating agent(s) results in activation of one or more (e.g., two, three, four, five, six, seven, or eight) of: a receptor for IL-2, a receptor for IL-7, a receptor for IL-12, a receptor for IL-15, a receptor for IL- 18, a receptor for IL-21, a receptor for IL-33, CD16, CD69, CD25, CD59, CD352, NKp80, DNAM-1, 2B4, NKp30, NKp44, NKp46, NKG2D, KIR2DS1, KIR2Ds2/3, KIR2DL4, KIR2DS4, KIR2DS5, and KIR3DS1 (e.g., in an immune cell, e.g., a human immune cell, e.g., a human NK cell) as compared to the level of activation in the absence of
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL-2 is a soluble IL-2 or an agonistic antibody that binds specifically to an IL-2 receptor. In some embodiments, the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL-7 is a soluble IL-7 or an agonistic antibody that binds specifically to an IL-7 receptor. In some embodiments, the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL-12 is a soluble IL-12 or an agonistic antibody that binds specifically to an IL-12 receptor.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL-15 is a soluble IL-15 or an agonistic antibody that binds specifically to an IL-15 receptor.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL-21 is a soluble IL-21 or an agonistic antibody that binds specifically to an IL-21 receptor.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of a receptor for IL-33 is a soluble IL-33 or an agonistic antibody that binds specifically to an IL-33 receptor.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of CD16 is an agonistic antibody that binds specifically to CD16.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of CD69 is an agonistic antibody that binds specifically to CD69.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of CD25, CD59 is an agonistic antibody that binds specifically to CD25, CD59.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of CD352 is an agonistic antibody that binds specifically to CD352.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of NKp80 is an agonistic antibody that binds specifically to NKp80.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of DNAM-1 is an agonistic antibody that binds specifically to DNAM-1.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of 2B4 is an agonistic antibody that binds specifically to 2B4.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of NKp30 is an agonistic antibody that binds specifically to NKp30.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of NKp44 is an agonistic antibody that binds specifically to NKp44.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of NKp46 is an agonistic antibody that binds specifically to NKp46.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of NKG2D is an agonistic antibody that binds specifically to NKG2D.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of KIR2DS1 is an agonistic antibody that binds specifically to KIT2DS1.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of KIR2DS2/3 is an agonistic antibody that binds specifically to KIT2DS2/3.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of KIR2DL4 is an agonistic antibody that binds specifically to KIT2DL4. In some embodiments, the at least one of the one or more NK cell activating agent(s) that results in activation of KIR2DS4 is an agonistic antibody that binds specifically to KIT2DS4. In some embodiments, the at least one of the one or more NK cell activating agent(s) that results in activation of KIR2DS5 is an agonistic antibody that binds specifically to KIT2DS5.
  • the at least one of the one or more NK cell activating agent(s) that results in activation of KIR3DS1 is an agonistic antibody that binds specifically to KIT3DS1.
  • at least one (e.g., two, three, four, or five) of the one or more NK cell activating agent(s) results in a decrease in the activation of one or more of: PD-1, a TGF- ⁇ receptor, TIGIT, CD1, TIM-3, Siglec-7, IRP60, Tactile, IL1R8, NKG2A/KLRD1, KIR2DL1, KIR2DL2/3, KIR2DL5, KIR3DL1, KIR3DL2, ILT2/LIR-1, and LAG-2 (e.g., in an immune cell, e.g., a human immune cell, e.g., a human NK cell) as compared to the level of activation in the absence of
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of a TGF- ⁇ receptor is a soluble TGF- ⁇ receptor, an antibody that binds specifically to TGF- ⁇ , or an antagonistic antibody that binds specifically to a TGF- ⁇ receptor.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of TIGIT is an antagonistic antibody that binds specifically to TIGIT, a soluble TIGIT, or an antibody that binds specifically to a ligand of TIGIT.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of CD1 is an antagonistic antibody that binds specifically to CD1, a soluble CD1, or an antibody that binds specifically to a ligand of CD1.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of TIM-3 is an antagonistic antibody that binds specifically to TIM-3, a soluble TIM-3, or an antibody that binds specifically to a ligand of TIM-3.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of Siglec-7 is an antagonistic antibody that binds specifically to Siglec-7, a soluble Siglec-7, or an antibody that binds specifically to a ligand of Siglec-7.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of IRP-60 is an antagonistic antibody that binds specifically to IRP-60, a soluble IRP-60, or an antibody that binds specifically to a ligand of IRP-60.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of Tactile is an antagonistic antibody that binds specifically to Tactile, a soluble Tactile, or an antibody that binds specifically to a ligand of Tactile.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of IL1R8 is an antagonistic antibody that binds specifically to IL1R8, a soluble IL1R8, or an antibody that binds specifically to a ligand of IL1R8.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of NKG2A/KLRD1 is an antagonistic antibody that binds specifically to NKG2A/KLRD1, a soluble NKG2A/KLRD1, or an antibody that binds specifically to a ligand of NKG2A/KLRD1.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of KIR2DL1 is an antagonistic antibody that binds specifically to KIR2DL1, a soluble KIR2DL1, or an antibody that binds specifically to a ligand of KIR2DL1.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of KIR2DL2/3 is an antagonistic antibody that binds specifically to KIR2DL2/3, a soluble KIR2DL2/3, or an antibody that binds specifically to a ligand of KIR2DL2/3.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of KIR2DL5 is an antagonistic antibody that binds specifically to KIR2DL5, a soluble KIR2DL5, or an antibody that binds specifically to a ligand of KIR2DL5.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of KIR3DL1 is an antagonistic antibody that binds specifically to KIR3DL1, a soluble KIR3DL1, or an antibody that binds specifically to a ligand of KIR3DL1.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of KIR3DL2 is an antagonistic antibody that binds specifically to KIR3DL2, a soluble KIR3DL2, or an antibody that binds specifically to a ligand of KIR3DL2.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of ILT2/LIR-1 is an antagonistic antibody that binds specifically to ILT2/LIR-1, a soluble ILT2/LIR-1, or an antibody that binds specifically to a ligand of ILT2/LIR-1.
  • the at least one of the one or more NK cell activating agent(s) that results in a decrease in the activation of LAG2 is an antagonistic antibody that binds specifically to LAG2, a soluble LAG2, or an antibody that binds specifically to a ligand of LAG2.
  • Non-limiting examples of NK cell activating agents are described below and can be used in any combination.
  • an NK cell activating agents can be a soluble PD-1, a soluble PD-L1, a soluble TIGIT, a soluble CD1, or a soluble TIM-3.
  • soluble PD-1, PD-L1, TIGIT, CD1, and TIM-3 are provided below.
  • Human Soluble PD-1 (SEQ ID NO: 73) pgwfldspdr pwnpptfspa llvvtegdna tftcsfsnts esfvlnwyrm spsnqtdkla afpedrsqpg qdcrfrvtql pngrdfhmsv vrarrndsgt ylcgaislap kaqikeslra elrvterrae vptahpspsp rpagqfqtlv vgvvggllgs lvllvwvlav icsraargti garrtgqplk edpsavpvfs vdygeldfqw rektpeppvp cvpeqteyat ivfpsgmgts sparrgsadg prsaqplrpe
  • a soluble PD-L1 protein can include a sequence that is at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 74.
  • a soluble TIGIT protein can include a sequence that is at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 75.
  • a soluble CD1A protein can include a sequence that is at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 76.
  • a soluble TIM3 protein can include a sequence that is at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 77.
  • NK activating agent can be: an agonistic antibody that binds specifically to an IL-2 receptor (see, e.g., those described in Gaulton et al., Clinical Immunology and Immunopathology 36(1):18-29, 1985), an agonistic antibody that binds specifically to an IL-7 receptor, an agonistic antibody that binds specifically to IL-12 receptor (see, e.g., those described in Rogge et al., J. Immunol.162(7): 3926-3932, 1999), an agonistic antibody that binds specifically to an IL-15 receptor, an agonistic antibody that binds specifically to an IL-21 receptor (see, e.g., those described in U.S.
  • Patent Application Publication No.2006/159655 an agonistic antibody that binds specifically to an IL-33 receptor (see, e.g., those described in U.S. Patent Application Publication No. 2007/160579), an antagonistic antibody that binds specifically to PD-1 (see, e.g., those described in U.S. Patent No.7,521,051), an antibody that binds specifically to PD-L1 (see, e.g., those described in U.S.
  • Patent No.8,217,149 an antibody that binds specifically to TGF- ⁇ , an antagonistic antibody that binds specifically to TGF- ⁇ receptor (see, e.g., those described in European Patent Application Publication No.1245676 A1), an antagonistic antibody that binds specifically to TIGIT (see, e.g., those described in WO 2017/053748), an antibody that binds specifically to a ligand of TIGIT (see, e.g., those described in WO 2011/127324), an antagonistic antibody that binds specifically to CD1 (see, e.g., those described in Szalay et al., J.
  • Immunol.162(12):6955-6958, 1999) an antibody that binds specifically to a ligand of CD1 (see, e.g., those described in Kain et al., Immunity 41(4):543-554, 2014), an antagonistic antibody that binds specifically to TIM-3 (see, e.g., those described in U.S. Patent Application Publication No. 2015/218274), an antibody that binds specifically to a ligand of TIM-3 (see, e.g., those described in U.S.
  • Patent Application Publication No.2017/283499 an agonistic antibody that binds specifically to CD69 (see, e.g., those described in Moretta et al., Journal of Experimental Medicine 174:1393, 1991), an agonistic antibody that binds specifically to CD25, CD59, an agonistic antibody that binds specifically to CD352 (see, e.g., those described in Yigit et al., Oncotarget 7:26346-26360, 2016), an agonistic antibody that binds specifically to NKp80 (see, e.g., those described in Peipp et al., Oncotarget 6:32075-32088, 2015), an agonistic antibody that binds specifically to DNAM-1, an agonistic antibody that binds specifically to 2B4 (see, e.g., those described in Sandusky et al., European J.
  • Immunol.36:3268-3276, 2006 an agonistic antibody that binds specifically to NKp30 (see, e.g., those described in Kellner et al., OncoImmunology 5:1-12, 2016), an agonistic antibody that binds specifically to NKp44, an agonistic antibody that binds specifically to NKp46 (see, e.g., those described in Xiong et al., J. Clin.
  • an agonistic antibody that binds specifically to NKG2D see, e.g., those described in Kellner et al., OncoImmunology 5:1- 12, 2016
  • an agonistic antibody that binds specifically to KIR2DS1 see, e.g., those described in Xiong et al., J. Clin. Invest.123:4264-4272, 2013
  • an agonistic antibody that binds specifically to KIR2Ds2/3 see, e.g., those described in Borgerding et al., Exp.
  • an agonistic antibody that binds specifically to KIR2DL4 see, e.g., those described in Miah et al., J. Immunol.180:2922-32, 2008
  • an agonistic antibody that binds specifically to KIR2DS4 see, e.g., those described in Czaja et al., Genes and Immunity 15:33-37, 2014
  • an agonistic antibody that binds specifically to KIR2DS5 see, e.g., those described in Czaja et al., Genes and Immunity 15:33-37, 2014
  • an agonistic antibody that binds specifically to KIR3DS1 see, e.g., those described in Czaja et al., Genes and Immunity 15:33-37, 2014
  • an antagonistic antibody that binds specifically to Siglec-7 see, e.g., those described in Hudak et al., Nature Chemical Biology 10:69-
  • a recombinant antibody that is an NK cell activating agent can be any of exemplary types of antibodies (e.g., a human or humanized antibody) or any of the exemplary antibody fragments described herein.
  • a recombinant antibody that is an NK cell activating agent can include, e.g., any of the antigen-binding domains described herein.
  • Recombinant Interleukins or Cytokines can be, e.g., a soluble IL-2, a soluble IL- 7, a soluble IL-12, a soluble IL-15, a soluble IL-21, and a soluble IL-33.
  • Non-limiting examples of soluble IL-12, IL-15, IL-21, and IL-33. are provided below.
  • Human Soluble IL-2 (SEQ ID NO: 78) aptssstkkt qlqlehllld lqmilnginn yknpkltrml tfkfympkka telkhlqcle eelkpleevl nlaqsknfhl rprdlisnin vivlelkgse ttfmceyade tativeflnr witfcqsiis tlt Human Soluble IL-7 (SEQ ID NO: 79) dcdiegkdgkqyesv lmvsidqlld smkeigsncl nnefnffkrh icdankegmf lfraarklrq flkmnstgdf dlhllkv
  • a soluble IL-7 protein can include a sequence that is at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 79.
  • a soluble IL-2 protein includes a sequence that is at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 80 and a sequence that is at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 81.
  • a soluble IL-15 protein can include a sequence that is at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 82.
  • a soluble IL-21 protein can include a sequence that is at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 83.
  • a soluble IL-33 protein can include a sequence that is at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 84.
  • Soluble Cytokine or Interleukin Receptors In some examples of any of the soluble cytokine or interleukin receptors described herein, the soluble cytokine or interleukin receptors can be a soluble TGF- ⁇ receptor.
  • the soluble TGF- ⁇ receptor is a soluble TGF- ⁇ receptor I (TGF- ⁇ RI) (see, e.g., those described in Docagne et al., Journal of Biological Chemistry 276(49):46243-46250, 2001), a soluble TGF- ⁇ receptor II (TGF- ⁇ RII) (see, e.g., those described in Yung et al., Am. J. Resp. Crit. Care Med.194(9):1140-1151, 2016), a soluble TGF- ⁇ RIII (see, e.g., those described in Heng et al., Placenta 57:320, 2017).
  • TGF- ⁇ RI soluble TGF- ⁇ receptor I
  • TGF- ⁇ RII soluble TGF- ⁇ receptor II
  • TGF- ⁇ RIII see, e.g., those described in Heng et al., Placenta 57:320, 2017.
  • the soluble TGF- ⁇ receptor is a receptor “trap” for TGF- ⁇ (see, e.g., those described in Zwaagstra et al., Mol. Cancer Ther.11(7):1477-1487, 2012, and those described in De Crescenzo et al. Transforming Growth Factor- ⁇ in Cancer Therapy, Volume II, pp 671-684). Additional examples of soluble cytokine or soluble interleukin receptors are known in the art.
  • NK cell activating agents are single-chain chimeric polypeptides that include: (i) a first target-binding domain (e.g., any of the target-binding domains described herein or known in the art), (ii) a soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein or known in the art), and (iii) as second target-binding domain (e.g., any of the target-binding domains described herein or known in the art).
  • a first target-binding domain e.g., any of the target-binding domains described herein or known in the art
  • a soluble tissue factor domain e.g., any of the exemplary soluble tissue factor domains described herein or known in the art
  • second target-binding domain e.g., any of the target-binding domains described herein or known in the art
  • the single-chain chimeric polypeptide can have a total length of about 50 amino acids to about 3000 amino acids, about 50 amino acids to about 2500 amino acids, about 50 amino acids to about 2000 amino acids, about 50 amino acids to about 1500 amino acids, about 50 amino acids to about 1000 amino acids, about 50 amino acids to about 950 amino acids, about 50 amino acids to about 900 amino acids, about 50 amino acids to about 850 amino acids, about 50 amino acids to about 800 amino acids, about 50 amino acids to about 750 amino acids, about 50 amino acids to about 700 amino acids, about 50 amino acids to about 650 amino acids, about 50 amino acids to about 600 amino acids, about 50 amino acids to about 550 amino acids, about 50 amino acids to about 500 amino acids, about 50 amino acids to about 480 amino acids, about 50 amino acids to about 460 amino acids, about 50 amino acids to about 440 amino acids, about 50 amino acids to about 420 amino acids, about 50 amino acids to about 400 amino acids, about 50 amino acids to about 50 amino acids to about 50 amino acids to about 3000 amino acids, about 50 amino acids
  • the first target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the soluble tissue factor domain e.g., any of the exemplary soluble tissue factor domains described herein
  • the single-chain chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the first target- binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) and the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein).
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the soluble tissue factor domain e.g., any of the exemplary soluble tissue factor domains described herein
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the single-chain chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein) and the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art).
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the first target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the single-chain chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) and the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art).
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the soluble tissue factor domain e.g., any of the exemplary soluble tissue factor domains described herein
  • the single-chain chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) and the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein or known in the art).
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • a single-chain chimeric polypeptide can include a sequence that is at least 70% identical (e.g., at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical) to QIVLTQSPAIMSASPGEKVTMTCSASSSVSYMNWYQQKSGTSPKRWIYDTSKLA SGVPAHFRGSGSGTSYSLTISGMEAEDAATYYCQQWSSNPFTFGSGTKLEINRGG GGSGGGGSGGGGSQVQLQQSGAELARPGASVKMSCKASGYTFTRYTMHWVKQ RPGQGLEWIGYINPSRGYTNYNQKFKDKATLTTDKSSSTAYMQLSSLTSEDSAV YYCARYYDDHYCLDYWGQGTTLTVSSSGTTNTVAAYNLTWKSTNFKTILEWEP KPVNQVYTVQISTKSGDWKSKCFYTT
  • a single-chain chimeric polypeptide is encoded by a nucleic acid that includes a sequence that is at least 70% identical (e.g., at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical) to CAGATCGTGCTGACCCAAAGCCCCGCCATCATGAGCGCTAGCCCCGGTGAGA AGGTGACCATGACATGCTCCGCTTCCAGCTCCGTGTCCTACATGAACTGGTAT CAGCAGAAAAGCGGAACCAGCCCCAAAAGGTGGATCTACGACACCAGCAAG CTGGCCTCCGGAGTGCCCGCTCATTTCCGGGGCTCTGGATCCGGCACCAGCTA CTCTTTAACCATTTCCGGCATGGAAGCTGAAGACGCTGCCACCTACTATTGCC AGCAATGGAGCAGCAACCCCTTCACATTCGGATCTGGCACCAAGCTCGAAAT CAATCGTGGAGGAGGTGGCAGCGGCGGCGGTGGAT CAATCGT
  • a single-chain chimeric polypeptide can include a sequence that is at least 70% identical (e.g., at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical) to MKWVTFISLLFLFSSAYSQIVLTQSPAIMSASPGEKVTMTCSASSSVSYMNWYQQ KSGTSPKRWIYDTSKLASGVPAHFRGSGSGTSYSLTISGMEAEDAATYYCQQWS SNPFTFGSGTKLEINRGGGGSGGGGSGGGGSQVQLQQSGAELARPGASVKMSCK ASGYTFTRYTMHWVKQRPGQGLEWIGYINPSRGYTNYNQKFKDKATLTTDKSS STAYMQLSSLTSEDSAVYYCARYYDDHYCLDYWGQGTTLTVSSSGTTNTVAAY NLTWKSTNFKTILEWEPKPVNQVYTVQISTK
  • a single-chain chimeric polypeptide is encoded by a nucleic acid that includes a sequence that is at least 70% identical (e.g., at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical) to ATGAAGTGGGTGACCTTCATCAGCTTATTATTTTTATTCAGCTCCGCCTATTCC CAGATCGTGCTGACCCAAAGCCCCGCCATCATGAGCGCTAGCCCCGGTGAGA AGGTGACCATGACATGCTCCGCTTCCAGCTCCGTGTCCTACATGAACTGGTAT CAGCAGAAAAGCGGAACCAGCCCCAAAAGGTGGATCTACGACACCAGCAAG CTGGCCTCCGGAGTGCCCGCTCATTTCCGGGGCTCTGGATCCGGCACCAGCTA CTCTTTAACCATTTCCGGCATGGAAGCTGAAGACGCTGCCACCTACTATTGCC AGCAATGGAGCAGCAACCCCTTCACATTCGGATCTGGC
  • a single-chain chimeric polypeptide can include a sequence that is at least 70% identical (e.g., at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical) to VQLQQSGPELVKPGASVKMSCKASGYTFTSYVIQWVKQKPGQGLEWIGSINPYN DYTKYNEKFKGKATLTSDKSSITAYMEFSSLTSEDSALYYCARWGDGNYWGRG TTLTVSSGGGGSGGGGSGGGGSDIEMTQSPAIMSASLGERVTMTCTASSSVSSSY FHWYQQKPGSSPKLCIYSTSNLASGVPPRFSGSGSTSYSLTISSMEAEDAATYFCH QYHRSPTFGGGTKLETKRSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTV QISTKSGDWKSKCFYTTDTECDLTDEIVKDVKQTYLAR
  • a single-chain chimeric polypeptide is encoded by a nucleic acid that includes a sequence that is at least 70% identical (e.g., at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical) to GTGCAGCTGCAGCAGTCCGGACCCGAACTGGTCAAGCCCGGTGCCTCCGTGA AAATGTCTTGTAAGGCTTCTGGCTACACCTTTACCTCCTACGTCATCCAATGG GTGAAGCAGAAGCCCGGTCAAGGTCTCGAGTGGATCGGCAGCATCAATCCCT ACAACGATTACACCAAGTATAACGAAAAGTTTAAGGGCAAGGCCACTCTGAC AAGCGACAAGCTCCATTACCGCCTACATGGAGTTTTCCTCTTTAACTTCTG AGGACTCCGCTTTATACTATTGCGCTCGTTGGGGCGATGGCAATTATTGGGGC CGGGGAACTACTTTAACAGTGAGCTCCGGCGGCGG
  • a single-chain chimeric polypeptide can include a sequence that is at least 70% identical (e.g., at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical) to MKWVTFISLLFLFSSAYSVQLQQSGPELVKPGASVKMSCKASGYTFTSYVIQWV KQKPGQGLEWIGSINPYNDYTKYNEKFKGKATLTSDKSSITAYMEFSSLTSEDSA LYYCARWGDGNYWGRGTTLTVSSGGGGSGGGGSGGGGSDIEMTQSPAIMSASL GERVTMTCTASSSVSSSYFHWYQQKPGSSPKLCIYSTSNLASGVPPRFSGSGSTSY SLTISSMEAEDAATYFCHQYHRSPTFGGGTKLETKRSGTTNTVAAYNLTWKSTN FKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTD
  • a single-chain chimeric polypeptide is encoded by a nucleic acid that includes a sequence that is at least 70% identical (e.g., at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical) to ATGAAATGGGTCACCTTCATCTCTTTACTGTTTTTATTTAGCAGCGCCTACAG CGTGCAGCTGCAGCAGTCCGGACCCGAACTGGTCAAGCCCGGTGCCTCCGTG AAAATGTCTTGTAAGGCTTCTGGCTACACCTTTACCTCCTACGTCATCCAATG GGTGAAGCAGAAGCCCGGTCAAGGTCTCGAGTGGATCGGCAGCATCAATCCC TACAACGATTACACCAAGTATAACGAAAAGTTTAAGGGCAAGGCCACTCTGA CAAGCGACAAGCTCCATTACCGCCTACATGGAGTTTTCCTCTTTAACTTCT GAGGACTCCGCTTTATACTATTGCTCGTTGGGGCGAT
  • any of the single-chain chimeric polypeptides described herein can further include one or more (e.g., two, three, four, five, six, seven, eight, nine, or ten) additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) at its N- and/or C-terminus.
  • the single-chain chimeric polypeptides can include one or more (e.g., two, three, four, five, six, seven, eight, nine, or ten) additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) at its N-terminus.
  • one of the one or more additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) at the N-terminus of the single-chain chimeric polypeptide can directly abut the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), or the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein).
  • the first target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the soluble tissue factor domain e.g., any of the exemplary soluble tissue factor domains described herein.
  • the single-chain chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between one of the at least one additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) at the N-terminus of the single-chain chimeric polypeptide and the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), or the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein).
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the single-chain chimeric polypeptide includes one or more (e.g., two, three, four, five, six, seven, eight, nine, or ten) additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) at its C-terminus.
  • additional target-binding domains e.g., any of the exemplary target-binding domains described herein or known in the art
  • one of the one or more additional target-binding domains at the C-terminus of the single-chain chimeric polypeptide directly abuts the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), or the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein or known in the art).
  • the first target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the soluble tissue factor domain e.g., any of the exemplary soluble tissue factor domains described herein or known in the art.
  • the single-chain chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between one of the at least one additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) at the C-terminus of the single-chain chimeric polypeptide and the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), or the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein).
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the single-chain chimeric polypeptide comprises one or more (e.g., two, three, four, five, six, seven, eight, nine, or ten) additional target binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) at its N- terminus and its C-terminus.
  • additional target binding domains e.g., any of the exemplary target-binding domains described herein or known in the art
  • one of the one or more additional antigen binding domains at the N-terminus of the single-chain chimeric polypeptide directly abuts the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), or the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein).
  • the first target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the soluble tissue factor domain e.g., any of the exemplary soluble tissue factor domains described herein.
  • the single-chain chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between one of the one or more additional antigen-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) at the N-terminus and the first target-binding domain (e.g., any of the exemplary target- binding domains described herein or known in the art), the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), or the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains).
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • one of the one or more additional antigen binding domains at the C-terminus directly abuts the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), or the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains).
  • the single-chain chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between one of the one or more additional antigen-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) at the C-terminus and the first target-binding domain(e.g., any of the exemplary target-binding domains described herein or known in the art), the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art), or the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein).
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the first target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more additional target-binding domains bind specifically to the same antigen.
  • two or more (e.g., three, four, five, six, seven, eight, nine, or ten) of the first target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more additional target-binding domains bind specifically to the same epitope.
  • two or more (e.g., three, four, five, six, seven, eight, nine, or ten) of the first target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more additional target-binding domains include the same amino acid sequence.
  • the first target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more e.g., two, three, four, five, six, seven, eight, nine, or ten
  • additional target-binding domains e.g., any of the exemplary target-binding domains described herein or known in the art
  • the first target- binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target- binding domains described herein or known in the art
  • the one or more e.g., two, three, four, five, six, seven, eight, nine, or ten
  • additional target-binding domains e.g., any of the exemplary target-binding domains described herein or known in the art
  • the first target-binding domain, the second target-binding domain, and the one or more (e.g., two, three, four, five, six, seven, eight, nine, or ten) additional target-binding domains each comprise the same amino acid sequence.
  • the first target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more e.g., two, three, four, five, six, seven, eight, nine, or ten
  • additional target-binding domains e.g., any of the exemplary target-binding domains described herein or known in the art
  • one or more of the first target-binding domain, the second target-binding domain, and the one or more target-binding domains is an antigen-binding domain (e.g., any of the exemplary antigen-binding domains described herein or known in the art).
  • the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains are each an antigen-binding domain (e.g., any of the exemplary antigen-binding domains described herein or known in the art).
  • the antigen- binding domain can include a scFv or a single domain antibody.
  • one or more of the first target-binding domain e.g., any of the exemplary target- binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more additional target-binding domains e.g., any of the exemplary target- binding domains described herein or known in the art
  • a soluble interleukin or cytokine protein is a soluble interleukin or cytokine protein.
  • Non-limiting examples of soluble interleukin proteins and soluble cytokine proteins include: IL-1, IL-2, IL-3, IL-7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or more of the first target-binding domain e.g., any of the exemplary target- binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more additional target-binding domains e.g., any of the exemplary target- binding domains described herein or known in the art
  • a soluble interleukin or cytokine receptor is a soluble interleukin or cytokine receptor.
  • Non-limiting examples of soluble interleukin receptors and soluble cytokine receptors include: a soluble TGF- ⁇ receptor II (TGF- ⁇ RII), a soluble TGF- ⁇ RIII, a soluble NKG2D, a soluble NKP30, a soluble NKp44, a soluble NKp46, a soluble DNAM1, a scMHCI, a scMHCII, a scTCR, a soluble CD155, a soluble CD122, a soluble CD3, or a soluble CD28.
  • TGF- ⁇ RII soluble TGF- ⁇ receptor II
  • a soluble TGF- ⁇ RIII soluble NKG2D
  • a soluble NKP30 a soluble NKp44, a soluble NKp46
  • a soluble DNAM1 a scMHCI, a scMHCII, a scTCR
  • a soluble CD155 a
  • the first target-binding domain e.g., any of the target-binding domains described herein
  • the second target-binding domain e.g., any of the target-binding domains described herein
  • the one or more additional target-binding domains e.g., any of the target-binding domains described herein
  • one or more of the first target-binding domain e.g., any of the exemplary target- binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more additional target-binding domains e.g., any of the exemplary target- binding domains described herein or known in the art
  • a soluble interleukin or cytokine protein is a soluble interleukin or cytokine protein.
  • the soluble interleukin or cytokine protein is selected from the group of: IL-1, IL- 2, IL-3, IL-7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or more of the first target-binding domain e.g., any of the exemplary target- binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more additional target-binding domains e.g., any of the exemplary target- binding domains described herein or known in the art
  • a soluble interleukin or cytokine receptor is a soluble interleukin or cytokine receptor.
  • the soluble receptor is a soluble TGF- ⁇ receptor II (TGF- ⁇ RII) a soluble TGF- ⁇ RIII, a soluble receptor for TNF ⁇ , a soluble receptor for IL-4, or a soluble receptor for IL-10.
  • TGF- ⁇ RII soluble TGF- ⁇ receptor II
  • soluble TGF- ⁇ RIII soluble TGF- ⁇ RIII
  • soluble receptor for TNF ⁇ a soluble receptor for IL-4
  • a soluble receptor for IL-10 a soluble receptor for IL-10.
  • Non-limiting examples of NK cell activating agents are multi-chain chimeric polypeptides that include: (a) a first chimeric polypeptide including: (i) a first target- binding domain; (ii) a soluble tissue factor domain; and (iii) a first domain of a pair of affinity domains; and (b) a second chimeric polypeptide including: (i) a second domain of a pair of affinity domains; and (ii) a second target-binding domain, where the first chimeric polypeptide and the second chimeric polypeptide associate through the binding of the first domain and the second domain of the pair of affinity domains.
  • the total length of first chimeric polypeptide and/or the second chimeric polypeptide can each independently be about 50 amino acids to about 3000 amino acids, about 50 amino acids to about 2500 amino acids, about 50 amino acids to about 2000 amino acids, about 50 amino acids to about 1500 amino acids, about 50 amino acids to about 1000 amino acids, about 50 amino acids to about 950 amino acids, about 50 amino acids to about 900 amino acids, about 50 amino acids to about 850 amino acids, about 50 amino acids to about 800 amino acids, about 50 amino acids to about 750 amino acids, about 50 amino acids to about 700 amino acids, about 50 amino acids to about 650 amino acids, about 50 amino acids to about 600 amino acids, about 50 amino acids to about 550 amino acids, about 50 amino acids to about 500 amino acids, about 50 amino acids to about 480 amino acids, about 50 amino acids to about 460 amino acids, about 50 amino acids to about 440 amino acids, about 50 amino acids to about 420 amino acids, about 50 amino acids to about 400
  • the first target-binding domain e.g., any of the first target-binding domains described herein
  • the soluble tissue factor domain e.g., any of the exemplary soluble tissue factor domains described herein
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the first target-binding domain (e.g., any of the exemplary first target-binding domains described herein) and the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein) in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the soluble tissue factor domain e.g., any of the exemplary soluble tissue factor domains described herein
  • the first domain of the pair of affinity domains e.g., any of the exemplary first domains of any of the exemplary pairs of affinity domains described herein
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein) and the first domain of the pair of affinity domains (e.g., any of the exemplary first domains of any of the exemplary pairs of affinity domains described herein) in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the second domain of the pair of affinity domains e.g., any of the exemplary second domains of any of the exemplary pairs of affinity domains described herein
  • the second target-binding domain e.g., any of the exemplary second target-binding domains described herein
  • the second chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the second domain of the pair of affinity domains (e.g., any of the exemplary second domains of any of the exemplary pairs of affinity domains described herein) and the second target- binding domain (e.g., any of the exemplary second target-binding domains described herein) in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the first chimeric polypeptide further includes one or more (e.g., two, three, four, five, six, seven, eight, nine, or ten) additional target-binding domain(s) (e.g., any of the exemplary target- binding domains described herein or known in the art), where at least one of the one or more additional antigen-binding domain(s) is positioned between the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein or known in the art) and the first domain of the pair of affinity domains (e.g., any of the exemplary first domains of any of the exemplary pairs of affinity domains described herein).
  • additional target-binding domain(s) e.g., any of the exemplary target- binding domains described herein or known in the art
  • the first chimeric polypeptide can further include a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein) and the at least one of the one or more additional target- binding domain(s) (e.g., any of the exemplary target-binding domains described herein or known in the art), and/or a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the at least one of the one or more additional target-binding domain(s) (e.g., any of the exemplary target-binding domains described herein or known in the art) and the first domain of the pair of affinity domains (e.g., any of the exemplary first domains described herein of any of the exemplary pairs of affinity domains described herein).
  • a linker sequence e.g., any of the exemplary linker sequences
  • the first chimeric polypeptide further includes one or more (e.g., two, three, four, five, six, seven, eight, nine, or ten) additional target-binding domains at the N-terminal and/or C-terminal end of the first chimeric polypeptide.
  • At least one of the one or more additional target-binding domains directly abuts the first domain of the pair of affinity domains (e.g., any of the exemplary first domains described herein of any of the exemplary pairs of affinity domains described herein) in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the at least one of the one or more additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) and the first domain of the pair of affinity domains (e.g., any of the exemplary first domains described herein of any of the exemplary pairs of affinity domains described herein).
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the at least one of the one or more additional target-binding domains directly abuts the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the at least one of the one or more additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) and the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art).
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • At least one of the one or more additional target-binding domains is disposed at the N- and/or C-terminus of the first chimeric polypeptide, and at least one of the one or more additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) is positioned between the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein or known in the art) and the first domain of the pair of affinity domains (e.g., any of the exemplary first domains of any of the exemplary pairs of affinity domains described herein) in the first chimeric polypeptide.
  • the soluble tissue factor domain e.g., any of the exemplary soluble tissue factor domains described herein or known in the art
  • affinity domains e.g., any of the exemplary first domains of any of the exemplary pairs of affinity domains described herein
  • the at least one additional target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) of the one or more additional target-binding domains disposed at the N-terminus directly abuts the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) or the first domain of the pair of affinity domains (e.g., any of the exemplary first domains described herein of any of the exemplary pairs of affinity domains described herein) in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the linker sequences described herein or known in the art) disposed between the at least one additional target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) and the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) or the first domain of the pair of affinity domains (e.g., any of the exemplary first domains described herein of any of the exemplary pairs of affinity domains described herein) in the first chimeric polypeptide.
  • a linker sequence e.g., any of the linker sequences described herein or known in the art
  • the at least one additional target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) of the one or more additional target-binding domains disposed at the C-terminus directly abuts the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) or the first domain of the pair of affinity domains (e.g., any of the exemplary first domains of any of the exemplary pairs of affinity domains described herein) in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) disposed between the at least one additional target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) and the first target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) or the first domain of the pair of affinity domains (e.g., any of the exemplary first domains described herein of any of the exemplary pairs of affinity domains described herein) in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the at least one of the one or more additional target-binding domains positioned between the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein) and the first domain of the pair of affinity domains (e.g., any of the first domains described herein or any of the exemplary pairs of affinity domains described herein), directly abuts the soluble tissue factor domain and/or the first domain of the pair of affinity domains.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) disposed (i) between the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein) and the at least one of the one or more additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) positioned between the soluble tissue factor domain (e.g., any of the exemplary soluble tissue factor domains described herein) and the first domain of the pair of affinity domains (e.g., any of the exemplary first domains of any of the exemplary pairs of affinity domains described herein), and/or (ii) between the first domain of the pair of affinity domains and the at least one of the one or more additional target-binding domains positioned between the soluble tissue factor domain and the first domain of the pair of affinity domains.
  • a linker sequence e.g.,
  • the second chimeric polypeptide further includes one or more (e.g., two, three, four, five, six, seven, eight, nine, or ten) additional target-binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) at the N- terminal end and/or the C-terminal end of the second chimeric polypeptide.
  • additional target-binding domains e.g., any of the exemplary target-binding domains described herein or known in the art
  • At least one of the one or more additional target-binding domains directly abuts the second domain of the pair of affinity domains (e.g., any of the exemplary second domains of any of the exemplary pairs of affinity domains described herein) in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between at least one of the one or more additional target- binding domains (e.g., any of the exemplary target-binding domains described herein or known in the art) and the second domain of the pair of affinity domains (e.g., any of the second domains described herein of any of the exemplary pairs of affinity domains described herein) in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • At least one of the one or more additional target-binding domains directly abuts the second target- binding domain (e.g., any of the target-binding domains described herein or known in the art) in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between at least one of the one or more additional target-binding domains (e.g., any of the exemplary target binding domains described herein or known in the art) and the second target-binding domain (e.g., any of the exemplary target binding domains described herein or known in the art) in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • two or more (e.g., three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more) of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to the same antigen.
  • two or more (e.g., three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more) of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to the same epitope.
  • two or more (e.g., three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more) of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains include the same amino acid sequence.
  • the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains each bind specifically to the same antigen.
  • the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains each bind specifically to the same epitope.
  • the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains each include the same amino acid sequence. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to different antigens.
  • one or more (e.g., two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more) of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains is an antigen-binding domain.
  • the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains are each an antigen-binding domain (e.g., a scFv or a single-domain antibody).
  • a target selected from the group consisting of: CD16a, CD28, CD3,
  • one or more of the first target-binding domain e.g., any of the exemplary target- binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more additional target-binding domains e.g., any of the exemplary target- binding domains described herein or known in the art
  • a soluble interleukin or cytokine protein is a soluble interleukin or cytokine protein.
  • Non-limiting examples of soluble interleukin proteins and soluble cytokine proteins include: IL-1, IL-2, IL-3, IL-7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or more of the first target-binding domain e.g., any of the exemplary target- binding domains described herein or known in the art
  • the second target-binding domain e.g., any of the exemplary target-binding domains described herein or known in the art
  • the one or more additional target-binding domains e.g., any of the exemplary target- binding domains described herein or known in the art
  • a soluble interleukin or cytokine receptor is a soluble interleukin or cytokine receptor.
  • Non-limiting examples of soluble interleukin receptors and soluble cytokine receptors include: a soluble TGF- ⁇ receptor II (TGF- ⁇ RII), a soluble TGF- ⁇ RIII, a soluble NKG2D, a soluble NKP30, a soluble NKp44, a soluble NKp46, a soluble DNAM1, a scMHCI, a scMHCII, a scTCR, a soluble CD155, a soluble CD122, a soluble CD3, or a soluble CD28.
  • TGF- ⁇ RII soluble TGF- ⁇ receptor II
  • a soluble TGF- ⁇ RIII soluble NKG2D
  • a soluble NKP30 a soluble NKp44, a soluble NKp46
  • a soluble DNAM1 a scMHCI, a scMHCII, a scTCR
  • a soluble CD155 a
  • the first target-binding domain e.g., any of the target-binding domains described herein, the second target-binding domain (e.g., any of the target-binding domains described herein), and the one or more additional target-binding domains (e.g., any of the target-binding domains described herein) can each, independently, bind specifically to a target selected from the group of: CD16a, CD33, CD20, CD19, CD22, CD123, PDL-1, TIGIT, PD-1, TIM3, CTLA4, MICA, MICB, IL-6, IL-8, TNF ⁇ , CD26, CD36, ULBP2, CD30, CD200, IGF-1R, MUC4AC, MUC5AC, Trop-2, CMET, EGFR, HER1, HER2, HER3, PSMA, CEA, B7H3, EPCAM, BCMA, P-cadherin, CEACAM
  • one or both of the first target-binding domain (e.g., any of the target-binding domains described herein), the second target-binding domain (e.g., any of the target- binding domains described herein), and the one or more additional binding domains (e.g., any of the target-binding described herein) is a soluble interleukin or cytokine protein.
  • the soluble interleukin or cytokine protein is selected from the group of: IL-1, IL-2, IL-3, IL- 7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin or cytokine receptor.
  • the soluble receptor is a soluble TGF- ⁇ receptor II (TGF- ⁇ RII) a soluble TGF- ⁇ RIII, a soluble receptor for TNF ⁇ , a soluble receptor for IL-4, or a soluble receptor for IL-10.
  • TGF- ⁇ RII soluble TGF- ⁇ receptor II
  • soluble TGF- ⁇ RIII soluble TGF- ⁇ RIII
  • soluble receptor for TNF ⁇ a soluble receptor for IL-4
  • a soluble receptor for IL-10 a soluble receptor for IL-10.
  • Non-limiting examples of NK cell activating agents are multi-chain chimeric polypeptides that include: (a) a first and second chimeric polypeptide each including: (i) a first target-binding domain; (ii) a Fc domain; and (iii) a first domain of a pair of affinity domains; and (b) a third and fourth chimeric polypeptide each including: (i) a second domain of a pair of affinity domains; and (ii) a second target-binding domain, where the first and second chimeric polypeptides and the third and fourth chimeric polypeptides associate through the binding of the first domain and the second domain of the pair of affinity domains, and the first and second chimeric polypeptides associate through their Fc domains.
  • the first target-binding domain e.g., any of the first target-binding domains described herein
  • the Fc domain e.g., any of the exemplary Fc domains described herein
  • the first and second chimeric polypeptides further comprise a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the first target- binding domain (e.g., any of the exemplary first target-binding domains described herein) and the Fc domain (e.g., any of the exemplary Fc domains described herein) in the first and second chimeric polypeptides.
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the Fc domain e.g., any of the exemplary Fc domains described herein
  • the first domain of the pair of affinity domains e.g., any of the exemplary first domains of any of the exemplary pairs of affinity domains described herein
  • the first and second chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the Fc domain (e.g., any of the exemplary Fc domains described herein) and the first domain of the pair of affinity domains (e.g., any of the exemplary first domains of any of the exemplary pairs of affinity domains described herein) in the first and second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linker sequences described herein or known in the art
  • the second domain of the pair of affinity domains e.g., any of the exemplary second domains of any of the exemplary pairs of affinity domains described herein
  • the second target-binding domain e.g., any of the exemplary second target-binding domains described herein
  • the third and fourth chimeric polypeptide further comprise a linker sequence (e.g., any of the exemplary linker sequences described herein or known in the art) between the second domain of the pair of affinity domains (e.g., any of the exemplary second domains of any of the exemplary pairs of affinity domains described herein) and the second target-binding domain (e.g., any of the exemplary second target-binding domains described herein) in the third and fourth chimeric polypeptide.
  • the first target-binding domain and the second target-binding domain bind specifically to the same antigen.
  • the first target-binding domain and the second target- binding domain bind specifically to the same epitope. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the first target-binding domain and the second target-binding domain include the same amino acid sequence. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the first target-binding domain and the second target-binding domain bind specifically to different antigens.
  • first target-binding domain and the second target- binding domain is an antigen-binding domain (e.g., any of the exemplary second target- binding domains described herein).
  • first target-binding domain and the second target-binding domain are each antigen-binding domains (e.g., any of the exemplary second target-binding domains described herein).
  • the antigen-binding domain (e.g., any of the exemplary second target-binding domains described herein) includes a scFv or a single domain antibody.
  • one or both of the first target-binding domain (e.g., any of the exemplary target- binding domains described herein or known in the art) and the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) bind specifically to a target selected from the group consisting of: CD16a, CD28, CD3, CD33, CD20, CD19, CD22, CD123, IL-1R, IL-1, VEGF, IL-6R, IL-4, IL-10, PDL- 1, TIGIT, PD-1, TIM3, CTLA4, MICA, MICB, IL-6, IL-8, TNF ⁇ , CD26, CD36, ULBP2, CD30, CD200, IGF-1R, MUC4AC, MUC5AC, Trop-2, CMET, EGFR, HER1, HER2, HER3, PSMA, CEA, B7H3, EP
  • one or both of the first target-binding domain (e.g., any of the exemplary target- binding domains described herein or known in the art) and the second target-binding domain (e.g., any of the exemplary target-binding domains described herein or known in the art) is a soluble interleukin or cytokine protein.
  • soluble interleukin proteins and soluble cytokine proteins include: IL-1, IL-2, IL-3, IL-7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin or cytokine receptor.
  • Non-limiting examples of soluble interleukin receptors and soluble cytokine receptors include: a soluble TGF- ⁇ receptor II (TGF- ⁇ RII), a soluble TGF- ⁇ RIII, a soluble NKG2D, a soluble NKp30, a soluble NKp44, a soluble NKp46, a soluble DNAM1, a scMHCI, a scMHCII, a scTCR, a soluble CD155, a soluble CD122, a soluble CD3, or a soluble CD28.
  • TGF- ⁇ RII soluble TGF- ⁇ receptor II
  • a soluble TGF- ⁇ RIII soluble NKG2D
  • a soluble NKp30 a soluble NKp44, a soluble NKp46
  • a soluble DNAM1 a scMHCI, a scMHCII, a scTCR
  • a soluble CD155 a
  • the first target-binding domain and the second target-binding domain can each, independently, bind specifically to a target selected from the group of: CD16a, CD33, CD20, CD19, CD22, CD123, PDL-1, TIGIT, PD-1, TIM3, CTLA4, MICA, MICB, IL-6, IL-8, TNF ⁇ , CD26, CD36, ULBP2, CD30, CD200, IGF-1R, MUC4AC, MUC5AC, Trop-2, CMET, EGFR, HER1, HER2, HER3, PSMA, CEA, B7H3, EPCAM, BCMA, P- cadherin, CEACAM5, a UL16-binding protein, HLA-DR, DLL4, TYRO3, AXL, MER, CD122, CD155, PDGF-DD, a ligand of TGF- ⁇ receptor II (TGF- ⁇ RII), a ligand of
  • one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin or cytokine protein.
  • the soluble interleukin or cytokine protein is selected from the group of: IL-1, IL-2, IL-3, IL-7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin or cytokine receptor.
  • the soluble receptor is a soluble TGF- ⁇ receptor II (TGF- ⁇ RII) a soluble TGF- ⁇ RIII, a soluble receptor for TNF ⁇ , a soluble receptor for IL-4, or a soluble receptor for IL-10.
  • Tissue Factor Human tissue factor is a 263 amino-acid transmembrane protein containing three domains: (1) a 219-amino acid N-terminal extracellular domain (residues 1-219); (2) a 22-amino acid transmembrane domain (residues 220-242); and (3) a 21-amino acid cytoplasmic C-terminal tail (residues 242-263) ((UniProtKB Identifier Number: P13726).
  • the cytoplasmic tail contains two phosphorylation sites at Ser253 and Ser258, and one S- palmitoylation site at Cys245. Deletion or mutation of the cytoplasmic domain was not found to affect tissue factor coagulation activity.
  • Tissue factor has one S-palmitoylation site in the intracellular domain of the protein at Cys245.
  • the Cys245 is located at the amino acid terminus of the intracellular domain and close to the membrane surface.
  • the tissue factor transmembrane domain is composed of a single-spanning ⁇ -helix.
  • the extracellular domain of tissue factor composed of two fibronectin type III domains, is connected to the transmembrane domain through a six-amino acid linker. This linker provides conformational flexibility to decouple the tissue factor extracellular domain from its transmembrane and cytoplasmic domains.
  • Each tissue factor fibronectin type III module is composed of two overlapping ⁇ sheets with the top sheet domain containing three antiparallel ⁇ -strands and the bottom sheet containing four ⁇ -strands.
  • the ⁇ -strands are connected by ⁇ -loops between strand ⁇ A and ⁇ B, ⁇ C and ⁇ D, and ⁇ E and ⁇ F, all of which are conserved in conformation in the two modules.
  • a unique feature of tissue factor is a 17- amino acid ⁇ -hairpin between strand ⁇ 10 and strand ⁇ 11, which is not a common element of the fibronectin superfamily.
  • the N-terminal domain also contains a 12 amino acid loop between ⁇ 6F and ⁇ 7G that is not present in the C-terminal domain and is unique to tissue factor.
  • a fibronectin type III domain structure is a feature of the immunoglobulin-like family of protein folds and is conserved among a wide variety of extracellular proteins.
  • the zymogen FVII is rapidly converted to FVIIa by limited proteolysis once it binds to tissue to form the active tissue factor-FVIIa complex.
  • the FVIIa which circulates as an enzyme at a concentration of approximately 0.1 nM (1% of plasma FVII), can also bind directly to tissue factor.
  • tissue factor and FVIIa on the tissue factor-FVIIa complex greatly increases the enzymatic activity of FVIIa: an approximate 20- to 100-fold increase in the rate of hydrolysis of small, chromogenic peptidyl substrates, and nearly a million-fold increase in the rate of activation of the natural macromolecular substrates FIX and FX.
  • tissue factor-FVIIa complex on phospholipid bilayer i.e., upon exposure of phosphatidyl-L-serine on membrane surfaces
  • FIX or FX activation increases the rate of FIX or FX activation, in a Ca 2+ -dependent manner, an additional 1,000-fold.
  • the roughly million-fold overall increase in FX activation by tissue factor-FVIIa-phospholipid complex relative to free FVIIa is a critical regulatory point for the coagulation cascade.
  • FVII is a ⁇ 50 kDa, single-chain polypeptide consisting of 406 amino acid residues, with an N-terminal ⁇ -carboxyglutamate-rich (GLA) domain, two epidermal growth factor-like domains (EGF1 and EFG2), and a C-terminal serine protease domain.
  • GLA N-terminal ⁇ -carboxyglutamate-rich
  • EGF1 and EFG2 epidermal growth factor-like domains
  • C-terminal serine protease domain is activated to FVIIa by a specific proteolytic cleavage of the Ile- 154 -Arg 152 bond in the short linker region between the EGF2 and the protease domain. This cleavage results in the light and heavy chains being held together by a single disulfide bond of Cys 135 and Cys 262 .
  • FVIIa binds phospholipid membrane in a Ca 2+ -dependent manner through its N- terminal GLA-domain.
  • GLA domain Immediately C-terminal to the GLA domain is an aromatic stack and two EGF domains.
  • the aromatic stack connects the GLA to EGF1 domain which binds a single Ca 2+ ion. Occupancy of this Ca 2+ -binding site increases FVIIa amidolytic activity and tissue factor association.
  • the catalytic triad consist of His 193 , Asp 242 , and Ser 344 , and binding of a single Ca 2+ ion within the FVIIa protease domain is critical for its catalytic activity.
  • FVIIa Proteolytic activation of FVII to FVIIa frees the newly formed amino terminus at Ile 153 to fold back and be inserted into the activation pocket forming a salt bridge with the carboxylate of Asp 343 to generate the oxyanion hole. Formation of this salt bridge is critical for FVIIa activity. However, oxyanion hole formation does not occur in free FVIIa upon proteolytic activation. As a result, FVIIa circulates in a zymogen-like state that is poorly recognized by plasma protease inhibitors, allowing it to circulate with a half-life of approximately 90 minutes. Tissue factor-mediated positioning of the FVIIa active site above the membrane surface is important for FVIIa towards cognate substrates.
  • Free FVIIa adopts a stable, extended structure when bound to the membrane with its active site positioned ⁇ 80 ⁇ above the membrane surface.
  • the FVa active site Upon FVIIa binding to tissue factor, the FVa active site is repositioned ⁇ 6 ⁇ closer to the membrane. This modulation may aid in a proper alignment of the FVIIa catalytic triad with the target substrate cleavage site.
  • GLA- domainless FVIIa it has been shown that the active site was still positioned a similar distance above the membrane, demonstrating that tissue factor is able to fully support FVIIa active site positioning even in the absence of FVIIa-membrane interaction.
  • tissue factor supported full FVIIa proteolytic activity as long as the tissue factor extracellular domain was tethered in some way to the membrane surface.
  • raising the active site of FVIIa greater than 80 ⁇ above the membrane surface greatly reduced the ability of the tissue factor-FVIIa complex to activate FX but did not diminish tissue factor-FVIIa amidolytic activity.
  • Alanine scanning mutagenesis has been used to assess the role of specific amino acid side chains in the tissue factor extracellular domain for interaction with FVIIa (Gibbs et al., Biochemistry 33(47): 14003-14010, 1994; Schullek et al., J Biol Chem 269(30): 19399-19403, 1994).
  • Thr 60 is only partially solvent-exposed and may play a local structural role rather than making a significant contact with ligand.
  • the binding site extends onto the concave side of the intermodule angle involving Glu 24 and Gln 110 , and potentially the more distant residue Val 207 .
  • the binding region extends from Asp58 onto a convex surface area formed by Lys 48 , Lys 46 , Gln 37 , Asp 44 , and Trp 45 .
  • Trp 45 and Asp 44 do not interact independently with FVIIa, indicating that the mutational effect at the Trp 45 position may reflect a structural importance of this side chain for the local packing of the adjacent Asp 44 and Gln 37 side chain.
  • the interactive area further includes two surface- exposed aromatic residues, Phe 76 and Tyr 78 , which form part of the hydrophobic cluster in the N-module.
  • the known physiologic substrates of tissue factor-FVIIa are FVII, FIX, and FX and certain proteinase-activated receptors.
  • Mutational analysis has identified a number of residues that, when mutated, support full FVIIa amidolytic activity towards small peptidyl substrates but are deficient in their ability to support macromolecular substrate (i.e., FVII, FIX, and FX) activation (Ruf et al., J Biol Chem 267(31): 22206-22210, 1992; Ruf et al., J Biol Chem 267(9): 6375-6381, 1992; Huang et al., J Biol Chem 271(36): 21752-21757, 1996; Kirchhofer et al., Biochemistry 39(25): 7380-7387, 2000).
  • macromolecular substrate i.e., FVII, FIX, and FX
  • tissue factor loop region at residues 159-165, and residues in or adjacent to this flexible loop have been shown to be critical for the proteolytic activity of the tissue factor-FVIIa complex.
  • the residues Lys 165 and Lys 166 have also been demonstrated to be important for substrate recognition and binding.
  • Lys 165 and Lys 166 face away from each other, with Lys 165 pointing towards FVIIa in most tissue factor-FVIIa structures, and Lys 166 pointing into the substrate binding exosite region in the crystal structure. Putative salt bridge formation between Lys 165 of and Gla 35 of FVIIa would support the notion that tissue factor interaction with the GLA domain of FVIIa modulates substrate recognition.
  • the soluble tissue factor domain can be a wildtype tissue factor polypeptide lacking the signal sequence, the transmembrane domain, and the intracellular domain.
  • the soluble tissue factor domain can be a tissue factor mutant, wherein a wildtype tissue factor polypeptide lacking the signal sequence, the transmembrane domain, and the intracellular domain, and has been further modified at selected amino acids.
  • the soluble tissue factor domain can be a soluble human tissue factor domain.
  • the soluble tissue factor domain can be a soluble mouse tissue factor domain.
  • the soluble tissue factor domain can be a soluble rat tissue factor domain.
  • Soluble Human Tissue Factor Domain (SEQ ID NO: 93) SGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTD TECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQ PTIQSFEQVGTKVNVTVEDERTLVRRNNTFLSLRDVFGKDLIYTLYYWKSSSSGK KTAKTNTNEFLIDVDKGENYCFSVQAVIPSRTVNRKSTDSPVECMGQEKGEFRE
  • Nucleic Acid Encoding Soluble Human Tissue Factor Domain (SEQ ID NO: 94) AGCGGCACAACCAACACAGTCGCTGCCTATAACCTCACTTGGAAGAGCACCA ACTTCAAAACCATCCTCGAATGGGAACCCAAACCCGTTAACCAAGTTTACACC GTGCAGATCAGCACCAAGTCCGGCGACTGGAAGTCCAAATGTTTCTATACCAC CGACACCGA
  • a soluble tissue factor domain can include a sequence of SEQ ID NO: 93, 95, 96, 97, or 98, with one to twenty amino acids (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) amino acids removed from its N-terminus and/or one to twenty amino acids (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) amino acids removed from its C-terminus.
  • amino acids e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20
  • the soluble tissue factor domain is not capable of binding to Factor VIIa. In some examples of any of the multi-chain chimeric polypeptides described herein, the soluble tissue factor domain does not convert inactive Factor X into Factor Xa.
  • the multi-chain chimeric polypeptide does not stimulate blood coagulation in a mammal.
  • the soluble tissue factor domain can be a soluble human tissue factor domain.
  • the soluble tissue factor domain can be a soluble mouse tissue factor domain.
  • the soluble tissue factor domain can be a soluble rat tissue factor domain.
  • the soluble tissue factor domain does not include one or more (e.g., two, three, four, five, six, or seven) of: a lysine at an amino acid position that corresponds to amino acid position 20 of mature wildtype human tissue factor protein; an isoleucine at an amino acid position that corresponds to amino acid position 22 of mature wildtype human tissue factor protein; a tryptophan at an amino acid position that corresponds to amino acid position 45 of mature wildtype human tissue factor protein; an aspartic acid at an amino acid position that corresponds to amino acid position 58 of mature wildtype human tissue factor protein; a tyrosine at an amino acid position that corresponds to amino acid position 94 of mature wildtype human tissue factor protein; an arginine at an amino acid position that corresponds to amino acid position 135 of mature wildtype human tissue factor protein; and a phenylalanine at an amino acid position that corresponds to amino acid position 140 of mature wildtype human tissue factor protein.
  • a lysine at an amino acid position that corresponds to amino acid position
  • the mutant soluble tissue factor possesses the amino acid sequence of SEQ ID NO: 97 or SEQ ID NO: 98.
  • the soluble tissue factor domain can be encoded by a nucleic acid including a sequence that is at least 70% identical, at least 72% identical, at least 74% identical, at least 76% identical, at least 78% identical, at least 80% identical, at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical to SEQ ID NO: 94.
  • the soluble tissue factor domain can have a total length of about 20 amino acids to about 220 amino acids, about 20 amino acids to about 215 amino acids, about 20 amino acids to about 210 amino acids, about 20 amino acids to about 205 amino acids, about 20 amino acids to about 200 amino acids, about 20 amino acids to about 195 amino acids, about 20 amino acids to about 190 amino acids, about 20 amino acids to about 185 amino acids, about 20 amino acids to about 180 amino acids, about 20 amino acids to about 175 amino acids, about 20 amino acids to about 170 amino acids, about 20 amino acids to about 165 amino acids, about 20 amino acids to about 160 amino acids, about 20 amino acids to about 155 amino acids, about 20 amino acids to about 150 amino acids, about 20 amino acids to about 145 amino acids, about 20 amino acids to about 140 amino acids, about 20 amino acids to about 135 amino acids, about 20 amino acids to about 130 amino acids, about 20 amino acids to about 125 amino acids, about 20 amino acids to about 120 amino acids, about 20 amino acids to about 115 amino acids, about 20 amino acids to about
  • the linker sequence can be a flexible linker sequence.
  • linker sequences that can be used are described in Klein et al., Protein Engineering, Design & Selection 27(10):325–330, 2014; Priyanka et al., Protein Sci.22(2):153–167, 2013.
  • the linker sequence is a synthetic linker sequence.
  • any of the single-chain chimeric polypeptides described herein can include one, two, three, four, five, six, seven, eight, nine, or ten linker sequence(s) (e.g., the same or different linker sequences, e.g., any of the exemplary linker sequences described herein or known in the art).
  • any of the single-chain chimeric polypeptides described herein can include one, two, three, four, five, six, seven, eight, nine, or ten linker sequence(s) (e.g., the same or different linker sequences, e.g., any of the exemplary linker sequences described herein or known in the art).
  • the first chimeric polypeptide can include one, two, three, four, five, six, seven, eight, nine, or ten linker sequence(s) (e.g., the same or different linker sequences, e.g., any of the exemplary linker sequences described herein or known in the art).
  • the second chimeric polypeptide can include one, two, three, four, five, six, seven, eight, nine, or ten linker sequence(s) (e.g., the same or different linker sequences, e.g., any of the exemplary linker sequences described herein or known in the art).
  • a linker sequence can have a total length of 1 amino acid to about 100 amino acids, 1 amino acid to about 90 amino acids, 1 amino acid to about 80 amino acids, 1 amino acid to about 70 amino acids, 1 amino acid to about 60 amino acids, 1 amino acid to about 50 amino acids, 1 amino acid to about 45 amino acids, 1 amino acid to about 40 amino acids, 1 amino acid to about 35 amino acids, 1 amino acid to about 30 amino acids, 1 amino acid to about 25 amino acids, 1 amino acid to about 24 amino acids, 1 amino acid to about 22 amino acids, 1 amino acid to about 20 amino acids, 1 amino acid to about 18 amino acids, 1 amino acid to about 16 amino acids, 1 amino acid to about 14 amino acids, 1 amino acid to about 12 amino acids, 1 amino acid to about 10 amino acids, 1 amino acid to about 8 amino acids, 1 amino acid to about 6 amino acids, 1 amino acid to about 4 amino acids, about 2 amino acids to about 100 amino acids, about 2 amino acids to about 90 amino acids, about 2 amino acids to about 80 amino acids, about 2 amino acids to about 70 amino acids,
  • the linker is rich in glycine (Gly or G) residues. In some embodiments, the linker is rich in serine (Ser or S) residues. In some embodiments, the linker is rich in glycine and serine residues. In some embodiments, the linker has one or more glycine-serine residue pairs (GS), e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 or more GS pairs. In some embodiments, the linker has one or more Gly-Gly-Gly-Ser (GGGS) (SEQ ID NO: 99) sequences, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 or more GGGS (SEQ ID NO: 99) sequences.
  • GS glycine-serine residue pairs
  • GGGS Gly-Gly-Gly-Ser
  • the linker has one or more Gly-Gly-Gly-Gly-Ser (GGGGS) sequences, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 or more GGGGS (SEQ ID NO: 100) sequences.
  • the linker has one or more Gly-Gly-Ser-Gly (GGSG) (SEQ ID NO: 101) sequences, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 or more GGSG (SEQ ID NO: 101) sequences.
  • the linker comprises GGSSRSSSSGGGGSGGGG (SEQ ID NO: 222).
  • the linker sequence can comprise or consist of GGGGSGGGGSGGGGS (SEQ ID NO: 102). In some embodiments, the linker sequence can be encoded by a nucleic acid comprising or consisting of: GGCGGTGGAGGATCCGGAGGAGGTGGCTCCGGCGGCGGAGGATCT (SEQ ID NO: 103).
  • the linker sequence can comprise or consist of: GGGSGGGS (SEQ ID NO: 104), Target-Binding Domains
  • the first target-binding domain, the second target-binding domain, and/or the additional one or more target-binding domains can be an antigen-binding domain (e.g., any of the exemplary antigen-binding domains described herein or known in the art), a soluble interleukin or cytokine protein (e.g., any of the exemplary soluble interleukin proteins or soluble cytokine proteins described herein), and a soluble interleukin or cytokine receptor (e.g., any of the exemplary soluble interleukin receptors or soluble cytokine receptors described herein).
  • an antigen-binding domain e.g., any of the exemplary antigen-binding domains described herein or known in the art
  • a soluble interleukin or cytokine protein e.g., any of the exemplary
  • the first target-binding domain, the second target-binding domain, and/or the one or more additional target-binding domains can each independent have a total number of amino acids of about 5 amino acids to about 1000 amino acids, about 5 amino acids to about 950 amino acids, about 5 amino acids to about 900 amino acids, about 5 amino acids to about 850 amino acids, about 5 amino acids to about 800 amino acids, about 5 amino acids to about 750 amino acids, about 5 amino acids to about 700 amino acids, about 5 amino acids to about 650 amino acids, about 5 amino acids to about 600 amino acids, about 5 amino acids to about 550 amino acids, about 5 amino acids to about 500 amino acids, about 5 amino acids to about 450 amino acids, about 5 amino acids to about 400 amino acids, about 5 amino acids to about 350 amino acids, about 5 amino acids to about 300 amino acids, about 5 amino acids to about 280 amino acids, about 5 amino acids to about 260 amino acids, about 5 amino acids to about 240 amino acids, about
  • any of the target-binding domains described herein can bind to its target with a dissociation equilibrium constant (K D ) of less than 1 x 10 -7 M, less than 1 x 10 -8 M, less than 1 x 10 -9 M, less than 1 x 10 -10 M, less than 1 x 10 -11 M, less than 1 x 10 -12 M, or less than 1 x 10 -13 M.
  • K D dissociation equilibrium constant
  • the antigen-binding protein construct provided herein can bind to an identifying antigen with a KD of about 1 x 10 -3 M to about 1 x 10 -5 M, about 1 x 10 -4 M to about 1 x 10 -6 M, about 1 x 10 -5 M to about 1 x 10 -7 M, about 1 x 10 -6 M to about 1 x 10 -8 M, about 1 x 10 -7 M to about 1 x 10 -9 M, about 1 x 10 -8 M to about 1 x 10 -10 M, or about 1 x 10 -9 M to about 1 x 10 -11 M (inclusive).
  • any of the target-binding domains described herein can bind to its target with a K D of between about 1 pM to about 30 nM (e.g., about 1 pM to about 25 nM, about 1 pM to about 20 nM, about 1 pM to about 15 nM, about 1 pM to about 10 nM, about 1 pM to about 5 nM, about 1 pM to about 2 nM, about 1 pM to about 1 nM, about 1 pM to about 950 pM, about 1 pM to about 900 pM, about 1 pM to about 850 pM, about 1 pM to about 800 pM, about 1 pM to about 750 pM, about 1 pM to about 700 pM, about 1 pM to about 650 pM, about 1 pM to about 600 pM, about 1 pM to about 550 pM, about 1 pM to about 500 pM, about 1
  • any of the target-binding domains described herein can bind to its target with a KD of between about 1 nM to about 10 nM (e.g., about 1 nM to about 9 nM, about 1 nM to about 8 nM, about 1 nM to about 7 nM, about 1 nM to about 6 nM, about 1 nM to about 5 nM, about 1 nM to about 4 nM, about 1 nM to about 3 nM, about 1 nM to about 2 nM, about 2 nM to about 10 nM, about 2 nM to about 9 nM, about 2 nM to about 8 nM, about 2 nM to about 7 nM, about 2 nM to about 6 nM, about 2 nM to about 5 nM, about 2 nM to about 4 nM, about 2 nM to about 3 nM, about 3 nM to about 10 nM, about 3 nM to about 10
  • any of the antigen-binding protein constructs described herein e.g., an electrophoretic mobility shift assay, a filter binding assay, surface plasmon resonance, and a biomolecular binding kinetics assay, etc.
  • Antigen-Binding Domains In some embodiments of any of the single-chain or multi-chain chimeric polypeptides described herein, the first target-binding domain and the second target- binding domain bind specifically to the same antigen. In some embodiments of these single-chain or multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain bind specifically to the same epitope.
  • the first target- binding domain and the second target-binding domain include the same amino acid sequence. In some embodiments of any of the single-chain or multi-chain chimeric polypeptides described herein, the first target-binding domain and the second target- binding domain bind specifically to different antigens. In some embodiments of any of the single-chain or multi-chain chimeric polypeptides described herein, one or both of the first target-binding domain and the second target-binding domain is an antigen-binding domain.
  • the first target-binding domain and the second target-binding domain are each antigen-binding domains.
  • the antigen-binding domain includes or is a scFv or a single domain antibody (e.g., a VaHH or a VNAR domain).
  • an antigen-binding domain e.g., any of the antigen-binding domains described herein
  • Patent No.9,035,026) CD28 (see, e.g., those described in U.S. Patent No.7,723,482), CD3 (see, e.g., those described in U.S. Patent No.9,226,962), CD33 (see, e.g., those described in U.S. Patent No.8,759,494), CD20 (see, e.g., those described in WO 2014/026054), CD19 (see, e.g., those described in U.S.
  • Patent No.9,701,758 CD22 (see, e.g., those described in WO 2003/104425), CD123 (see, e.g., those described in WO 2014/130635), IL-1R (see, e.g., those described in U.S. Patent No.8,741,604), IL- 1 (see, e.g., those described in WO 2014/095808), VEGF (see, e.g., those described in U.S. Patent No.9,090,684), IL-6R (see, e.g., those described in U.S. Patent No. 7,482,436), IL-4 (see, e.g., those described in U.S. Patent Application Publication No.
  • IL-10 see, e.g., those described in U.S. Patent Application Publication No.2016/0340413
  • PDL-1 see, e.g., those described in Drees et al., Protein Express. Purif.94:60-66, 2014
  • TIGIT see, e.g., those described in U.S. Patent Application Publication No.2017/0198042
  • PD-1 see, e.g., those described in U.S. Patent No. 7,488,802
  • TIM3 see, e.g., those described in U.S.
  • Patent No.8,552,156 CTLA4 (see, e.g., those described in WO 2012/120125), MICA (see, e.g., those described in WO 2016/154585), MICB (see, e.g., those described in U.S. Patent No.8,753,640), IL-6 (see, e.g., those described in Gejima et al., Human Antibodies 11(4):121-129, 2002), IL-8 (see, e.g., those described in U.S. Patent No.6,117,980), TNF ⁇ (see, e.g., those described in Geng et al., Immunol.
  • CD26 see, e.g., those described in WO 2017/189526
  • CD36 see, e.g., those described in U.S. Patent Application Publication No.2015/0259429
  • ULBP2 see, e.g., those described in U.S. Patent No.9,273,136
  • CD30 see, e.g., those described in Homach et al., Scand. J. Immunol.48(5):497-501, 1998)
  • CD200 see, e.g., those described in U.S. Patent No.9,085,623
  • IGF-1R see, e.g., those described in U.S.
  • MUC4AC see, e.g., those described in WO 2012/170470
  • MUC5AC see, e.g., those described in U.S. Patent No.9,238,084
  • Trop-2 see, e.g., those described in WO 2013/068946
  • CMET see, e.g., those described in Edwardraja et al., Biotechnol. Bioeng.106(3):367- 375, 2010
  • EGFR see, e.g., those described in Akbari et al., Protein Expr. Purif.127:8- 15, 2016
  • HER1 see, e.g., those described in U.S.
  • HER2 see, e.g., those described in Cao et al., Biotechnol. Lett. 37(7):1347-1354, 2015
  • HER3 see, e.g., those described in U.S. Patent No.9,505,843
  • PSMA see, e.g., those described in Parker et al., Protein Expr. Purif.89(2):136-145, 2013
  • CEA see, e.g., those described in WO 1995/015341
  • B7H3 see, e.g., those described in U.S.
  • Patent No.9,371,395) EPCAM (see, e.g., those described in WO 2014/159531), BCMA (see, e.g., those described in Smith et al., Mol. Ther.26(6):1447- 1456, 2018), P-cadherin (see, e.g., those described in U.S. Patent No.7,452,537), CEACAM5 (see, e.g., those described in U.S. Patent No.9,617,345), a UL16-binding protein (see, e.g., those described in WO 2017/083612), HLA-DR (see, e.g., Pistillo et al., Exp. Clin.
  • any of the antigen-binding domains present in any of the single-chain or multi-chain chimeric polypeptides described herein are each independently selected from the group consisting of: a VHH domain, a VNAR domain, and a scFv.
  • any of the antigen-binding domains described herein is a BiTe, a (scFv)2, a nanobody, a nanobody-HSA, a DART, a TandAb, a scDiabody, a scDiabody-CH3, scFv-CH-CL- scFv, a HSAbody, scDiabody-HAS, or a tandem-scFv.
  • a VHH domain is a single monomeric variable antibody domain that can be found in camelids.
  • a VNAR domain is a single monomeric variable antibody domain that can be found in cartilaginous fish. Non-limiting aspects of VHH domains and VNAR domains are described in, e.g., Cromie et al., Curr. Top. Med. Chem.15:2543-2557, 2016; De Genst et al., Dev. Comp.
  • each of the antigen-binding domains in the single-chain or multi-chain chimeric polypeptides described herein are both VHH domains, or at least one antigen-binding domain is a VHH domain. In some embodiments, each of the antigen-binding domains in the single-chain or multi-chain chimeric polypeptides described herein are both VNAR domains, or at least one antigen-binding domain is a VNAR domain. In some embodiments, each of the antigen-binding domains in the single-chain or multi-chain chimeric polypeptides described herein are both scFv domains, or at least one antigen-binding domain is a scFv domain.
  • two or more of polypeptides present in the single-chain or multi-chain chimeric polypeptide can assemble (e.g., non-covalently assemble) to form any of the antigen-binding domains described herein, e.g., an antigen-binding fragment of an antibody (e.g., any of the antigen-binding fragments of an antibody described herein), a VHH-scAb, a VHH-Fab, a Dual scFab, a F(ab’)2, a diabody, a crossMab, a DAF (two- in-one), a DAF (four-in-one), a DutaMab, a DT-IgG, a knobs-in-holes common light chain, a knobs-in-holes assembly, a charge pair, a Fab-arm exchange, a SEEDbody, a LUZ-Y, a Fcab, a ⁇ -body, an orthogonal
  • Non-limiting examples of an antigen-binding fragment of an antibody include an Fv fragment, a Fab fragment, a F(ab')2 fragment, and a Fab' fragment.
  • an antigen-binding fragment of an antibody is an antigen-binding fragment of an IgG (e.g., an antigen-binding fragment of IgG1, IgG2, IgG3, or IgG4) (e.g., an antigen-binding fragment of a human or humanized IgG, e.g., human or humanized IgG1, IgG2, IgG3, or IgG4); an antigen-binding fragment of an IgA (e.g., an antigen-binding fragment of IgA1 or IgA2) (e.g., an antigen-binding fragment of a human or humanized IgA, e.g., a human or humanized IgA1 or IgA2); an antigen-binding fragment of an IgD (e.g., an antigen-binding fragment of a human or humanized IgD); an antigen-binding fragment of an IgE (e.g., an antigen-binding fragment of a human
  • An “Fv” fragment includes a non-covalently-linked dimer of one heavy chain variable domain and one light chain variable domain.
  • a “Fab” fragment includes, the constant domain of the light chain and the first constant domain (CH1) of the heavy chain, in addition to the heavy and light chain variable domains of the Fv fragment.
  • a “F(ab') 2 ” fragment includes two Fab fragments joined, near the hinge region, by disulfide bonds.
  • a “dual variable domain immunoglobulin” or “DVD-Ig” refers to multivalent and multispecific binding proteins as described, e.g., in DiGiammarino et al., Methods Mol.
  • DARTs are described in, e.g., Garber, Nature Reviews Drug Discovery 13:799- 801, 2014.
  • any of the antigen-binding domains described herein can bind to an antigen selected from the group consisting of: a protein, a carbohydrate, a lipid, and a combination thereof.
  • one or both of the first target-binding domain and the second target-binding domain can be a soluble interleukin protein or soluble cytokine protein.
  • the soluble interleukin or soluble cytokine protein is selected from the group of: IL-2, IL-3, IL-7, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, IL- 21, PDGF-DD, and SCF.
  • Non-limiting examples of soluble IL-2, IL-3, IL-7, IL-8, IL-10, IL-15, IL-17, IL-18, IL-21, PDGF-DD, and SCF are provided below.
  • Human Soluble IL-3 (SEQ ID NO: 105) apmtqttplkt swvncsnmid eiithlkqpp lplldfnnln gedqdilmen nlrrpnleaf nravkslqna saiesilknl lpclplataa ptrhpihikd gdwnefrrkl tfylktlena qaqqttlsla if Human Soluble IL-8 (SEQ ID NO: 106) egavlprsak elrcqcikty skpfhpkfik elrviesgph cantei
  • Soluble Receptor In some embodiments of any of the single-chain or multi-chain chimeric polypeptides described herein, one or both of the first target-binding domain and the second target-binding domain is a soluble interleukin receptor or a soluble cytokine receptor. In some embodiments, the soluble receptor is a soluble TGF- ⁇ receptor II (TGF- ⁇ RII) (see, e.g., those described in Yung et al., Am. J. Resp. Crit. Care Med.
  • TGF- ⁇ RII soluble TGF- ⁇ receptor II
  • a soluble TGF- ⁇ RIII see, e.g., those described in Heng et al., Placenta 57:320, 2017
  • a soluble NKG2D see, e.g., Cosman et al., Immunity 14(2):123- 133, 2001; Costa et al., Front. Immunol., Vol.9, Article 1150, May 29, 2018; doi: 10.3389/fimmu.2018.01150
  • a soluble NKp30 see, e.g., Costa et al., Front.
  • a soluble NKp44 see, e.g., those described in Costa et al., Front. Immunol., Vol.9, Article 1150, May 29, 2018; doi: 10.3389/fimmu.2018.01150
  • a soluble NKp46 see, e.g., Mandelboim et al., Nature 409:1055-1060, 2001; Costa et al., Front.
  • a soluble DNAM1 see, e.g., those described in Costa et al., Front. Immunol., Vol.9, Article 1150, May 29, 2018; doi: 10.3389/fimmu.2018.01150
  • a scMHCI see, e.g., those described in Washburn et al., PLoS One 6(3):e18439, 2011
  • a scMHCII see, e.g., those described in Bishwajit et al., Cellular Immunol.170(1):25-33, 1996)
  • a scTCR see, e.g., those described in Weber et al., Nature 356(6372):793-796, 1992
  • a soluble CD155 see, e.g., those described in Tahara-Hanaoka et al
  • a multi-chain chimeric polypeptide includes: 1) a first chimeric polypeptide that includes a first domain of a pair of affinity domains, and 2) a second chimeric polypeptide that includes a second domain of a pair of affinity domains such that the first chimeric polypeptide and the second chimeric polypeptide associate through the binding of the first domain and the second domain of the pair of affinity domains.
  • the pair of affinity domains is a sushi domain from an alpha chain of human IL-15 receptor (IL-15R ⁇ ) and a soluble IL-15.
  • a sushi domain also known as a short consensus repeat or type 1 glycoprotein motif, is a common motif in protein-protein interaction.
  • Sushi domains have been identified on a number of protein-binding molecules, including complement components C1r, C1s, factor H, and C2m, as well as the nonimmunologic molecules factor XIII and ⁇ 2-glycoprotein.
  • a typical Sushi domain has approximately 60 amino acid residues and contains four cysteines (Ranganathan, Pac. Symp Biocomput.2000:155-67). The first cysteine can form a disulfide bond with the third cysteine, and the second cysteine can form a disulfide bridge with the fourth cysteine.
  • the soluble IL-15 has a D8N or D8A amino acid substitution.
  • one member of the pair of affinity domains is an alpha chain of human IL-15 receptor (IL-15R ⁇ )
  • the human IL- 15R ⁇ is a mature full-length IL-15R ⁇ .
  • the pair of affinity domains is barnase and barnstar.
  • the pair of affinity domains is a PKA and an AKAP.
  • the pair of affinity domains is an adapter/docking tag module based on mutated RNase I fragments (Rossi, Proc Natl Acad Sci USA.103:6841-6846, 2006; Sharkey et al., Cancer Res.68:5282-5290, 2008; Rossi et al., Trends Pharmacol Sci.33:474-481, 2012) or SNARE modules based on interactions of the proteins syntaxin, synaptotagmin, synaptobrevin, and SNAP25 (Deyev et al., Nat Biotechnol.1486-1492, 2003).
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide includes a first domain of a pair of affinity domains and a second chimeric polypeptide of the multi-chain chimeric polypeptide includes a second domain of a pair of affinity domains, wherein the first domain of the pair of affinity domains and the second domain of the pair of affinity domains bind to each other with a dissociation equilibrium constant (KD) of less than 1 x 10 -7 M, less than 1 x 10 -8 M, less than 1 x 10 -9 M, less than 1 x 10 -10 M, less than 1 x 10 -11 M, less than 1 x 10 -12 M, or less than 1 x 10 -13 M.
  • KD dissociation equilibrium constant
  • the first domain of the pair of affinity domains and the second domain of the pair of affinity domains bind to each other with a KD of about 1 x 10 -4 M to about 1 x 10 -6 M, about 1 x 10 -5 M to about 1 x 10 -7 M, about 1 x 10 -6 M to about 1 x 10 -8 M, about 1 x 10 -7 M to about 1 x 10 -9 M, about 1 x 10 -8 M to about 1 x 10 -10 M, about 1 x 10 -9 M to about 1 x 10 -11 M, about 1 x 10 -10 M to about 1 x 10 -12 M, about 1 x 10 -11 M to about 1 x 10 -13 M, about 1 x 10 -4 M to about 1 x 10 -5 M, about 1 x 10 -5 M to about 1 x 10- 6 M, about 1 x 10 -6 M to about 1 x 10 -7 M, about 1 x 10 -7 M to about 1 x 10 -8 M, about 1 x 10 -4
  • any of a variety of different methods known in the art can be used to determine the K D value of the binding of the first domain of the pair of affinity domains and the second domain of the pair of affinity domains (e.g., an electrophoretic mobility shift assay, a filter binding assay, surface plasmon resonance, and a biomolecular binding kinetics assay, etc.).
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide includes a first domain of a pair of affinity domains and a second chimeric polypeptide of the multi-chain chimeric polypeptide includes a second domain of a pair of affinity domains, wherein the first domain of the pair of affinity domains, the second domain of the pair of affinity domains, or both is about 10 to 100 amino acids in length.
  • a first domain of a pair of affinity domains, a second domain of a pair of affinity domains, or both can be about 10 to 100 amino acids in length, about 15 to 100 amino acids in length, about 20 to 100 amino acids in length, about 25 to 100 amino acids in length, about 30 to 100 amino acids in length, about 35 to 100 amino acids in length, about 40 to 100 amino acids in length, about 45 to 100 amino acids in length, about 50 to 100 amino acids in length, about 55 to 100 amino acids in length, about 60 to 100 amino acids in length, about 65 to 100 amino acids in length, about 70 to 100 amino acids in length, about 75 to 100 amino acids in length, about 80 to 100 amino acids in length, about 85 to 100 amino acids in length, about 90 to 100 amino acids in length, about 95 to 100 amino acids in length, about 10 to 95 amino acids in length, about 10 to 90 amino acids in length, about 10 to 85 amino acids in length, about 10 to 80 amino acids in length, about 10 to 75 amino acids in length, about 10 to 70 amino acids in length, about 10 to 65 amino acids in length
  • a first domain of a pair of affinity domains, a second domain of a pair of affinity domains, or both is about 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 amino acids in length.
  • any of the first and/or second domains of a pair of affinity domains disclosed herein can include one or more additional amino acids (e.g., 1, 2, 3, 5, 6, 7, 8, 9, 10, or more amino acids) at its N-terminus and/or C-terminus, so long as the function of the first and/or second domains of a pair of affinity domains remains intact.
  • a sushi domain from an alpha chain of human IL-15 receptor can include one or more additional amino acids at the N-terminus and/or the C-terminus, while still retaining the ability to bind to a soluble IL-15.
  • a soluble IL-15 can include one or more additional amino acids at the N-terminus and/or the C-terminus, while still retaining the ability to bind to a sushi domain from an alpha chain of human IL-15 receptor (IL-15R ⁇ ).
  • a non-limiting example of a sushi domain from an alpha chain of IL-15 receptor alpha can include a sequence that is at least 70% identical, at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical to ITCPPPMSVEHADIWVKSYSLYSRERYICNSGFKRKAGTSSLTECVLNKATNVAH WTTPSLKCIR (SEQ ID NO: 113).
  • a sushi domain from an alpha chain of IL-15R ⁇ can be encoded by a nucleic acid including ATTACATGCCCCCCTCCCATGAGCGTGGAGCACGCCGACATCTGGGTGAAGAG CTATAGCCTCTACAGCCGGGAGAGGTATATCTGTAACAGCGGCTTCAAGAGGA AGGCCGGCACCAGCAGCCTCACCGAGTGCGTGCTGAATAAGGCTACCAACGT GGCTCACTGGACAACACCCTCTTTAAAGTGCATCCGG (SEQ ID NO: 114).
  • a soluble IL-15 can include a sequence that is at least 70% identical, at least 75% identical, at least 80% identical, at least 85% identical, at least 90% identical, at least 95% identical, at least 99% identical, or 100% identical to NWVNVISDLKKIEDLIQSMHIDATLYTESDVHPSCKVTAMKCFLLELQVISLESGD ASIHDTVENLIILANNSLSSNGNVTESGCKECEELEEKNIKEFLQSFVHIVQMFINT S (SEQ ID NO: 115).
  • a soluble IL-15 can be encoded by a nucleic acid including the sequence of AACTGGGTGAACGTCATCAGCGATTTAAAGAAGATCGAAGATTTAATTCAGTC CATGCATATCGACGCCACTTTATACACAGAATCCGACGTGCACCCCTCTTGTAA GGTGACCGCCATGAAATGTTTTTTACTGGAGCTGCAAGTTATCTCTTTAGAGAG CGGAGACGCTAGCATCCACGACACCGTGGAGAATTTAATCATTTTAGCCAATA ACTCTTTATCCAGCAACGGCAACGTGACAGAGTCCGGCTGCAAGGAGTGCGA AGAGCTGGAGGAGAAGAACATCAAGGAGTTTCTGCAATCCTTTGTGCACATTG TCCAGATGTTCATCAATACCTCC (SEQ ID NO: 116).
  • a single-chain chimeric polypeptide comprises a signal sequence at its N-terminal end.
  • a multi-chain chimeric polypeptide includes a first chimeric polypeptide that includes a signal sequence at its N- terminal end.
  • a multi-chain chimeric polypeptide includes a second chimeric polypeptide that includes a signal sequence at its N-terminal end.
  • both the first chimeric polypeptide of a multi-chain chimeric polypeptide and a second chimeric polypeptide of the multi-chain chimeric polypeptide include a signal sequence.
  • a signal sequence is an amino acid sequence that is present at the N-terminus of a number of endogenously produced proteins that directs the protein to the secretory pathway (e.g., the protein is directed to reside in certain intracellular organelles, to reside in the cell membrane, or to be secreted from the cell).
  • Signal sequences are heterogeneous and differ greatly in their primary amino acid sequences. However, signal sequences are typically 16 to 30 amino acids in length and include a hydrophilic, usually positively charged N-terminal region, a central hydrophobic domain, and a C-terminal region that contains the cleavage site for signal peptidase.
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide includes a signal sequence having an amino acid sequence MKWVTFISLLFLFSSAYS (SEQ ID NO: 117).
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide includes a signal sequence encoded by the nucleic acid sequence ATGAAATGGGTGACCTTTATTTCTTTACTGTTCCTCTTTAGCAGCGCCTACTCC (SEQ ID NO: 118), ATGAAGTGGGTCACATTTATCTCTTTACTGTTCCTCTTCTCCAGCGCCTACAGC (SEQ ID NO: 119), or ATGAAATGGGTGACCTTTATTTCTTTACTGTTCCTCTTTAGCAGCGCCTACTCC (SEQ ID NO: 120).
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide includes a signal sequence having an amino acid sequence MKCLLYLAFLFLGVNC (SEQ ID NO: 121).
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide includes a signal sequence having an amino acid sequence MGQIVTMFEALPHIIDEVINIVIIVLIIITSIKAVYNFATCGILALVSFLFLAGRSCG (SEQ ID NO: 122).
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide includes a signal sequence having an amino acid sequence: MPNHQSGSPTGSSDLLLSGKKQRPHLALRRKRRREMRKINRKVRRMNLAPIKEK TAWQHLQALISEAEEVLKTSQTPQNSLTLFLALLSVLGPPVTG (SEQ ID NO: 123).
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide includes a signal sequence having an amino acid sequence MDSKGSSQKGSRLLLLLVVSNLLLCQGVVS (SEQ ID NO: 124).
  • SEQ ID NO: 124 amino acid sequence MDSKGSSQKGSRLLLLLVVSNLLLCQGVVS
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide includes a signal sequence that is about 10 to 100 amino acids in length.
  • a signal sequence can be about 10 to 100 amino acids in length, about 15 to 100 amino acids in length, about 20 to 100 amino acids in length, about 25 to 100 amino acids in length, about 30 to 100 amino acids in length, about 35 to 100 amino acids in length, about 40 to 100 amino acids in length, about 45 to 100 amino acids in length, about 50 to 100 amino acids in length, about 55 to 100 amino acids in length, about 60 to 100 amino acids in length, about 65 to 100 amino acids in length, about 70 to 100 amino acids in length, about 75 to 100 amino acids in length, about 80 to 100 amino acids in length, about 85 to 100 amino acids in length, about 90 to 100 amino acids in length, about 95 to 100 amino acids in length, about 10 to 95 amino acids in length, about 10 to 90 amino acids in length, about 10 to 85 amino acids in length, about 10 to 80 amino acids in length, about 10 to 75 amino acids in length, about 10 to 70 amino acids in length, about 10 to 65 amino acids in length, about 10 to 60 amino acids in length, about 10 to 55 amino acids in length, about 10 to to 100
  • a signal sequence is about 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 amino acids in length.
  • any of the signal sequences disclosed herein can include one or more additional amino acids (e.g., 1, 2, 3, 5, 6, 7, 8, 9, 10, or more amino acids) at its N-terminus and/or C-terminus, so long as the function of the signal sequence remains intact.
  • a signal sequence having the amino acid sequence MKCLLYLAFLFLGVNC can include one or more additional amino acids at the N-terminus or C-terminus, while still retaining the ability to direct the a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide, to the secretory pathway.
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide includes a signal sequence that directs the multi-chain chimeric polypeptide into the extracellular space. Such embodiments are useful in producing single-chain or multi-chain chimeric polypeptides that are relatively easy to be isolated and/or purified.
  • Peptide Tags In some embodiments, a single-chain chimeric polypeptide includes a peptide tag (e.g., at the N-terminal end or the C-terminal end of the chimeric polypeptide).
  • a multi-chain chimeric polypeptide includes a first chimeric polypeptide that includes a peptide tag (e.g., at the N-terminal end or the C-terminal end of the first chimeric polypeptide).
  • a multi-chain chimeric polypeptide includes a second chimeric polypeptide that includes a peptide tag (e.g., at the N-terminal end or the C-terminal end of the second chimeric polypeptide).
  • both the first chimeric polypeptide of a multi-chain chimeric polypeptide and a second chimeric polypeptide of the multi-chain chimeric polypeptide include a peptide tag.
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single- chain chimeric polypeptide includes two or more peptide tags.
  • Exemplary peptide tags that can be included in a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide include, without limitation, AviTag (GLNDIFEAQKIEWHE; SEQ ID NO: 126), a calmodulin-tag (KRRWKKNFIAVSAANRFKKISSSGAL; SEQ ID NO: 127), a polyglutamate tag (EEEEEE; SEQ ID NO: 128), an E-tag (GAPVPYPDPLEPR; SEQ ID NO: 129), a FLAG-tag (DYKDDDDK; SEQ ID NO: 130), an HA-tag, a peptide from hemagglutinin (YPYDVPDYA; SEQ ID NO: 131), a his-tag (HHHHH (SEQ ID NO: 132); HHHHHH (SEQ ID NO: 133);
  • tissue factor protein is a peptide tag.
  • Peptide tags that can be included in a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide can be used in any of a variety of applications related to the multi-chain or single-chain chimeric polypeptide, respectively.
  • a peptide tag can be used in the purification of a multi-chain or single-chain chimeric polypeptide.
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide e.g., a recombinantly expressed first chimeric polypeptide
  • a second chimeric polypeptide of the multi-chain chimeric polypeptide e.g., a recombinantly expressed second chimeric polypeptide
  • a single-chain chimeric polypeptide can include a myc tag; the multi-chain chimeric polypeptide that includes the myc-tagged first chimeric polypeptide, the myc-tagged second chimeric polypeptide, or both, or the myc-tagged single-chain chimeric polypeptide can be purified using an antibody that recognizes the myc tag(s).
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide e.g., a recombinantly expressed first chimeric polypeptide
  • a second chimeric polypeptide of the multi-chain chimeric polypeptide e.g., a recombinantly expressed second chimeric polypeptide
  • a single-chain chimeric polypeptide can include a histidine tag; the multi-chain chimeric polypeptide that includes the histidine-tagged first chimeric polypeptide, the histidine-tagged second chimeric polypeptide, or both, or the histidine- tagged single-chain chimeric polypeptide can be purified using a nickel or cobalt chelate.
  • a peptide tag is removed from the first chimeric polypeptide and/or the second chimeric polypeptide of the multi-chain chimeric polypeptide, or the single- chain chimeric polypeptide after purification. In some embodiments, a peptide tag is not removed from the first chimeric polypeptide and/or the second chimeric polypeptide of the multi-chain chimeric polypeptide, or the single-chain chimeric polypeptide, after purification.
  • Peptide tags that can be included in a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide can be used, for example, in immunoprecipitation of the multi-chain chimeric polypeptide or single-chain chimeric polypeptide, respectively, imaging of the multi-chain chimeric polypeptide or single- chain chimeric polypeptide, respectively (e.g., via Western blotting, ELISA, flow cytometry, and/or immunocytochemistry), and/or solubilization of the multi-chain chimeric polypeptide or single-chain chimeric polypeptide, respectively.
  • a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide includes a peptide tag that is about 10 to 100 amino acids in length.
  • a peptide tag can be about 10 to 100 amino acids in length, about 15 to 100 amino acids in length, about 20 to 100 amino acids in length, about 25 to 100 amino acids in length, about 30 to 100 amino acids in length, about 35 to 100 amino acids in length, about 40 to 100 amino acids in length, about 45 to 100 amino acids in length, about 50 to 100 amino acids in length, about 55 to 100 amino acids in length, about 60 to 100 amino acids in length, about 65 to 100 amino acids in length, about 70 to 100 amino acids in length, about 75 to 100 amino acids in length, about 80 to 100 amino acids in length, about 85 to 100 amino acids in length, about 90 to 100 amino acids in length, about 95 to 100 amino acids in length, about 10 to 95 amino acids in length, about 10 to 90 amino acids in length, about 10 to 85 amino acids in length, about 10 to 80 amino acids in length, about 10 to 75 amino acids in length, about 10 to 70 amino acids in length, about 10 to 65 amino acids in length, about 10 to 60 amino acids in length, about 10 to 55 amino acids in length, about
  • a peptide tag is about 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 amino acids in length.
  • Peptide tags included in a first chimeric polypeptide of a multi-chain chimeric polypeptide, a second chimeric polypeptide of the multi-chain chimeric polypeptide, or both, or a single-chain chimeric polypeptide can be of any suitable length.
  • peptide tags can be 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more amino acids in length.
  • a single-chain or multi-chain chimeric polypeptide includes two or more peptide tags
  • the two or more peptide tags can be of the same or different lengths.
  • any of the peptide tags disclosed herein may include one or more additional amino acids (e.g., 1, 2, 3, 5, 6, 7, 8, 9, 10, or more amino acids) at the N-terminus and/or C-terminus, so long as the function of the peptide tag remains intact.
  • a myc tag having the amino acid sequence EQKLISEEDL can include one or more additional amino acids (e.g., at the N-terminus and/or the C- terminus of the peptide tag), while still retaining the ability to be bound by an antibody (e.g., 9E10).
  • an antibody e.g. 9E10
  • Exemplary Embodiments of Single-Chain Chimeric Polypeptides- Type A in some embodiments of any of the single-chain chimeric polypeptides described herein, the first target-binding domain and/or the second target-binding domain can independently bind specifically to CD3 (e.g., human CD3) or CD28 (e.g., human CD28).
  • the first target-binding domain binds specifically to CD3 (e.g., human CD3) and the second target-binding domain binds specifically to CD28 (e.g., human CD28). In some embodiments, the first target-binding domain binds specifically to CD28 (e.g., human CD28) and the second target-binding domain binds specifically to CD3 (e.g., human CD3). In some embodiments of these single-chain chimeric polypeptides, the first target- binding domain and the soluble tissue factor domain directly abut each other.
  • the single-chain chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain.
  • the soluble tissue factor domain and the second target-binding domain directly abut each other.
  • the single-chain chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the second target-binding domain.
  • one or both of the first target-binding domain and the second target-binding domain is an antigen- binding domain.
  • the first target-binding domain and the second target-binding domain are each an antigen- binding domain (e.g., any of the exemplary antigen-binding domains described herein).
  • the antigen-binding domain includes a scFv or a single domain antibody.
  • a non-limiting example of an scFv that binds specifically to CD3 can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QIVLTQSPAIMSASPGEKVTMTCSASSSVSYMNWYQQKSGTSPKRWIYDTSKLA SGVPAHFRGSGSGTSYSLTISGMEAEDAATYYCQQWSSNPFTFGSGTKLEINRGG GGSGGGGSGGGGSQVQLQQSGAELARPGASVKMSCKASGYTFTRYTMHWVKQ RPGQGLEWIGYINPSRGYTNYNQKFKDKATLTTDKSSSTAYMQLSSLTSEDSAV YYCARYYDDHYCLDYWGQGTTLTVSS (S
  • an scFv that binds specifically to CD3 can be encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGATCGTGCTGACCCAAAGCCCCGCCATCATGAGCGCTAGCCCCGGTGAGA AGGTGACCATGACATGCTCCGCTTCCAGCTCCGTGTCCTACATGAACTGGTAT CAGCAGAAAAGCGGAACCAGCCCCAAAAGGTGGATCTACGACACCAGCAAG CTGGCCTCCGGAGTGCCCGCTCATTTCCGGGGCTCTGGATCCGGCACCAGCTA CTCTTTAACCATTTCCGGCATGGAAGCTGAAGACGCTGCCACCTACTATTGCC AGCAATGGAGCAGCAACCCCTTCACATTCGGATCTGGCACCAAGC
  • a non-limiting example of an scFv that binds specifically to CD28 can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: VQLQQSGPELVKPGASVKMSCKASGYTFTSYVIQWVKQKPGQGLEWIGSINPYN DYTKYNEKFKGKATLTSDKSSITAYMEFSSLTSEDSALYYCARWGDGNYWGRG TTLTVSSGGGGSGGGGSGGGGSDIEMTQSPAIMSASLGERVTMTCTASSSVSSSY FHWYQQKPGSSPKLCIYSTSNLASGVPPRFSGSGSTSYSLTISSMEAEDAATYFCH QYHRSPTFGGGTKLETKR (SEQ ID NO: 153).
  • an scFv that binds specifically to CD28 can be encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GTCCAGCTGCAGCAGAGCGGACCCGAACTCGTGAAACCCGGTGCTTCCGTGA AAATGTCTTGTAAGGCCAGCGGATACACCTTCACCTCCTATGTGATCCAGTGG GTCAAACAGAAGCCCGGACAAGGTCTCGAGTGGATCGGCAGCATCAACCCTT ACAACGACTATACCAAATACAACGAGAAGTTTAAGGGAAAGGCTACTTTAAC CTCCGACAAAAGCTCCATCACAGCCTACATGGAGTTCAGCTCTTTAACATCCG AGGACAGCGCTCTGTACTATTGCCCGGTGGGGCGACGGCAATT
  • the first target- binding domain and/or the second target-binding domain is a soluble receptor (e.g., a soluble CD28 receptor or a soluble CD3 receptor).
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • a single-chain chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QIVLTQSPAIMSASPGEKVTMTCSASSSVSYMNWYQQKSGTSPKRWIYDTSKLA SGVPAHFRGSGSGTSYSLTISGMEAEDAATYYCQQWSSNPFTFGSGTKLEINRGG GGSGGGGSGGGGSQVQLQQSGAELARPGASVKMSCKASGYTFTRYTMHWVKQ RPGQGLEWIGYINPSRGYTNYNQKFKDKATLTTDKSSSTAYMQLSSLTSEDSAV YYCARYYDDHYCLDYWGQGTTLTVSSSGTTNTVAAYNLT
  • a single-chain chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGATCGTGCTGACCCAAAGCCCCGCCATCATGAGCGCTAGCCCCGGTGAGA AGGTGACCATGACATGCTCCGCTTCCAGCTCCGTGTCCTACATGAACTGGTAT CAGCAGAAAAGCGGAACCAGCCCCAAAAGGTGGATCTACGACACCAGCAAG CTGGCCTCCGGAGTGCCCGCTCATTTCCGGGGCTCTGGATCCGGCACCAGCTA CTCTTTAACCATTTCCGGCATGGAAGCTGAAGACGCTGCCACCTACTATTGCC AGCAATGGAGCAGCAACCCCTTCACATTCGGATCTGGCACCAAGCTCGAAAT
  • a single-chain chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSQIVLTQSPAIMSASPGEKVTMTCSASSSVSYMNWYQQ KSGTSPKRWIYDTSKLASGVPAHFRGSGSGTSYSLTISGMEAEDAATYYCQQWS SNPFTFGSGTKLEINRGGGGSGGGGSGGGGSQVQLQQSGAELARPGASVKMSCK ASGYTFTRYTMHWVKQRPGQGLEWIGYINPSRGYTNYNQKFKDKATLTTDKSS STAYMQLSSLTSEDSAVYYCARYYDDHYCLDYWGQGTTLTV
  • a single-chain chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCTTATTATTTTTATTCAGCTCCGCCTATTCC CAGATCGTGCTGACCCAAAGCCCCGCCATCATGAGCGCTAGCCCCGGTGAGA AGGTGACCATGACATGCTCCGCTTCCAGCTCCGTGTCCTACATGAACTGGTAT CAGCAGAAAAGCGGAACCAGCCCCAAAAGGTGGATCTACGACACCAGCAAG CTGGCCTCCGGAGTGCCCGCTCATTTCCGGGGCTCTGGATCCGGCACCAGCTA CTCTTTAACCATTTCCGGCATGGAAGCTGAAGACGCTGCCACCTACTATTGCC A
  • the first target-binding domain and/or the second target-binding domain can independently bind specifically to an IL-2 receptor (e.g., human IL-2 receptor).
  • an IL-2 receptor e.g., human IL-2 receptor
  • the first target- binding domain and the soluble tissue factor domain directly abut each other.
  • the single-chain chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain.
  • the soluble tissue factor domain and the second target-binding domain directly abut each other.
  • the single-chain chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the second target-binding domain.
  • the first target- binding domain and the second target-binding domain is a soluble human IL-2 protein.
  • a non-limiting example of an IL-2 protein that binds specifically to an IL-2 receptor can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: APTSSSTKKTQLQLEHLLLDLQMILNGINNYKNPKLTRMLTFKFYMPKKATELK HLQCLEEELKPLEEVLNLAQSKNFHLRPRDLISNINVIVLELKGSETTFMCEYADE TATIVEFLNRWITFCQSIISTLT (SEQ ID NO: 78).
  • an IL-2 protein that binds specifically to an IL-2 receptor can be encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GCACCTACTTCAAGTTCTACAAAGAAAACACAGCTACAACTGGAGCATTTAC TGCTGGATTTACAGATGATTTTGAATGGAATTAATAATTACAAGAATCCCAA ACTCACCAGGATGCTCACATTTAAGTTTTACATGCCCAAGAAGGCCACAGAA CTGAAACATCTTCAGTGTCTAGAAGAAGAACTCAAACCTCTGGAGGAAGTGC TAAATTTAGCTCAAAGCAAAAACTTTCACTTAAGACCCAGGGACTTAATCAG CAATATCAACGTAATAGTTCTGGAACTAAAGGGATCTGAAACAAC
  • an IL-2 protein that binds specifically to an IL-2 receptor can be encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GCCCCCACCTCCTCCTCCACCAAGAAGACCCAGCTGCAGCTGGAGCATTTAC TGCTGGATTTACAGATGATTTTAAACGGCATCAACAACTACAAGAACCCCAA GCTGACTCGTATGCTGACCTTCAAGTTCTACATGCCCAAGAAGGCCACCGAG CTGAAGCATTTACAGTGTTTAGAGGAGGAGCTGAAGCCCCTCGAGGAGGTGC TGAATTTAGCCCAGTCCAAGAATTTCCATTTAAGGCCCCGGGATTTAATCAGC AACATCAACGTTTTAAAGGGCTCCGAGACC
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • a single-chain chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: APTSSSTKKTQLQLEHLLLDLQMILNGINNYKNPKLTRMLTFKFYMPKKATELK HLQCLEEELKPLEEVLNLAQSKNFHLRPRDLISNINVIVLELKGSETTFMCEYADE TATIVEFLNRWITFCQSIISTLTSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQV YTVQISTKSGDWKSKCFYTTDTECDLTDEIVKDV
  • a single-chain chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GCCCCCACCTCCTCCTCCACCAAGAAGACCCAGCTGCAGCTGGAGCATTTAC TGCTGGATTTTAAACGGCATCAACAACTACAAGAACCCCAA GCTGACTCGTATGCTGACCTTCAAGTTCTACATGCCCAAGAAGGCCACCGAG CTGAAGCATTTACAGTGTTTAGAGGAGGAGCTGAAGCCCCTCGAGGAGGTGC TGAATTTAGCCCAGTCCAAGAATTTCCATTTAAGGCCCCGGGATTTAATCAGC AACATCAACGTTTTAAAGGGCTCCGAGACCACCTTCATGT GCG
  • a single-chain chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSAPTSSSTKKTQLQLEHLLLDLQMILNGINNYKNPKLTR MLTFKFYMPKKATELKHLQCLEEELKPLEEVLNLAQSKNFHLRPRDLISNINVIVL ELKGSETTFMCEYADETATIVEFLNRWITFCQSIISTLTSGTTNTVAAYNLTWKST NFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVKDVKQTY LARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPTIQSFEQVG
  • a single-chain chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CGCCCCCACCTCCTCCTCCACCAAGAAGACCCAGCTGCAGCTGGAGCATTTA CTGCTGGATTTACAGATGATTTTAAACGGCATCAACAACTACAAGAACCCCA AGCTGACTCGTATGCTGACCTTCAAGTTCTACATGCCCAAGAAGGCCACCGA GCTGAAGCATTTACAGTGTTTAGAGGAGGAGCTGAAGCCCCTCGAGGAGGTG CTGAATTTAGCCCAGTCCAAGAATTTCCATTTAAGGCCCCGGGATTTAATCAG CAA
  • the first target-binding domain and/or the second target-binding domain can independently bind specifically to an IL-15 receptor (e.g., a human IL-15 receptor).
  • an IL-15 receptor e.g., a human IL-15 receptor
  • the first target- binding domain and the soluble tissue factor domain directly abut each other.
  • the single-chain chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain.
  • the soluble tissue factor domain and the second target-binding domain directly abut each other.
  • the single-chain chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the second target-binding domain.
  • the first target- binding domain and the second target-binding domain is a soluble human IL-15 protein.
  • a non-limiting example of an IL-15 protein that binds specifically to an IL-15 receptor can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: NWVNVISDLKKIEDLIQSMHIDATLYTESDVHPSCKVTAMKCFLLELQVISLESG DASIHDTVENLIILANNSLSSNGNVTESGCKECEELEEKNIKEFLQSFVHIVQMFIN TS (SEQ ID NO: 82).
  • an IL-15 protein that binds specifically to an IL-15 receptor can be encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: AACTGGGTGAACGTGATCAGCGATTTAAAGAAGATCGAGGATTTAATCCAGA GCATGCACATCGACGCCACTCTGTACACTGAGAGCGACGTGCACCCTAGCTG CAAGGTGACTGCCATGAAGTGCTTTTTACTGGAGCTGCAAGTTATCTTTAG AGAGCGGCGATGCCAGCATCCACGACACTGTGGAGAATTTAATCATTTTAGC CAACAACTCTTTAAGCAGCAACGGCAACGTGACAGAGAGCGGCTGCAAGGA GTGCGAGGAGCTGGAGGAGAAGAACATCAAGGAGTTTTTACAGA
  • an IL-15 protein that binds specifically to an IL-15 receptor can be encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: AACTGGGTGAACGTCATCAGCGATTTAAAGAAGATCGAAGATTTAATTCAGT CCATGCATATCGACGCCACTTTATACACAGAATCCGACGTGCACCCCTCTTGT AAGGTGACCGCCATGAAATGTTTTTTACTGGAGCTGCAAGTTATCTTTAGA GAGCGGAGACGCTAGCATCCACGACACCGTGGAGAATTTAATCATTTTAGCC AATAACTCTTTATCCAGCAACGGCAACGTGACAGAGTCCGGCTGCAAGGAGT GCGAAGAGCTGGAGGAGAAGAACATCAAGGAGTTTCTGCA
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • a single-chain chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: NWVNVISDLKKIEDLIQSMHIDATLYTESDVHPSCKVTAMKCFLLELQVISLESG DASIHDTVENLIILANNSLSSNGNVTESGCKECEELEEKNIKEFLQSFVHIVQMFIN TSSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYT TDTECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSAGE
  • a single-chain chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: AACTGGGTGAACGTGATCAGCGATTTAAAGAAGATCGAGGATTTAATCCAGA GCATGCACATCGACGCCACTCTGTACACTGAGAGCGACGTGCACCCTAGCTG CAAGGTGACTGCCATGAAGTGCTTTTTACTGGAGCTGCAAGTTATCTTTAG AGAGCGGCGATGCCAGCATCCACGACACTGTGGAGAATTTAATCATTTTAGC CAACAACTCTTTAAGCAGCAACGGCAACGTGACAGAGAGCGGCTGCAAGGA GTGCGAGGAGCTGGAGGAGAAGAACATCAAGGAGTTTTTACAGAGCTTCGTG CACAT
  • a single-chain chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSNWVNVISDLKKIEDLIQSMHIDATLYTESDVHPSCKVT AMKCFLLELQVISLESGDASIHDTVENLIILANNSLSSNGNVTESGCKECEELEEK NIKEFLQSFVHIVQMFINTSSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYT VQISTKSGDWKSKCFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSA GEPLYENSPEFTPYLETNLGQPTIQSFEQVGTKVNVTVEDERTLVRRNNTFLSLRD V
  • a single-chain chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CAACTGGGTGAACGTGATCAGCGATTTAAAGAAGATCGAGGATTTAATCCAG AGCATGCACATCGACGCCACTCTGTACACTGAGAGCGACGTGCACCCTAGCT GCAAGGTGACTGCCATGAAGTGCTTTTTACTGGAGCTGCAAGTTATCTTTA GAGAGCGGCGATGCCAGCATCCACGACACTGTGGAGAATTTAATCATTTTAG CCAACAACTCTTTAAGCAGCAACGGCAACGTGACAGAGAGCGGCTGCAAGG AGT
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to a receptor of IL-18 or a receptor of IL-12.
  • the first target-binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • one or both of the first target-binding domain and the second target-binding domain is an agonistic antigen-binding domain.
  • the first target-binding domain and the second target-binding domain are each agonistic antigen-binding domains.
  • the antigen-binding domain includes a scFv or single-domain antibody.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble IL-15 or a soluble IL-18.
  • the first target-binding domain and the second target-binding domain are each independently a soluble IL-15 or a soluble IL-18. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain both bind specifically to a receptor of IL-18 or a receptor of IL-12. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain bind specifically to the same epitope. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain include the same amino acid sequence.
  • the first target- binding domain binds specifically to a receptor for IL-12, and the second target-binding domain binds specifically to a receptor for IL-18. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain binds specifically to a receptor for IL-18, and the second target-binding domain bind specifically to a receptor for IL-12. In some embodiments of these multi-chain chimeric polypeptides, the first target- binding domain includes a soluble IL-18 (e.g., a soluble human IL-18).
  • the soluble human IL-18 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: YFGKLESKLSVIRNLNDQVLFIDQGNRPLFEDMTDSDCRDNAPRTIFIISMYKDSQ PRGMAVTISVKCEKISTLSCENKIISFKEMNPPDNIKDTKSDIIFFQRSVPGHDNKM QFESSSYEGYFLACEKERDLFKLILKKEDELGDRSIMFTVQNED (SEQ ID NO: 109).
  • the soluble human IL-18 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: TACTTCGGCAAACTGGAATCCAAGCTGAGCGTGATCCGGAATTTAAACGACC AAGTTCTGTTTATCGATCAAGGTAACCGGCCTCTGTTCGAGGACATGACCGAC TCCGATTGCCGGGACAATGCCCCCCGGACCATCTTCATTATCTCCATGTACAA GGACAGCCAGCCCCGGGGCATGGCTGTGACAATTAGCGTGAAGTGTGAGAA AATCAGCACTTTATCTTGTGAGAACAAGATCATCTCCTTTAAGGAAATGAACC CCCCCGATAACATCAAGGACACCAAGTCCGATAACATCAAGGACACCAAGTCCGATAACATCAAGGACA
  • the second target-binding domain includes a soluble IL-12 (e.g., a soluble human IL-12).
  • the soluble human IL-15 includes a sequence of soluble human IL-12 ⁇ (p40) and a sequence of soluble human IL- 12 ⁇ (p35).
  • the soluble IL-15 human IL-15 further includes a linker sequence (e.g., any of the exemplary linker sequences described herein) between the sequence of soluble IL-12 ⁇ (p40) and the sequence of soluble human IL-12 ⁇ (p35).
  • the linker sequence comprises GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the sequence of soluble human IL-12 ⁇ (p40) comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IWELKKDVYVVELDWYPDAPGEMVVLTCDTPEEDGITWTLDQSSEVLGSGKTLT IQVKEFGDAGQYTCHKGGEVLSHSLLLLHKKEDGIWSTDILKDQKEPKNKTFLR CEAKNYSGRFTCWWLTTISTDLTFSVKSSRGSSDPQGVTCGAATLSAERVRGDN KEYEYSVECQEDSACPAAEE
  • the soluble human IL-12 ⁇ (p40) is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTTGGGAACTGAAGAAGGACGTCTACGTGGTCGAACTGGACTGGTATCCCG ATGCTCCCGGCGAAATGGTGGTGCTCACTTGTGACACCCCCGAAGAAGACGG CATCACTTGGACCCTCGATCAGAGCAGCGAGGTGCTGGGCTCCGGAAAGACC CTCACAATCCAAGTTAAGGAGTTCGGAGACGCTGGCCAATACACATGCCACA AGGGAGGCGAGGTGCTCAGCCATTCCTTATTATTATTACACAAGAAGGAAGA CGGAATCTGGTCCACCGACATTTTAAAAGATCAGA
  • the soluble human IL-12 ⁇ (p35) includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: RNLPVATPDPGMFPCLHHSQNLLRAVSNMLQKARQTLEFYPCTSEEIDHEDITKD KTSTVEACLPLELTKNESCLNSRETSFITNGSCLASRKTSFMMALCLSSIYEDLKM YQVEFKTMNAKLLMDPKRQIFLDQNMLAVIDELMQALNFNSETVPQKSSLEEPD FYKTKIKLCILLHAFRIRAVTIDRVMSYLNAS (SEQ ID NO: 80).
  • the soluble human IL-12 ⁇ (p35) is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CGTAACCTCCCCGTGGCTACCCCCGATCCCGGAATGTTCCCTTGTTTACACCA CAGCCAGAATTTACTGAGGGCCGTGAGCAACATGCTGCAGAAAGCTAGGCAG ACTTTAGAATTTTACCCTTGCACCAGCGAGGAGATCGACCATGAAGATATCA CCAAGGACAAGACATCCACCGTGGAGGCTTGTTTACCTCTGGAGCTGACAAA GAACGAGTCTTGTCTCAACTCTCGTGAAACCAGCTTCATCACAAATGGCTCTT GTTTAGCTTCCCGGAAGACCTCCT
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: YFGKLESKLSVIRNLNDQVLFIDQGNRPLFEDMTDSDCRDNAPRTIFIISMYKDSQ PRGMAVTISVKCEKISTLSCENKIISFKEMNPPDNIKDTKSDIIFFQRSVPGHDNKM QFESSSYEGYFLACEKERDLFKLILKKEDELGDRSIMFTVQNEDSGTTNTVAAYN LTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVKD VKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPT
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: TACTTCGGCAAACTGGAATCCAAGCTGAGCGTGATCCGGAATTTAAACGACC AAGTTCTGTTTATCGATCAAGGTAACCGGCCTCTGTTCGAGGACATGACCGAC TCCGATTGCCGGGACAATGCCCCGGACCATCTTCATTATCTCCATGTACAA GGACAGCCAGCCCCGGGGCATGGCTGTGACAATTAGCGTGAAGTGTGAGAA AATCAGCACTTTATCTTGTGAGAACAAGATCATCTCCTTTAAGGAAATGAACC CCCCCGATAACATCAAGGACACCAAGTCCGATATCATCTTCTTCCAGCGGTCC
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSYFGKLESKLSVIRNLNDQVLFIDQGNRPLFEDMTDSD CRDNAPRTIFIISMYKDSQPRGMAVTISVKCEKISTLSCENKIISFKEMNPPDNIKD TKSDIIFFQRSVPGHDNKMQFESSSYEGYFLACEKERDLFKLILKKEDELGDRSIM FTVQNEDSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKS KCFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSAGEPLY
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTCACATTTATCTCTTTACTGTTCCTCTTCTCCAGCGCCTACAGC TACTTCGGCAAACTGGAATCCAAGCTGAGCGTGATCCGGAATTTAAACGACC AAGTTCTGTTTATCGATCAAGGTAACCGGCCTCTGTTCGAGGACATGACCGAC TCCGATTGCCGGGACAATGCCCCCCGGACCATCTTCATTATCTCCATGTACAA GGACAGCCAGCCCCGGGGCATGGCTGTGACAATTAGCGTGAAGTGTGAGAA AATCAGCACTTTATCTTGTGAGAACAAGATCATCTCCTTTAAGGAAATGAACC
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IWELKKDVYVVELDWYPDAPGEMVVLTCDTPEEDGITWTLDQSSEVLGSGKTLT IQVKEFGDAGQYTCHKGGEVLSHSLLLLHKKEDGIWSTDILKDQKEPKNKTFLR CEAKNYSGRFTCWWLTTISTDLTFSVKSSRGSSDPQGVTCGAATLSAERVRGDN KEYEYSVECQEDSACPAAEESLPIEVMVDAVHKLKYENYTSSFFIRDIIKPDPPKN LQLKPLKNSRQVEVSWEYPDTWSTPHSYFSLTFCVQVQGKSKREKKDRVF
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTTGGGAACTGAAGAAGGACGTCTACGTGGTCGAACTGGACTGGTATCCCG ATGCTCCCGGCGAAATGGTGGTGCTCACTTGTGACACCCCCGAAGAAGACGG CATCACTTGGACCCTCGATCAGAGCAGCGAGGTGCTGGGCTCCGGAAAGACC CTCACAATCCAAGTTAAGGAGTTCGGAGACGCTGGCCAATACACATGCCACA AGGGAGGCGAGGTGCTCAGCCATTCCTTATTATTATTACACAAGAAGGAAGA CGGAATCTGGTCCACCGACATTTTAAAAGATCAGAAGGAGCCCAAGAATAAG ACCTTTAAGGT
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIWELKKDVYVVELDWYPDAPGEMVVLTCDTPEEDGI TWTLDQSSEVLGSGKTLTIQVKEFGDAGQYTCHKGGEVLSHSLLLLHKKEDGIW STDILKDQKEPKNKTFLRCEAKNYSGRFTCWWLTTISTDLTFSVKSSRGSSDPQG VTCGAATLSAERVRGDNKEYEYSVECQEDSACPAAEESLPIEVMVDAVHKLKYE NYTSSFFIRDIIKPDPPKNLQLKPLKNSRQVEVSWEYPDTWSTPHSYFSLTFC
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAATGGGTGACCTTTATTTCTTTACTGTTCCTCTTTAGCAGCGCCTACTCC ATTTGGGAACTGAAGAAGGACGTCTACGTGGTCGAACTGGACTGGTATCCCG ATGCTCCCGGCGAAATGGTGGTGCTCACTTGTGACACCCCCGAAGAAGACGG CATCACTTGGACCCTCGATCAGAGCAGCGAGGTGCTGGGCTCCGGAAAGACC CTCACAATCCAAGTTAAGGAGTTCGGAGACGCTGGCCAATACACATGCCACA AGGGAGGCGAGGTGCTCAGCCATTCCTTATTATTACACAAGAAGGAAGA CGGAATCTGGT
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to a receptor of IL-21 or to TGF- ⁇ .
  • the first target-binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble IL-21 (e.g., a soluble human IL-21 polypeptide) or a soluble TGF- ⁇ receptor (e.g., a soluble TGFR ⁇ RII receptor).
  • the first target-binding domain and the second target-binding domain are each independently a soluble IL-21 or a soluble TGF- ⁇ receptor (e.g., a soluble TGFR ⁇ RII receptor).
  • the first target-binding domain and the second target-binding domain both bind specifically to a receptor of IL-21 or to TGF- ⁇ . In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain bind specifically to the same epitope. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain include the same amino acid sequence. In some embodiments of these multi-chain chimeric polypeptides, the first target- binding domain binds specifically to a receptor for IL-21, and the second target-binding domain binds specifically to TGF- ⁇ .
  • the first target-binding domain binds specifically to TGF- ⁇ , and the second target-binding domain bind specifically to a receptor for IL-21.
  • the first target- binding domain includes a soluble IL-21 (e.g., a soluble human IL-21).
  • the soluble human IL-21 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDS (SEQ ID NO: 83).
  • the soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTA
  • the second target-binding domain includes a soluble TGF- ⁇ receptor (e.g., a soluble TGFR ⁇ RII receptor (e.g., a soluble human TGFR ⁇ RII receptor)).
  • a soluble TGFR ⁇ RII receptor e.g., a soluble human TGFR ⁇ RII receptor
  • the soluble human TGFR ⁇ RII includes a first sequence of soluble human TGFR ⁇ RII and a second sequence of soluble human TGFR ⁇ RII.
  • the soluble human TGFR ⁇ RII includes a linker disposed between the first sequence of soluble human TGFR ⁇ RII and the second sequence of soluble human TGFR ⁇ RII.
  • the linker includes the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the first sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 183).
  • the second sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 184).
  • the first sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCACGATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCT
  • the second sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTCCTCCCCACGTGCAGAAGAGCGTGAATAATGACATGATCGTGACCGATA ACAATGGCGCCGTGAAATTTCCCCAGCTGTGCAAATTCTGCGATGTGAGGTTT TCCACCTGCGACAACCAGAAGTCCTGTATGAGCAACTGCACAATCACCTCCA TCTGTGAGAAGCCTCAGGAGGTGTGCGTGGCTGTCTGGCGGAAGAATGACGA GAATATCACCCTGGAAACCGTCTGCCACGATCCCAAGCTGCCCTACCACGAT TTCATCCTGGAAGACGCC
  • the soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCACGATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCACA
  • the human TGF ⁇ RII receptor includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSD
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDSSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQ VYTVQISTKSGDWKSKCFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVEST GSAGEPLYENSPEFTPYLETNLGQPTIQSFEQVGTKVNVTVEDERTLVRRNNTFLS
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTACGAGAAGAAGCC CCAAG
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSQGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPED VETNCEWSAFSCFQKAQLKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLT CPSCDSYEKKPPKEFLERFKSLLQKMIHQHLSSRTHGSEDSSGTTNTVAAYNLTW KSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVKDVKQ TYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPTIQSF
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CCAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTC GACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCC CCGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAA GGCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGT GAGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAAACAACGAGCGGATCATCAACGT GAG
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSDECND NIIFSEEYNTSNPD
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCACGATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGGAAGAAG AAGC
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSI TSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKK PGETFFMCSC
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGAC AACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTT CAGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCACGATCACCTCC ATCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGAC GAGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACG ACTTCCCACCCCAA
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to a receptor of IL-7 or a receptor of IL-21.
  • the first target-binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble IL-21 (e.g., a soluble human IL-21 polypeptide) or a soluble IL-7 (e.g., a soluble human IL-7 polypeptide).
  • the first target-binding domain and the second target-binding domain are each independently a soluble IL-21 or a soluble IL-7.
  • the first target-binding domain and the second target-binding domain both bind specifically to a receptor of IL-21 or a receptor of IL-7.
  • the first target-binding domain and the second target-binding domain bind specifically to the same epitope. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain include the same amino acid sequence. In some embodiments of these multi-chain chimeric polypeptides, the first target- binding domain binds specifically to a receptor for IL-21, and the second target-binding domain binds specifically to a receptor for IL-7.
  • the first target-binding domain binds specifically to a receptor for IL-7, and the second target-binding domain binds specifically to a receptor for IL-21.
  • the first target- binding domain includes a soluble IL-21 (e.g., a soluble human IL-21).
  • the soluble human IL-21 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDS (SEQ ID NO: 83).
  • the soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAAGGTCAAGATCGCCACATGATTAGAATGCGTCAACTTATAGATATTGTTG ATCAGCTGAAAAATTATGTGAATGACTTGGTCCCTGAATTTCTGCCAGCTCCA GAAGATGTAGAGACAAACTGTGAGTGGTCAGCTTTTTCCTGTTTTCAGAAGG CCCAACTAAAGTCAGCAAATACAGGAAACAATGAAAGGATAATCAATGTATC AATTAAAAAGCTGAAGAGGAAACCACCTTCCACAAATGCAGGGAGAAGACA GAAACACAGACTAACATGCCCTTCATGTGATTCTTATGA
  • the soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTA
  • the sequence of soluble human IL-7 comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEH (SEQ ID NO: 79).
  • the soluble human IL-7 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGTGATATTGAAGGTAAAGATGGCAAACAATATGAGAGTGTTCTAATGG TCAGCATCGATCAATTATTGGACAGCATGAAAGAAATTGGTAGCAATTGCCT GAATAATGAATTTAACTTTTTTAAAAGACATATCTGTGATGCTAATAAGGAA GGTATGTTTTTATTCCGTGCTGCTCGCAAGTTGAGGCAATTTCTTAAAATGAA TAGCACTGGTGATTTTGATCTCCACTTATTAAAAGTTTCAGAAGGCACAACAA TACTGTTGAACTGCACTGGCCAGGTTAAAGGAAGAAAACC
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDSSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQ VYTVQISTKSGDWKSKCFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVEST GSAGEPLYENSPEFTPYLETNLGQPTIQSFEQVGTKVNVTVEDERTLVRRNNTFLS
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAAGGTCAAGATCGCCACATGATTAGAATGCGTCAACTTATAGATATTGTTG ATCAGCTGAAAAATTATGTGAATGACTTGGTCCCTGAATTTCTGCCAGCTCCA GAAGATGTAGAGACAAACTGTGAGTGGTCAGCTTTTTCCTGTTTTCAGAAGG CCCAACTAAAGTCAGCAAATACAGGAAACAATGAAAGGATAATCAATGTATC AATTAAAAAGCTGAAGAGGAAACCACCTTCCACAAATGCAGGGAGAAGACA GAAACACAGACTAACATGCCCTTCATGTGATTCTTATGAAAAAAAACCACCC AAAGAATTCC
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MGVKVLFALICIAVAEAQGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPED VETNCEWSAFSCFQKAQLKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLT CPSCDSYEKKPPKEFLERFKSLLQKMIHQHLSSRTHGSEDSSGTTNTVAAYNLTW KSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVKDVKQ TYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPTIQSFEQ
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGGGAGTGAAAGTTCTTTTTGCCCTTATTTGTATTGCTGTGGCCGAGGCCCA AGGTCAAGATCGCCACATGATTAGAATGCGTCAACTTATAGATATTGTTGATC AGCTGAAAAATTATGTGAATGACTTGGTCCCTGAATTTCTGCCAGCTCCAGAA GATGTAGAGACAAACTGTGAGTGGTCAGCTTTTTCCTGTTTTCAGAAGGCCCA ACTAAAGTCAGCAAATACAGGAAACAATGAAAGGATAATCAATGTATCAATT AAAAAGCTGAAGAGGAAACCACCTTCCACAAATGCAGGGAGAAGACAGAAA CACA
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MGVKVLFALICIAVAEADCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNN EFNFFKRHICDANKEGMFLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNC TGQVKGRKPAALGEAQPTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKIL MGTKEHITCPPPMSVEHADIWVKSYSLYSRERYICNSGFKRKAGTSSLTECVLNK ATNVAHWTTPSLKCIR (SEQ ID NO: 205).
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGGGAGTGAAAGTTCTTTTTTTGCCCTTATTTGTATTGCTGTGGCCGAGGCCGA TTGTGATATTGAAGGTAAAGATGGCAAACAATATGAGAGTGTTCTAATGGTC AGCATCGATCAATTATTGGACAGCATGAAAGAAATTGGTAGCAATTGCCTGA ATAATGAATTTAACTTTTTTAAAAGACATATCTGTGATGCTAATAAGGAAGGT ATGTTTTTATTCCGTGCTGCTCGCAAGTTGAGGCAATTTCTTAAAATGAATAG CACTGGTGATTTTGATCTCCACTTATTAAAAGTTTCAGAAGGCACAACAATAC TGTT
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to a receptor of IL-7 or a receptor of IL-21.
  • the first target-binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble IL-21 (e.g., a soluble human IL-21 polypeptide) or a soluble IL-7 (e.g., a soluble human IL-7 polypeptide).
  • the first target-binding domain and the second target-binding domain are each independently a soluble IL-21 or a soluble IL-7.
  • the first target-binding domain and the second target-binding domain both bind specifically to a receptor of IL-21 or a receptor of IL-7.
  • the first target-binding domain and the second target-binding domain bind specifically to the same epitope. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain include the same amino acid sequence. In some embodiments of these multi-chain chimeric polypeptides, the first target- binding domain binds specifically to a receptor for IL-21, and the second target-binding domain binds specifically to a receptor for IL-7.
  • the first target-binding domain binds specifically to a receptor for IL-7
  • the second target-binding domain binds specifically to a receptor for IL-21.
  • the soluble human IL-21 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDS (SEQ ID NO: 83).
  • the soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAAGGTCAAGATCGCCACATGATTAGAATGCGTCAACTTATAGATATTGTTG ATCAGCTGAAAAATTATGTGAATGACTTGGTCCCTGAATTTCTGCCAGCTCCA GAAGATGTAGAGACAAACTGTGAGTGGTCAGCTTTTTCCTGTTTTCAGAAGG CCCAACTAAAGTCAGCAAATACAGGAAACAATGAAAGGATAATCAATGTATC AATTAAAAAGCTGAAGAGGAAACCACCTTCCACAAATGCAGGGAGAAGACA GAAACACAGACTAACATGCCCTTCATGTGATTCTTATGA
  • the soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTA
  • the sequence of soluble human IL-7 comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEH (SEQ ID NO: 79).
  • the soluble human IL-7 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGTGATATTGAAGGTAAAGATGGCAAACAATATGAGAGTGTTCTAATGG TCAGCATCGATCAATTATTGGACAGCATGAAAGAAATTGGTAGCAATTGCCT GAATAATGAATTTAACTTTTTTAAAAGACATATCTGTGATGCTAATAAGGAA GGTATGTTTTTATTCCGTGCTGCTCGCAAGTTGAGGCAATTTCTTAAAATGAA TAGCACTGGTGATTTTGATCTCCACTTATTAAAAGTTTCAGAAGGCACAACAA TACTGTTGAACTGCACTGGCCAGGTTAAAGGAAGAAAACC
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEHSGTTNTVAAY NLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVK DVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPTIQSFEQVG
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGATG GTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGCC TCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGGA GGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGATG AACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACCA CCATCCTGCTGAACTGCACCGGACAGGTGAAGGGCCGGAAACCTGCTGCTCT GGGAGAGA
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSDCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNN EFNFFKRHICDANKEGMFLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNC TGQVKGRKPAALGEAQPTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKIL MGTKEHSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKS KCFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTP
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CGATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGAT GGTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGC CTCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGG AGGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGAT GAACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACC ACCATCCTG
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDSITCPPPMSVEHADIWVKSYSLYSRERYICNSGF KRKAGTSSLTECVLNKATNVAHWTTPSLKCIR (SEQ ID NO: 211).
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTACGAGAAGAAGCC CCAAG
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSQGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPED VETNCEWSAFSCFQKAQLKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLT CPSCDSYEKKPPKEFLERFKSLLQKMIHQHLSSRTHGSEDSITCPPPMSVEHADIW VKSYSLYSRERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIR (SEQ ID NO: 213).
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CCAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTC GACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCC CCGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAA GGCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGT GAGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAAACAACGAGCGGATCATCAACGT GAG
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to a receptor for IL-18 (e.g., a soluble human IL-18), a receptor for IL-12 (e.g., a soluble human IL-12), or CD16 (e.g., an anti-CD16 scFv).
  • a receptor for IL-18 e.g., a soluble human IL-18
  • a receptor for IL-12 e.g., a soluble human IL-12
  • CD16 e.g., an anti-CD16 scFv
  • the first chimeric polypeptide further includes the additional target-binding domain.
  • the second chimeric polypeptide further includes the additional target-binding domain.
  • the additional target-binding domain binds specifically to CD16 or a receptor for IL-12.
  • the first target- binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the second chimeric polypeptide further comprises one or more additional target-binding domains at the N-terminal end or the C-terminal end of the second chimeric polypeptide.
  • the additional target-binding domain and the second domain of the pair of affinity domains directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the additional target-binding domain in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the additional target-binding domain and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second target-binding domain and the additional target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • one or more of the first target-binding domain, the second target-binding domain and the additional antigen-binding domain is an agonistic antigen-binding domain.
  • the first target-binding domain, the second target-binding domain, and the additional antigen-binding domain are each agonistic antigen-binding domains.
  • the antigen-binding domain includes a scFv or single-domain antibody.
  • one or both of the first target-binding domain and the second target-binding domain is a soluble IL-15 or a soluble IL-18.
  • the first target-binding domain and the second target-binding domain are each independently a soluble IL-15 or a soluble IL-18.
  • the first target-binding domain and the second target-binding domain both bind specifically to a receptor of IL-18 or a receptor of IL-12.
  • the first target-binding domain and the second target-binding domain bind specifically to the same epitope. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain include the same amino acid sequence. In some embodiments of these multi-chain chimeric polypeptides, the first target- binding domain binds specifically to a receptor for IL-12, and the second target-binding domain binds specifically to a receptor for IL-18.
  • the first target-binding domain binds specifically to a receptor for IL-18, and the second target-binding domain bind specifically to a receptor for IL-12. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain binds specifically to CD16, and the second target-binding domain binds specifically to a receptor for IL-18. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain binds specifically to a receptor for IL-18, and the second target-binding domain bind specifically to CD16.
  • two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to the same antigen. In some embodiments, two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to the same epitope. In some embodiments, two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains comprise the same amino acid sequence.
  • the first target- binding domain includes a soluble IL-18 (e.g., a soluble human IL-18).
  • the soluble human IL-18 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: YFGKLESKLSVIRNLNDQVLFIDQGNRPLFEDMTDSDCRDNAPRTIFIISMYKDSQ PRGMAVTISVKCEKISTLSCENKIISFKEMNPPDNIKDTKSDIIFFQRSVPGHDNKM QFESSSYEGYFLACEKERDLFKLILKKEDELGDRSIMFTVQNED (SEQ ID NO: 109).
  • the soluble human IL-18 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: TACTTCGGCAAACTGGAATCCAAGCTGAGCGTGATCCGGAATTTAAACGACC AAGTTCTGTTTATCGATCAAGGTAACCGGCCTCTGTTCGAGGACATGACCGAC TCCGATTGCCGGGACAATGCCCCCCGGACCATCTTCATTATCTCCATGTACAA GGACAGCCAGCCCCGGGGCATGGCTGTGACAATTAGCGTGAAGTGTGAGAA AATCAGCACTTTATCTTGTGAGAACAAGATCATCTCCTTTAAGGAAATGAACC CCCCCGATAACATCAAGGACACCAAGTCCGATAACATCAAGGACACCAAGTCCGATAACATCAAGGACA
  • the second target-binding domain includes a soluble IL-12 (e.g., a soluble human IL-12).
  • the soluble human IL-15 includes a sequence of soluble human IL-12 ⁇ (p40) and a sequence of soluble human IL- 12 ⁇ (p35).
  • the soluble IL-15 (e.g., soluble human IL-15) further includes a linker sequence (e.g., any of the exemplary linker sequences described herein) between the sequence of soluble IL-12 ⁇ (p40) and the sequence of soluble human IL-12 ⁇ (p35).
  • the linker sequence comprises GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the sequence of soluble human IL-12 ⁇ (p40) comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IWELKKDVYVVELDWYPDAPGEMVVLTCDTPEEDGITWTLDQSSEVLGSGKTLT IQVKEFGDAGQYTCHKGGEVLSHSLLLLHKKEDGIWSTDILKDQKEPKNKTFLR CEAKNYSGRFTCWWLTTISTDLTFSVKSSRGSSDPQGVTCGAATLSAERVRGDN KEYEYSVECQEDSACPAAEESLPIEVMVDAVHKLKYENYTSSFFIRDIIKPDPPKN LQLKPLKNSRQVEVSWEYPDTWS
  • the soluble human IL-12 ⁇ (p40) is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTTGGGAACTGAAGAAGGACGTCTACGTGGTCGAACTGGACTGGTATCCCG ATGCTCCCGGCGAAATGGTGGTGCTCACTTGTGACACCCCCGAAGAAGACGG CATCACTTGGACCCTCGATCAGAGCAGCGAGGTGCTGGGCTCCGGAAAGACC CTCACAATCCAAGTTAAGGAGTTCGGAGACGCTGGCCAATACACATGCCACA AGGGAGGCGAGGTGCTCAGCCATTCCTTATTATTATTACACAAGAAGGAAGA CGGAATCTGGTCCACCGACATTTTAAAAGATCAGA
  • the soluble human IL-12 ⁇ (p35) includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: RNLPVATPDPGMFPCLHHSQNLLRAVSNMLQKARQTLEFYPCTSEEIDHEDITKD KTSTVEACLPLELTKNESCLNSRETSFITNGSCLASRKTSFMMALCLSSIYEDLKM YQVEFKTMNAKLLMDPKRQIFLDQNMLAVIDELMQALNFNSETVPQKSSLEEPD FYKTKIKLCILLHAFRIRAVTIDRVMSYLNAS (SEQ ID NO: 80).
  • the soluble human IL-12 ⁇ (p35) is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CGTAACCTCCCCGTGGCTACCCCCGATCCCGGAATGTTCCCTTGTTTACACCA CAGCCAGAATTTACTGAGGGCCGTGAGCAACATGCTGCAGAAAGCTAGGCAG ACTTTAGAATTTTACCCTTGCACCAGCGAGGAGATCGACCATGAAGATATCA CCAAGGACAAGACATCCACCGTGGAGGCTTGTTTACCTCTGGAGCTGACAAA GAACGAGTCTTGTCTCAACTCTCGTGAAACCAGCTTCATCACAAATGGCTCTT GTTTAGCTTCCCGGAAGACCTCCT
  • the additional target-binding domain includes an scFv that specifically binds to CD16 (e.g., an anti- CD16 scFv).
  • the scFv that binds specifically to CD16 includes a light chain variable domain that includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: SELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQKPGQAPVLVIYGKNNRPS GIPDRFSGSSSGNTASLTITGAQAEDEADYYCNSRDSSGNHVVFGGGTKLTVGH (SEQ ID NO: 215).
  • the scFv that binds specifically to CD16 is encoded by a light chain variable domain sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: TCCGAGCTGACCCAGGACCCTGCTGTGTCCGTGGCTCTGGGCCAGACCGTGA GGATCACCTGCCAGGGCGACTCCCTGAGGTCCTACTACGCCTCCTGGTACCA GCAGAAGCCCGGCCAGGCTCCTGTGCTGGTGATCTACGGCAAGAACAACAGG CCCTCCGGCATCCCTGACAGGTTCTCCGGATCCTCCTCCGGCAACACCGCCTC CCTGACCATCACAGGCGCTCAGGCCGAGGACGAGGCTGACTACTACTGCAAC TCCAGGGACTCC
  • the scFv that binds specifically to CD16 includes a heavy chain variable domain that includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: EVQLVESGGGVVRPGGSLRLSCAASGFTFDDYGMSWVRQAPGKGLEWVSGINW NGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARGRSLLFDY WGQGTLVTVSR (SEQ ID NO: 217).
  • the scFv that binds specifically to CD16 is encoded by a heavy chain variable domain sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GAGGTGCAGCTGGTGGAGTCCGGAGGAGGAGTGGTGAGGCCTGGAGGCTCC CTGAGGCTGAGCTGTGCTGCCTCCGGCTTCACCTTCGACGACTACGGCATGTC CTGGGTGAGGCAGGCTCCTGGAAAGGGCCTGGAGTGGGTGTCCGGCATCAAC TGGAACGGCGGATCCACCGGCTACGCCGATTCCGTGAAGGGCAGGTTCACCA TCAGCAGGGACAACGCCAAGAACTCCCTGTACCTGCAGATGAACTCCCTGAG GGCCGAGG
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: YFGKLESKLSVIRNLNDQVLFIDQGNRPLFEDMTDSDCRDNAPRTIFIISMYKDSQ PRGMAVTISVKCEKISTLSCENKIISFKEMNPPDNIKDTKSDIIFFQRSVPGHDNKM QFESSSYEGYFLACEKERDLFKLILKKEDELGDRSIMFTVQNEDSGTTNTVAAYN LTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVKD VKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPT
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: TACTTCGGCAAACTGGAATCCAAGCTGAGCGTGATCCGGAATTTAAACGACC AAGTTCTGTTTATCGATCAAGGTAACCGGCCTCTGTTCGAGGACATGACCGAC TCCGATTGCCGGGACAATGCCCCGGACCATCTTCATTATCTCCATGTACAA GGACAGCCAGCCCCGGGGCATGGCTGTGACAATTAGCGTGAAGTGTGAGAA AATCAGCACTTTATCTTGTGAGAACAAGATCATCTCCTTTAAGGAAATGAACC CCCCCGATAACATCAAGGACACCAAGTCCGATATCATCTTCTTCCAGCGGTCC
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSYFGKLESKLSVIRNLNDQVLFIDQGNRPLFEDMTDSD CRDNAPRTIFIISMYKDSQPRGMAVTISVKCEKISTLSCENKIISFKEMNPPDNIKD TKSDIIFFQRSVPGHDNKMQFESSSYEGYFLACEKERDLFKLILKKEDELGDRSIM FTVQNEDSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKS KCFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSAGEPLY
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTCACATTTATCTCTTTACTGTTCCTCTTCTCCAGCGCCTACAGC TACTTCGGCAAACTGGAATCCAAGCTGAGCGTGATCCGGAATTTAAACGACC AAGTTCTGTTTATCGATCAAGGTAACCGGCCTCTGTTCGAGGACATGACCGAC TCCGATTGCCGGGACAATGCCCCCCGGACCATCTTCATTATCTCCATGTACAA GGACAGCCAGCCCCGGGGCATGGCTGTGACAATTAGCGTGAAGTGTGAGAA AATCAGCACTTTATCTTGTGAGAACAAGATCATCTCCTTTAAGGAAATGAACC
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IWELKKDVYVVELDWYPDAPGEMVVLTCDTPEEDGITWTLDQSSEVLGSGKTLT IQVKEFGDAGQYTCHKGGEVLSHSLLLLHKKEDGIWSTDILKDQKEPKNKTFLR CEAKNYSGRFTCWWLTTISTDLTFSVKSSRGSSDPQGVTCGAATLSAERVRGDN KEYEYSVECQEDSACPAAEESLPIEVMVDAVHKLKYENYTSSFFIRDIIKPDPPKN LQLKPLKNSRQVEVSWEYPDTWSTPHSYFSLTFCVQVQGKSKREKKDRVF
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTTGGGAACTGAAGAAGGACGTCTACGTGGTCGAACTGGACTGGTATCCCG ATGCTCCCGGCGAAATGGTGGTGCTCACTTGTGACACCCCCGAAGAAGACGG CATCACTTGGACCCTCGATCAGAGCAGCGAGGTGCTGGGCTCCGGAAAGACC CTCACAATCCAAGTTAAGGAGTTCGGAGACGCTGGCCAATACACATGCCACA AGGGAGGCGAGGTGCTCAGCCATTCCTTATTATTATTACACAAGAAGGAAGA CGGAATCTGGTCCACCGACATTTTAAAAGATCAGAAGGAGCCCAAGAATAAG ACCTTTAAGGT
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIWELKKDVYVVELDWYPDAPGEMVVLTCDTPEEDGI TWTLDQSSEVLGSGKTLTIQVKEFGDAGQYTCHKGGEVLSHSLLLLHKKEDGIW STDILKDQKEPKNKTFLRCEAKNYSGRFTCWWLTTISTDLTFSVKSSRGSSDPQG VTCGAATLSAERVRGDNKEYEYSVECQEDSACPAAEESLPIEVMVDAVHKLKYE NYTSSFFIRDIIKPDPPKNLQLKPLKNSRQVEVSWEYPDTWSTPHSYFSLTFC
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAATGGGTGACCTTTATTTCTTTACTGTTCCTCTTTAGCAGCGCCTACTCC ATTTGGGAACTGAAGAAGGACGTCTACGTGGTCGAACTGGACTGGTATCCCG ATGCTCCCGGCGAAATGGTGGTGCTCACTTGTGACACCCCCGAAGAAGACGG CATCACTTGGACCCTCGATCAGAGCAGCGAGGTGCTGGGCTCCGGAAAGACC CTCACAATCCAAGTTAAGGAGTTCGGAGACGCTGGCCAATACACATGCCACA AGGGAGGCGAGGTGCTCAGCCATTCCTTATTATTACACAAGAAGGAAGA CGGAATCTGGT
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to a receptor for IL-7 (e.g., a soluble human IL-7), CD16 (e.g., an anti-CD16 scFv), or a receptor for IL-21 (e.g., a soluble human IL-21).
  • a receptor for IL-7 e.g., a soluble human IL-7
  • CD16 e.g., an anti-CD16 scFv
  • a receptor for IL-21 e.g., a soluble human IL-21
  • the first chimeric polypeptide further includes the additional target-binding domain.
  • the second chimeric polypeptide further includes the additional target-binding domain.
  • the additional target-binding domain binds specifically to CD16 or a receptor for IL-21.
  • the first target- binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the second chimeric polypeptide further comprises one or more additional target-binding domains at the N-terminal end or the C-terminal end of the second chimeric polypeptide.
  • the additional target-binding domain and the second domain of the pair of affinity domains directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the additional target-binding domain in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the additional target-binding domain and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second target-binding domain and the additional target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • one or more of the first target-binding domain, the second target-binding domain and the additional antigen-binding domain is an agonistic antigen-binding domain.
  • the first target-binding domain, the second target-binding domain, and the additional antigen-binding domain are each agonistic antigen-binding domains.
  • the antigen-binding domain includes a scFv or single-domain antibody.
  • the first target-binding domain binds specifically to a receptor IL-7 and the second target-binding domain binds specifically to CD16 or a receptor for IL-21.
  • the first target-binding domain includes a soluble IL-7 protein.
  • the soluble IL-7 protein is a soluble human IL-7.
  • the second antigen-binding domain includes a target- binding domain that binds specifically to CD16.
  • the second target-binding domain includes an scFv that binds specifically to CD16. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the second target-binding domain binds specifically to a receptor for IL-21. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the second target-binding domain includes a soluble IL-21. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the soluble IL-21 is a soluble human IL-21.
  • the second chimeric polypeptide further includes an additional target-binding domain that binds specifically to a receptor for IL-21.
  • the additional target-binding domain includes a soluble IL-21.
  • the soluble IL-21 is a soluble human IL-21.
  • the second chimeric polypeptide further includes an additional target-binding domain that binds specifically to CD16.
  • two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to the same antigen. In some embodiments, two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to the same epitope. In some embodiments, two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains comprise the same amino acid sequence.
  • the first target- binding domain includes a soluble IL-7 (e.g., a soluble human IL-7).
  • the soluble human IL-7 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEH (SEQ ID NO: 79).
  • the soluble human IL-7 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGTGATATTGAAGGTAAAGATGGCAAACAATATGAGAGTGTTCTAATGG TCAGCATCGATCAATTATTGGACAGCATGAAAGAAATTGGTAGCAATTGCCT GAATAATGAATTTAACTTTTTTAAAAGACATATCTGTGATGCTAATAAGGAA GGTATGTTTTTATTCCGTGCTGCTCGCAAGTTGAGGCAATTTCTTAAAATGAA TAGCACTGGTGATTTTGATCTCCACTTATTAAAAGTTTCAGAAGGCACAACAA TACTGTTGAACTGCACTGGCCAGGTTAAAGGAAGAAAACC
  • the soluble human IL-7 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGATG GTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGCC TCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGGA GGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGATG AACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACCA CCATCCTGCTGAACTGCACCGGACAGGTGAAGGGCCGGAA
  • the sequence of soluble human IL-21 comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDS (SEQ ID NO: 83).
  • the soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAAGGTCAAGATCGCCACATGATTAGAATGCGTCAACTTATAGATATTGTTG ATCAGCTGAAAAATTATGTGAATGACTTGGTCCCTGAATTTCTGCCAGCTCCA GAAGATGTAGAGACAAACTGTGAGTGGTCAGCTTTTTCCTGTTTTCAGAAGG CCCAACTAAAGTCAGCAAATACAGGAAACAATGAAAGGATAATCAATGTATC AATTAAAAAGCTGAAGAGGAAACCACCTTCCACAAATGCAGGGAGAAGACA GAAACACAGACTAACATGCCCTTCATGTGATTCTTATGA
  • the soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTA
  • the scFv that binds specifically to CD16 includes a light chain variable domain that includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: SELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQKPGQAPVLVIYGKNNRPS GIPDRFSGSSSGNTASLTITGAQAEDEADYYCNSRDSSGNHVVFGGGTKLTVGH (SEQ ID NO: 215).
  • the scFv that binds specifically to CD16 is encoded by a light chain variable domain sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: TCCGAGCTGACCCAGGACCCTGCTGTGTCCGTGGCTCTGGGCCAGACCGTGA GGATCACCTGCCAGGGCGACTCCCTGAGGTCCTACTACGCCTCCTGGTACCA GCAGAAGCCCGGCCAGGCTCCTGTGCTGGTGATCTACGGCAAGAACAACAGG CCCTCCGGCATCCCTGACAGGTTCTCCGGATCCTCCTCCGGCAACACCGCCTC CCTGACCATCACAGGCGCTCAGGCCGAGGACGAGGCTGACTACTACTGCAAC TCCAGGGACTCC
  • the scFv that binds specifically to CD16 includes a heavy chain variable domain that includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: EVQLVESGGGVVRPGGSLRLSCAASGFTFDDYGMSWVRQAPGKGLEWVSGINW NGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARGRSLLFDY WGQGTLVTVSR (SEQ ID NO: 217).
  • the scFv that binds specifically to CD16 is encoded by a heavy chain variable domain sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GAGGTGCAGCTGGTGGAGTCCGGAGGAGGAGTGGTGAGGCCTGGAGGCTCC CTGAGGCTGAGCTGTGCTGCCTCCGGCTTCACCTTCGACGACTACGGCATGTC CTGGGTGAGGCAGGCTCCTGGAAAGGGCCTGGAGTGGGTGTCCGGCATCAAC TGGAACGGCGGATCCACCGGCTACGCCGATTCCGTGAAGGGCAGGTTCACCA TCAGCAGGGACAACGCCAAGAACTCCCTGTACCTGCAGATGAACTCCCTGAG GGCCGAGG
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEHSGTTNTVAAY NLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVK DVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPTIQSFEQVG
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGATG GTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGCC TCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGGA GGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGATG AACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACCA CCATCCTGCTGAACTGCACCGGACAGGTGAAGGGCCGGAAACCTGCTGCTCT GGGAGAGA
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSDCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNN EFNFFKRHICDANKEGMFLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNC TGQVKGRKPAALGEAQPTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKIL MGTKEHSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKS KCFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTP
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CGATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGAT GGTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGC CTCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGG AGGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGAT GAACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACC ACCATCCTG
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: SELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQKPGQAPVLVIYGKNNRPS GIPDRFSGSSSGNTASLTITGAQAEDEADYYCNSRDSSGNHVVFGGGTKLTVGHG GGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCAASGFTFDDYGMSWVRQ APGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTA VYYCARGRSLLFDYWGQGTLVTVSRITCPPPMSVEHADIWVKSYSLYSRERYIC NSGFK
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: TCCGAGCTGACCCAGGACCCTGCTGTGTCCGTGGCTCTGGGCCAGACCGTGA GGATCACCTGCCAGGGCGACTCCCTGAGGTCCTACTACGCCTCCTGGTACCA GCAGAAGCCCGGCCAGGCTCCTGTGCTGGTGATCTACGGCAAGAACAACAGG CCCTCCGGCATCCCTGACAGGTTCTCCGGATCCTCCTCCGGCAACACCGCCTC CCTGACCATCACAGGCGCTCAGGCCGAGGACGAGGCTGACTACTACTGCAAC TCCAGGGACTCCTCCGGCAACCATGTGGTGTTCGGCGGCGGCACCATGTGGTGTTCGGCGGCG
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSSELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQ KPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCNSRDSS GNHVVFGGGTKLTVGHGGGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSC AASGFTFDDYGMSWVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNA KNSLYLQMNSLRAEDTAVYYCARGRSLLFDYWGQGTLVTVSRITCPPPMSVEH ADIWVKSYS
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CTCCGAGCTGACCCAGGACCCTGCTGTGTCCGTGGCTCTGGGCCAGACCGTG AGGATCACCTGCCAGGGCGACTCCCTGAGGTCCTACTACGCCTCCTGGTACC AGCAGAAGCCCGGCCAGGCTCCTGTGCTGGTGATCTACGGCAAGAACAACAG GCCCTCCGGCATCCCTGACAGGTTCTCCGGATCCTCCTCCGGCAACACCGCCT CCCTGACCATCACAGGCGCTCAGGCCGAGGACGAGGCTGACTACTACTGCAA CTCCAGGG
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to TGF ⁇ (e.g., a human TGF ⁇ RII receptor), CD16 (e.g., an anti-CD16 scFv), or a receptor for IL-21 (e.g., a soluble human IL-21).
  • TGF ⁇ e.g., a human TGF ⁇ RII receptor
  • CD16 e.g., an anti-CD16 scFv
  • a receptor for IL-21 e.g., a soluble human IL-21
  • the first chimeric polypeptide further includes the additional target-binding domain.
  • the second chimeric polypeptide further includes the additional target-binding domain.
  • the additional target-binding domain binds specifically to CD16 or a receptor for IL-21.
  • the first target- binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the second chimeric polypeptide further comprises one or more additional target-binding domains at the N-terminal end or the C-terminal end of the second chimeric polypeptide.
  • the additional target-binding domain and the second domain of the pair of affinity domains directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the additional target-binding domain in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the additional target-binding domain and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second target-binding domain and the additional target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • one or more of the first target-binding domain, the second target-binding domain and the additional antigen-binding domain is an agonistic antigen-binding domain.
  • the first target-binding domain, the second target-binding domain, and the additional antigen-binding domain are each agonistic antigen-binding domains.
  • the antigen-binding domain includes a scFv or single-domain antibody.
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to TGF- ⁇ , CD16, or a receptor for IL-21. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the first target-binding domain binds specifically to a TGF- ⁇ and the second target-binding domain binds specifically to CD16 or a receptor of IL-21. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the first target-binding domain is a soluble TGF- ⁇ receptor.
  • soluble TGF- ⁇ receptor is a soluble TGF ⁇ RII receptor.
  • the second target-binding domain binds specifically to CD16.
  • the second antigen-binding domain includes an antigen-binding domain that binds specifically to CD16.
  • the second antigen-binding domain includes an scFv that binds specifically to CD16.
  • the second target-binding domain binds specifically to a receptor for IL-21. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the second target-binding domain includes a soluble IL-21. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the second target-binding domain includes a soluble human IL-21. In some embodiments of any of the multi-chain chimeric polypeptides described herein, the second chimeric polypeptide further includes an additional target-binding domain that binds specifically to a receptor for IL-21.
  • the additional target-binding domain includes a soluble IL-21.
  • the soluble IL-21 is a soluble human IL-21.
  • the second chimeric polypeptide further includes an additional target- binding domain that binds specifically to CD16. In some embodiments of these multi-chain chimeric polypeptides, two or more of the first target-binding domain, the second target-binding domain, and the one or more additional target-binding domains bind specifically to the same antigen.
  • the first target-binding domain includes a TGF ⁇ RII receptor (e.g., a soluble human TGF ⁇ RII receptor).
  • the soluble human TGFR ⁇ RII includes a first sequence of soluble human TGFR ⁇ RII and a second sequence of soluble human TGFR ⁇ RII.
  • the soluble human TGFR ⁇ RII includes a linker disposed between the first sequence of soluble human TGFR ⁇ RII and the second sequence of soluble human TGFR ⁇ RII.
  • the linker includes the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the first sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 183).
  • the second sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 184).
  • the first sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCT
  • the soluble TGF- ⁇ receptor includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSD
  • the soluble human TGF ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCAAA
  • the sequence of soluble human IL-21 comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDS (SEQ ID NO: 83).
  • the soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAAGGTCAAGATCGCCACATGATTAGAATGCGTCAACTTATAGATATTGTTG ATCAGCTGAAAAATTATGTGAATGACTTGGTCCCTGAATTTCTGCCAGCTCCA GAAGATGTAGAGACAAACTGTGAGTGGTCAGCTTTTTCCTGTTTTCAGAAGG CCCAACTAAAGTCAGCAAATACAGGAAACAATGAAAGGATAATCAATGTATC AATTAAAAAGCTGAAGAGGAAACCACCTTCCACAAATGCAGGGAGAAGACA GAAACACAGACTAACATGCCCTTCATGTGATTCTTATGA
  • the soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTA
  • the scFv that binds specifically to CD16 includes a light chain variable domain that includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: SELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQKPGQAPVLVIYGKNNRPS GIPDRFSGSSSGNTASLTITGAQAEDEADYYCNSRDSSGNHVVFGGGTKLTVGH (SEQ ID NO: 215).
  • the scFv that binds specifically to CD16 is encoded by a light chain variable domain sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: TCCGAGCTGACCCAGGACCCTGCTGTGTCCGTGGCTCTGGGCCAGACCGTGA GGATCACCTGCCAGGGCGACTCCCTGAGGTCCTACTACGCCTCCTGGTACCA GCAGAAGCCCGGCCAGGCTCCTGTGCTGGTGATCTACGGCAAGAACAACAGG CCCTCCGGCATCCCTGACAGGTTCTCCGGATCCTCCTCCGGCAACACCGCCTC CCTGACCATCACAGGCGCTCAGGCCGAGGACGAGGCTGACTACTACTGCAAC TCCAGGGACTCC
  • the scFv that binds specifically to CD16 includes a heavy chain variable domain that includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: EVQLVESGGGVVRPGGSLRLSCAASGFTFDDYGMSWVRQAPGKGLEWVSGINW NGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARGRSLLFDY WGQGTLVTVSR (SEQ ID NO: 217).
  • the scFv that binds specifically to CD16 is encoded by a heavy chain variable domain sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GAGGTGCAGCTGGTGGAGTCCGGAGGAGGAGTGGTGAGGCCTGGAGGCTCC CTGAGGCTGAGCTGTGCTGCCTCCGGCTTCACCTTCGACGACTACGGCATGTC CTGGGTGAGGCAGGCTCCTGGAAAGGGCCTGGAGTGGGTGTCCGGCATCAAC TGGAACGGCGGATCCACCGGCTACGCCGATTCCGTGAAGGGCAGGTTCACCA TCAGCAGGGACAACGCCAAGAACTCCCTGTACCTGCAGATGAACTCCCTGAG GGCCGAGG
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSDECND NIIFSEEYNTSNPD
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGGAAGAAG AAGC
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSI TSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKK PGETFFMCSC
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGAC AACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTT CAGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCC ATCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGGCGGAAAAATGAC GAGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACG ACTTCTCCCTTATCA
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: SELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQKPGQAPVLVIYGKNNRPS GIPDRFSGSSSGNTASLTITGAQAEDEADYYCNSRDSSGNHVVFGGGTKLTVGHG GGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCAASGFTFDDYGMSWVRQ APGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTA VYYCARGRSLLFDYWGQGTLVTVSRITCPPPMSVEHADIWVKSYSLYSRERYIC NSGFK
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: TCCGAGCTGACCCAGGACCCTGCTGTGTCCGTGGCTCTGGGCCAGACCGTGA GGATCACCTGCCAGGGCGACTCCCTGAGGTCCTACTACGCCTCCTGGTACCA GCAGAAGCCCGGCCAGGCTCCTGTGCTGGTGATCTACGGCAAGAACAACAGG CCCTCCGGCATCCCTGACAGGTTCTCCGGATCCTCCTCCGGCAACACCGCCTC CCTGACCATCACAGGCGCTCAGGCCGAGGACGAGGCTGACTACTACTGCAAC TCCAGGGACTCCTCCGGCAACCATGTGGTGTTCGGCGGCGGCACCATGTGGTGTTCGGCGGCG
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSSELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQ KPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCNSRDSS GNHVVFGGGTKLTVGHGGGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSC AASGFTFDDYGMSWVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNA KNSLYLQMNSLRAEDTAVYYCARGRSLLFDYWGQGTLVTVSRITCPPPMSVEH ADIWVKSYS
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CTCCGAGCTGACCCAGGACCCTGCTGTGTCCGTGGCTCTGGGCCAGACCGTG AGGATCACCTGCCAGGGCGACTCCCTGAGGTCCTACTACGCCTCCTGGTACC AGCAGAAGCCCGGCCAGGCTCCTGTGCTGGTGATCTACGGCAAGAACAACAG GCCCTCCGGCATCCCTGACAGGTTCTCCGGATCCTCCTCCGGCAACACCGCCT CCCTGACCATCACAGGCGCTCAGGCCGAGGACGAGGCTGACTACTACTGCAA CTCCAGGG
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to a receptor of IL-7.
  • the first target-binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • the first target- binding domain and the second target-binding domain each independently bind specifically to a receptor for IL-7. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain bind specifically to the same epitope. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain include the same amino acid sequence. In some embodiments of these multi-chain chimeric polypeptides, the first target- binding domain and the second target-binding domain include a soluble IL-7 (e.g., a soluble human IL-7).
  • a soluble IL-7 e.g., a soluble human IL-7
  • the soluble human IL-7 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEH (SEQ ID NO: 79).
  • the soluble human IL-7 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGATG GTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGCC TCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGGA GGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGATG AACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACCA CCATCCTGCTGAACTGCACCGGACAGGTGAAGGGCCGGAA
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEHSGTTNTVAAY NLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVK DVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPTIQSFEQVG
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGATG GTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGCC TCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGGA GGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGATG AACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACCA CCATCCTGCTGAACTGCACCGGACAGGTGAAGGGCCGGAAACCTGCTGCTCT GGGAGAGA
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSDCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNN EFNFFKRHICDANKEGMFLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNC TGQVKGRKPAALGEAQPTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKIL MGTKEHSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKS KCFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTP
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CGATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGAT GGTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGC CTCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGG AGGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGAT GAACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACC ACCATCCTG
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEHITCPPPMSVEH ADIWVKSYSLYSRERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIR (SEQ ID NO: 203).
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGTGATATTGAAGGTAAAGATGGCAAACAATATGAGAGTGTTCTAATGG TCAGCATCGATCAATTATTGGACAGCATGAAAGAAATTGGTAGCAATTGCCT GAATAATGAATTTAACTTTTTTAAAAGACATATCTGTGATGCTAATAAGGAA GGTATGTTTTTATTCCGTGCTGCTCGCAAGTTGAGGCAATTTCTTAAAATGAA TAGCACTGGTGATTTTGATCTCCACTTATTAAAAGTTTCAGAAGGCACAACAA TACTGTTGAACTGCACTGGCCAGGTTAAAGGAAGAAAACCAGCTGCCCTGGG TGAAGCC
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSDCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNN EFNFFKRHICDANKEGMFLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNC TGQVKGRKPAALGEAQPTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKIL MGTKEHITCPPPMSVEHADIWVKSYSLYSRERYICNSGFKRKAGTSSLTECVLNK ATNVAHWTTPSLKCIR (SEQ ID NO: 250).
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CGATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGAT GGTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGC CTCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGG AGGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGAT GAACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACC ACCATCCTG
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to TGF- ⁇ .
  • the first target-binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • the first target- binding domain and the second target-binding domain each independently bind specifically to TGF- ⁇ . In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain bind specifically to the same epitope. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain and the second target-binding domain include the same amino acid sequence.
  • the first target-binding domain and the second target-binding domain is a soluble TGF- ⁇ receptor (e.g., a soluble TGF ⁇ RII receptor, e.g., a soluble human TGF ⁇ RII).
  • the soluble human TGFR ⁇ RII includes a first sequence of soluble human TGFR ⁇ RII and a second sequence of soluble human TGFR ⁇ RII.
  • the soluble human TGFR ⁇ RII includes a linker disposed between the first sequence of soluble human TGFR ⁇ RII and the second sequence of soluble human TGFR ⁇ RII.
  • the linker includes the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the first sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 183).
  • the second sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 184).
  • the first sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCT
  • the second sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTCCTCCCCACGTGCAGAAGAGCGTGAATAATGACATGATCGTGACCGATA ACAATGGCGCCGTGAAATTTCCCCAGCTGTGCAAATTCTGCGATGTGAGGTTT TCCACCTGCGACAACCAGAAGTCCTGTATGAGCAACTGCTCCATCACCTCCAT CTGTGAGAAGCCTCAGGAGGTGTGCGTGGCTGTCTGGCGGAAGAATGACGAG AATATCACCCTGGAAACCGTCTGCCACGATCCCAAGCTGCCCTACCACGATTT CATCCTGGAAGACGCCGCCA
  • the soluble TGF- ⁇ receptor includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSD
  • the soluble TGF- ⁇ receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAA
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSDECND NIIFSEEYNTSNPD
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGGAAGAAG AAGC
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSI TSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKK PGETFFMCSC
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGAC AACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTT CAGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCC ATCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGGCGGAAAAATGAC GAGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACG ACTTCTCCCTTATCA
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSDECND NIIFSEEYNTSNPD
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGGAAGAAG AAGC
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSI TSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKK PGETFFMCSC
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGAC AACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTT CAGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCC ATCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGGCGGAAAAATGAC GAGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACG ACTTCTCCCTTATCA
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to a receptor of IL-7, a receptor of IL-21, or a receptor of CD137L.
  • the second chimeric polypeptide further includes the additional target-binding domain.
  • the additional target-binding domain binds specifically to a receptor for IL-21 (e.g., a soluble IL-21, e.g., a soluble human IL-21) or a receptor for CD137L (e.g., a soluble CD137L, e.g., a soluble human CD137L).
  • a receptor for IL-21 e.g., a soluble IL-21, e.g., a soluble human IL-21
  • CD137L e.g., a soluble CD137L, e.g., a soluble human CD137L
  • the first target- binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the second chimeric polypeptide further comprises one or more additional target-binding domains at the N-terminal end or the C-terminal end of the second chimeric polypeptide.
  • the additional target-binding domain and the second domain of the pair of affinity domains directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the additional target-binding domain in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the additional target-binding domain and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second target-binding domain and the additional target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • the second chimeric polypeptide can include an additional target-binding domain.
  • the additional target-binding domain and the In some embodiments of these multi-chain chimeric polypeptides one or more of the first target-binding domain, the second target-binding domain and the additional target-binding domain is an agonistic antigen-binding domain. In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain, the second target-binding domain, and the additional target-binding domain are each agonistic antigen-binding domains. In some embodiments of these multi-chain chimeric polypeptides, the antigen-binding domain includes a scFv or single-domain antibody.
  • the first target- binding domain binds specifically to a receptor for IL-7
  • the second target-binding domain binds specifically to a receptor for IL-21 or a receptor for CD137L.
  • the additional target-binding domain binds specifically to a receptor for IL- 21 or a receptor for CD137L.
  • the first target- binding domain is a soluble IL-7 (e.g., a soluble human IL-7).
  • the soluble human IL-7 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEH (SEQ ID NO: 79).
  • the soluble human IL-7 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGATG GTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGCC TCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGGA GGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGATG AACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACCA CCATCCTGCTGAACTGCACCGGACAGGTGAAGGGCCGGAA
  • the second target-binding domain or the additional target-binding domain binds specifically to a receptor for IL-21.
  • the second target-binding domain or the additional target-binding domain is a soluble IL-21 (e.g., a soluble human IL-21).
  • a soluble human IL-21 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDS (SEQ ID NO: 83).
  • a soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCC
  • the second target-binding domain binds specifically to a receptor for CD137L. In some embodiments of these multi-chain chimeric polypeptides, the second chimeric polypeptide further comprises an additional target-binding domain that binds specifically to a receptor for CD137L. In some embodiments of these multi-chain chimeric polypeptides, the second target-binding domain and/or the additional target-binding domain is a soluble CD137L (e.g., a soluble human CD137L).
  • a soluble human CD137L includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: REGPELSPDDPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGGL SYKEDTKELVVAKAGVYYVFFQLELRRVVAGEGSGSVSLALHLQPLRSAAGAA ALALTVDLPPASSEARNSAFGFQGRLLHLSAGQRLGVHLHTEARARHAWQLTQ GATVLGLFRVTPEIPAGLPSPRSE (SEQ ID NO: 260).
  • a soluble human CD137L is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CGCGAGGGTCCCGAGCTTTCGCCCGACGATCCCGCCGGCCTCTTGGACCTGC GGCAGGGCATGTTTGCGCAGCTGGTGGCCCAAAATGTTCTGCTGATCGATGG GCCCCTGAGCTGGTACAGTGACCCAGGCCTGGCAGGCGTGTCCCTGACGGGG GGCCTGAGCTACAAAGAGGACACGAAGGAGCTGGTGGTGGCCAAGGCTGGA GTCTACTATGTCTTCTTTCAACTAGAGCTGCGGCGCGTGGTGGCCGGCGAGGG CTCAGGCTCCGTTTCACTTGCGCTGC
  • a soluble human CD137L includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGGLSYKEDTKEL VVAKAGVYYVFFQLELRRVVAGEGSGSVSLALHLQPLRSAAGAAALALTVDLP PASSEARNSAFGFQGRLLHLSAGQRLGVHLHTEARARHAWQLTQGATVLGLFR VTPEI (SEQ ID NO: 262).
  • a soluble human CD137L is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATCCCGCCGGCCTCTTGGACCTGCGGCAGGGCATGTTTGCGCAGCTGGTGG CCCAAAATGTTCTGCTGATCGATGGGCCCCTGAGCTGGTACAGTGACCCAGG CCTGGCAGGCGTGTCCCTGACGGGGGGCCTGAGCTACAAAGAGGACACGAA GGAGCTGGTGGTGGCCAAGGCTGGAGTCTACTATGTCTTCTTTCAACTAGAGC TGCGGCGCGTGGTGGCCGGCGAGGGCTCAGGCTCCGTTTCACTTGCGCTGCA CCTGCAGCCACTGCTCTGCTGCTGCTGCTGCTGCTGCTGCTGCTGCTGCTGCTGCTGC
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEHSGTTNTVAAY NLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVK DVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPTIQSFEQVG
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGATG GTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGCC TCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGGA GGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGATG AACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACCA CCATCCTGCTGAACTGCACCGGACAGGTGAAGGGCCGGAAACCTGCTGCTCT GGGAGAGA
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSDCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNN EFNFFKRHICDANKEGMFLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNC TGQVKGRKPAALGEAQPTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKIL MGTKEHSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKS KCFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTP Y
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CGATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGAT GGTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGC CTCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGG AGGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGAT GAACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACC ACCATCCTG
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDSITCPPPMSVEHADIWVKSYSLYSRERYICNSGF KRKAGTSSLTECVLNKATNVAHWTTPSLKCIRGGGGSGGGGSGGGGSREGPELS PDDPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGGLSYKEDTK ELV
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTACGAGAAGAAGCC CCAAG
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSQGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPED VETNCEWSAFSCFQKAQLKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLT CPSCDSYEKKPPKEFLERFKSLLQKMIHQHLSSRTHGSEDSITCPPPMSVEHADIW VKSYSLYSRERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIRGGGGS GGGGSGGGGSREGPELSPDDPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPG
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CCAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTC GACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCC CCGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAA GGCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGT GAGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAAACAACGAGCGGATCATCAACGT GAG
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDSITCPPPMSVEHADIWVKSYSLYSRERYICNSGF KRKAGTSSLTECVLNKATNVAHWTTPSLKCIRGGGGSGGGGSGGGGSDPAGLL DLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGGLSYKEDTKELVVAKA GVYY
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTACGAGAAGAAGCC CCAAG
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSQGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPED VETNCEWSAFSCFQKAQLKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLT CPSCDSYEKKPPKEFLERFKSLLQKMIHQHLSSRTHGSEDSITCPPPMSVEHADIW VKSYSLYSRERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIRGGGGS GGGGSGGSDPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTG G
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTC CCAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTC GACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCC CCGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAA GGCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGT GAGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAAACAACGAGCGGATCATCAACGT GAG
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to a receptor of IL-7 or TGF- ⁇ .
  • the first target-binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • the first target- binding domain binds specifically to a receptor for IL-7, and the second target-binding domain binds specifically to TGF- ⁇ . In some embodiments of these multi-chain chimeric polypeptides, the first target-binding domain binds specifically to TGF- ⁇ , and the second target-binding domain binds specifically to a receptor for IL-7. In some embodiments of these multi-chain chimeric polypeptides, the first target- binding domain includes a soluble IL-7 protein (e.g., a soluble human IL-7 protein).
  • the soluble human IL-7 protein includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEH (SEQ ID NO: 79).
  • the soluble human IL-7 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGATG GTGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGCC TCAACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGGA GGGCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGATG AACTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACCA CCATCCTGCTGAACTGCACCGGACAGGTGAAGGGCCGGAA
  • the second target-binding domain comprises a target-binding domain that binds specifically to TGF- ⁇ .
  • the second target- binding domain is a soluble TGF- ⁇ receptor (e.g., a soluble TGF ⁇ RII receptor, e.g., a soluble human TGF ⁇ RII receptor).
  • the soluble human TGFR ⁇ RII includes a first sequence of soluble human TGFR ⁇ RII and a second sequence of soluble human TGFR ⁇ RII.
  • the soluble human TGFR ⁇ RII includes a linker disposed between the first sequence of soluble human TGFR ⁇ RII and the second sequence of soluble human TGFR ⁇ RII.
  • the linker includes the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the first sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 183).
  • the second sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 184).
  • the first sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCT
  • the second sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTCCTCCCCACGTGCAGAAGAGCGTGAATAATGACATGATCGTGACCGATA ACAATGGCGCCGTGAAATTTCCCCAGCTGTGCAAATTCTGCGATGTGAGGTTT TCCACCTGCGACAACCAGAAGTCCTGTATGAGCAACTGCTCCATCACCTCCAT CTGTGAGAAGCCTCAGGAGGTGTGCGTGGCTGTCTGGCGGAAGAATGACGAG AATATCACCCTGGAAACCGTCTGCCACGATCCCAAGCTGCCCTACCACGATTT CATCCTGGAAGACGCCGCCA
  • the soluble TGF- ⁇ receptor includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSD
  • the soluble TGF- ⁇ receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAA
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: DCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNNEFNFFKRHICDANKEGM FLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNCTGQVKGRKPAALGEAQ PTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKILMGTKEHSGTTNTVAAYN LTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVKD VKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPTIQSFEQVG
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGATGG TGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGCCTC AACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGGAGG GCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGATGAA CTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACCACC ATCCTGCTGAACTGCACCGGACAGGTGAAGGGCCGGAAACCTGCTGCTCTGG GAGAGG
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSDCDIEGKDGKQYESVLMVSIDQLLDSMKEIGSNCLNN EFNFFKRHICDANKEGMFLFRAARKLRQFLKMNSTGDFDLHLLKVSEGTTILLNC TGQVKGRKPAALGEAQPTKSLEENKSLKEQKKLNDLCFLKRLLQEIKTCWNKIL MGTKEHSGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSK CFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTP
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC GATTGCGACATCGAGGGCAAGGACGGCAAGCAGTACGAGAGCGTGCTGATGG TGTCCATCGACCAGCTGCTGGACAGCATGAAGGAGATCGGCTCCAACTGCCTC AACAACGAGTTCAACTTCTTCAAGCGGCACATCTGCGACGCCAACAAGGAGG GCATGTTCCTGTTCAGGGCCGCCAGGAAACTGCGGCAGTTCCTGAAGATGAA CTCCACCGGCGACTTCGACCTGCACCTGCTGAAGGTGTCCGAGGGCACCACC ATCC
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRKN DENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGS
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGAAGAAGAAGAAG
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSIT SICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKP GETFFMC
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACT
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to TGF- ⁇ , a receptor of IL-21, or a receptor of CD137L.
  • the second chimeric polypeptide further includes the additional target-binding domain.
  • the additional target-binding domain binds specifically to a receptor for IL-21 (e.g., a soluble IL-21, e.g., a soluble human IL-21) or a receptor for CD137L (e.g., a soluble CD137L, e.g., a soluble human CD137L).
  • a receptor for IL-21 e.g., a soluble IL-21, e.g., a soluble human IL-21
  • CD137L e.g., a soluble CD137L, e.g., a soluble human CD137L
  • the first target- binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the second chimeric polypeptide further comprises one or more additional target-binding domains at the N-terminal end or the C-terminal end of the second chimeric polypeptide.
  • the additional target-binding domain and the second domain of the pair of affinity domains directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the additional target-binding domain in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the additional target-binding domain and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second target-binding domain and the additional target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • one or more of the first target-binding domain, the second target-binding domain and the additional target-binding domain is an agonistic antigen-binding domain.
  • the first target-binding domain, the second target-binding domain, and the additional target-binding domain are each agonistic antigen-binding domains.
  • the antigen-binding domain includes a scFv or single-domain antibody.
  • the first target- binding domain binds specifically to TGF- ⁇ and the second target-binding domain binds specifically to a receptor for IL-21 or a receptor for CD137L.
  • the first target- binding domain is a soluble TGF- ⁇ receptor (e.g., a soluble TGF ⁇ RII receptor, e.g., a soluble human TGF ⁇ RII receptor).
  • the soluble human TGFR ⁇ RII includes a first sequence of soluble human TGFR ⁇ RII and a second sequence of soluble human TGFR ⁇ RII.
  • the soluble human TGFR ⁇ RII includes a linker disposed between the first sequence of soluble human TGFR ⁇ RII and the second sequence of soluble human TGFR ⁇ RII.
  • the linker includes the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the first sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 183).
  • the second sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 184).
  • the first sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCT
  • the second sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTCCTCCCCACGTGCAGAAGAGCGTGAATAATGACATGATCGTGACCGATA ACAATGGCGCCGTGAAATTTCCCCAGCTGTGCAAATTCTGCGATGTGAGGTTT TCCACCTGCGACAACCAGAAGTCCTGTATGAGCAACTGCTCCATCACCTCCAT CTGTGAGAAGCCTCAGGAGGTGTGCGTGGCTGTCTGGCGGAAGAATGACGAG AATATCACCCTGGAAACCGTCTGCCACGATCCCAAGCTGCCCTACCACGATTT CATCCTGGAAGACGCCGCCA
  • the soluble TGF- ⁇ receptor includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSD
  • the soluble TGF- ⁇ receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAA
  • the second target-binding domain or the additional target-binding domain binds specifically to a receptor for IL-21.
  • the second target-binding domain or the additional target-binding domain includes a soluble IL-21(e.g., a soluble human IL-21).
  • a soluble human IL-21 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDS (SEQ ID NO: 83).
  • the soluble human IL-21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTA
  • the second target-binding domain or the additional target-binding domain binds specifically to a receptor for CD137L.
  • the second target-binding domain and/or the additional target-binding domain includes a soluble CD137L (e.g., a soluble human CD137L).
  • a soluble CD137L includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: REGPELSPDDPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGGL SYKEDTKELVVAKAGVYYVFFQLELRRVVAGEGSGSVSLALHLQPLRSAAGAA ALALTVDLPPASSEARNSAFGFQGRLLHLSAGQRLGVHLHTEARARHAWQLTQ GATVLGLFRVTPEIPAGLPSPRSE (SEQ ID NO: 260).
  • a soluble CD137L is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CGCGAGGGTCCCGAGCTTTCGCCCGACGATCCCGCCGGCCTCTTGGACCTGC GGCAGGGCATGTTTGCGCAGCTGGTGGCCCAAAATGTTCTGCTGATCGATGG GCCCCTGAGCTGGTACAGTGACCCAGGCCTGGCAGGCGTGTCCCTGACGGGG GGCCTGAGCTACAAAGAGGACACGAAGGAGCTGGTGGTGGCCAAGGCTGGA GTCTACTATGTCTTCTTTCAACTAGAGCTGCGGCGCGTGGTGGCCGGCGAGGG CTCAGGCTCCGTTTCACTTGCGCTGCACC
  • a soluble human CD137L includes a sequence that is at least 80% identical (e.g., at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical) to: DPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGGLSYKEDTKEL VVAKAGVYYVFFQLELRRVVAGEGSGSVSLALHLQPLRSAAGAAALALTVDLP PASSEARNSAFGFQGRLLHLSAGQRLGVHLHTEARARHAWQLTQGATVLGLFR VTPEI (SEQ ID NO: 262).
  • a soluble human CD137L is encoded by a sequence that is at least 80% identical (e.g., at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical) to: GATCCCGCCGGCCTCTTGGACCTGCGGCAGGGCATGTTTGCGCAGCTGGTGG CCCAAAATGTTCTGCTGATCGATGGGCCCCTGAGCTGGTACAGTGACCCAGG CCTGGCAGGCGTGTCCCTGACGGGGGGCCTGAGCTACAAAGAGGACACGAA GGAGCTGGTGGTGGCCAAGGCTGGAGTCTACTATGTCTTCTTTCAACTAGAGC TGCGGCGCGTGGTGGCCGGCGAGGGCTCAGGCTCCGTTTCACTTGCGCTGCA CCTGCAGCCACTGCTCTGCTGCTGGGGCCGCCGCCCTGGCTTTGACCGTGG ACCTGCCTCCTCCGAGG
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRKN DENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGS
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGAAGAAGAAGAAG
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSIT SICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKP GETFFMC
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACT
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDSITCPPPMSVEHADIWVKSYSLYSRERYICNSGF KRKAGTSSLTECVLNKATNVAHWTTPSLKCIRGGGGSGGGGSGGGGSREGPELSP DDPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGGLSYKEDTKE LV
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTGA GCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGGC AGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTACGAGAAGAAGCCCCC CAAGGA
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTGA GCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGGC AGAAGC
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDSITCPPPMSVEHADIWVKSYSLYSRERYICNSGF KRKAGTSSLTECVLNKATNVAHWTTPSLKCIRGGGGSGGGGSGGGGSDPAGLLD LRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGGLSYKEDTKELVVAKAGV Y
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTGA GCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGGC AGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTACGAGAAGAAGCCCCC CAAGGA
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSQGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPED VETNCEWSAFSCFQKAQLKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLT CPSCDSYEKKPPKEFLERFKSLLQKMIHQHLSSRTHGSEDSITCPPPMSVEHADIW VKSYSLYSRERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIRGGGGS GGGGSGGSDPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGG
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTGA GCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGGC AGAAGC
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to TGF- ⁇ or a receptor of IL-21.
  • the second chimeric polypeptide further includes the additional target-binding domain.
  • the additional target-binding domain binds specifically to a receptor for IL-21 (e.g., a soluble IL-21, e.g., a soluble human IL-21) or a TGF- ⁇ (e.g., a soluble TGF ⁇ receptor, e.g., a soluble TGF ⁇ RII receptor).
  • IL-21 e.g., a soluble IL-21, e.g., a soluble human IL-21
  • TGF- ⁇ e.g., a soluble TGF ⁇ receptor, e.g., a soluble TGF ⁇ RII receptor
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the second chimeric polypeptide further comprises one or more additional target-binding domains at the N-terminal end or the C-terminal end of the second chimeric polypeptide.
  • the additional target-binding domain and the second domain of the pair of affinity domains directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the additional target-binding domain in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the additional target-binding domain and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second target-binding domain and the additional target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • the first target- binding domain binds specifically to TGF- ⁇
  • the second target-binding domain binds specifically to TGF- ⁇ or a receptor for IL-21.
  • the first target-binding domain is a soluble TGF- ⁇ receptor (e.g., a soluble TGF ⁇ RII receptor, e.g., a soluble human TGF ⁇ RII receptor).
  • the soluble human TGFR ⁇ RII includes a first sequence of soluble human TGFR ⁇ RII and a second sequence of soluble human TGFR ⁇ RII.
  • the soluble human TGFR ⁇ RII includes a linker disposed between the first sequence of soluble human TGFR ⁇ RII and the second sequence of soluble human TGFR ⁇ RII.
  • the linker includes the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the first sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 183).
  • the second sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 184).
  • the first sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCT
  • the second sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTCCTCCCCACGTGCAGAAGAGCGTGAATAATGACATGATCGTGACCGATA ACAATGGCGCCGTGAAATTTCCCCAGCTGTGCAAATTCTGCGATGTGAGGTTT TCCACCTGCGACAACCAGAAGTCCTGTATGAGCAACTGCTCCATCACCTCCAT CTGTGAGAAGCCTCAGGAGGTGTGCGTGGCTGTCTGGCGGAAGAATGACGAG AATATCACCCTGGAAACCGTCTGCCACGATCCCAAGCTGCCCTACCACGATTT CATCCTGGAAGACGCCGCCA
  • the soluble TGF- ⁇ receptor includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSD
  • the soluble TGF- ⁇ receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCAAATGCAT
  • the second target-binding domain binds specifically to a receptor for IL-21.
  • the second target-binding domain includes a soluble IL-21 (e.g., a human soluble IL-21).
  • the soluble IL-21 includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: QGQDRHMIRMRQLIDIVDQLKNYVNDLVPEFLPAPEDVETNCEWSAFSCFQKAQ LKSANTGNNERIINVSIKKLKRKPPSTNAGRRQKHRLTCPSCDSYEKKPPKEFLER FKSLLQKMIHQHLSSRTHGSEDS (SEQ ID NO: 83).
  • the soluble IL- 21 is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: CAGGGCCAGGACAGGCACATGATCCGGATGAGGCAGCTCATCGACATCGTCG ACCAGCTGAAGAACTACGTGAACGACCTGGTGCCCGAGTTTCTGCCTGCCCC CGAGGACGTGGAGACCAACTGCGAGTGGTCCGCCTTCTCCTGCTTTCAGAAG GCCCAGCTGAAGTCCGCCAACACCGGCAACAACGAGCGGATCATCAACGTG AGCATCAAGAAGCTGAAGCGGAAGCCTCCCTCCACAAACGCCGGCAGGAGG CAGAAGCACAGGCTGACCTGCCCCAGCTGTGACTCCTA
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRKN DENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGS
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGAAGAAGAAGAAG
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSIT SICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKP GETFFMC
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACT
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRKN DENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGS
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGAAGAAGAAGAAG
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSIT SICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKP GETFFMC
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACT
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to TGF- ⁇ or CD16.
  • the second chimeric polypeptide further includes the additional target-binding domain.
  • the additional target-binding domain binds specifically to CD16 (e.g., an anti-CD16 scFv) or a TGF- ⁇ (e.g., a soluble TGF ⁇ receptor, e.g., a soluble TGF ⁇ RII receptor).
  • CD16 e.g., an anti-CD16 scFv
  • TGF- ⁇ e.g., a soluble TGF ⁇ receptor, e.g., a soluble TGF ⁇ RII receptor
  • the first target- binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the second chimeric polypeptide further comprises one or more additional target-binding domains at the N-terminal end or the C-terminal end of the second chimeric polypeptide.
  • the additional target-binding domain and the second domain of the pair of affinity domains directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the additional target-binding domain in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the additional target-binding domain and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second target-binding domain and the additional target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • the first target- binding domain binds specifically to TGF- ⁇
  • the second target-binding domain binds specifically to TGF- ⁇ or CD16.
  • the first target-binding domain is a soluble TGF- ⁇ receptor (e.g., a soluble TGF ⁇ RII receptor, e.g., a soluble human TGF ⁇ RII receptor).
  • the soluble human TGFR ⁇ RII includes a first sequence of soluble human TGFR ⁇ RII and a second sequence of soluble human TGFR ⁇ RII.
  • the soluble human TGFR ⁇ RII includes a linker disposed between the first sequence of soluble human TGFR ⁇ RII and the second sequence of soluble human TGFR ⁇ RII.
  • the linker includes the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the first sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 183).
  • the second sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 184).
  • the first sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCT
  • the second sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTCCTCCCCACGTGCAGAAGAGCGTGAATAATGACATGATCGTGACCGATA ACAATGGCGCCGTGAAATTTCCCCAGCTGTGCAAATTCTGCGATGTGAGGTTT TCCACCTGCGACAACCAGAAGTCCTGTATGAGCAACTGCTCCATCACCTCCAT CTGTGAGAAGCCTCAGGAGGTGTGCGTGGCTGTCTGGCGGAAGAATGACGAG AATATCACCCTGGAAACCGTCTGCCACGATCCCAAGCTGCCCTACCACGATTT CATCCTGGAAGACGCCGCCA
  • the soluble TGF- ⁇ receptor includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSD
  • the soluble TGF- ⁇ receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCAAATGCAT
  • the second target-binding domain binds specifically to CD16. In some embodiments of these multi-chain chimeric polypeptides, the second target-binding domain includes an anti-CD16 scFv.
  • the scFv that binds specifically to CD16 includes a light chain variable domain that includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: SELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQKPGQAPVLVIYGKNNRPS GIPDRFSGSSSGNTASLTITGAQAEDEADYYCNSRDSSGNHVVFGGGTKLTVGH (SEQ ID NO: 215).
  • the scFv that binds specifically to CD16 is encoded by a light chain variable domain sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: TCCGAGCTGACCCAGGACCCTGCTGTGTCCGTGGCTCTGGGCCAGACCGTGA GGATCACCTGCCAGGGCGACTCCCTGAGGTCCTACTACGCCTCCTGGTACCA GCAGAAGCCCGGCCAGGCTCCTGTGCTGGTGATCTACGGCAAGAACAACAGG CCCTCCGGCATCCCTGACAGGTTCTCCGGATCCTCCTCCGGCAACACCGCCTC CCTGACCATCACAGGCGCTCAGGCCGAGGACGAGGCTGACTACTACTGCAAC TCCAGGGACTCC
  • the scFv that binds specifically to CD16 includes a heavy chain variable domain that includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: EVQLVESGGGVVRPGGSLRLSCAASGFTFDDYGMSWVRQAPGKGLEWVSGINW NGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARGRSLLFDY WGQGTLVTVSR (SEQ ID NO: 217).
  • the scFv that binds specifically to CD16 is encoded by a heavy chain variable domain sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: GAGGTGCAGCTGGTGGAGTCCGGAGGAGGAGTGGTGAGGCCTGGAGGCTCC CTGAGGCTGAGCTGTGCTGCCTCCGGCTTCACCTTCGACGACTACGGCATGTC CTGGGTGAGGCAGGCTCCTGGAAAGGGCCTGGAGTGGGTGTCCGGCATCAAC TGGAACGGCGGATCCACCGGCTACGCCGATTCCGTGAAGGGCAGGTTCACCA TCAGCAGGGACAACGCCAAGAACTCCCTGTACCTGCAGATGAACTCCCTGAG GGCCGAGG
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRKN DENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGS
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGAAGAAGAAGAAG
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSIT SICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKP GETFFMC
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACT
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRKN DENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGS
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGAAGAAGAAGAAG
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSIT SICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKP GETFFMC
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACT
  • the first target-binding domain and the second targeting-binding domain each independently bind specifically to TGF- ⁇ or a receptor of CD137L.
  • the second chimeric polypeptide further includes the additional target-binding domain.
  • the additional target-binding domain binds specifically to a receptor to TGF- ⁇ (e.g., a soluble TGF- ⁇ receptor, e.g., a soluble TGF ⁇ RII receptor) or CD137L.
  • the first target- binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the second chimeric polypeptide further comprises one or more additional target-binding domains at the N-terminal end or the C-terminal end of the second chimeric polypeptide.
  • the additional target-binding domain and the second domain of the pair of affinity domains directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the additional target-binding domain in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the additional target-binding domain and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second target-binding domain and the additional target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • the first target- binding domain binds specifically to TGF- ⁇
  • the second target-binding domain binds specifically to CD137L.
  • the first target-binding domain or the additional target-binding domain is a soluble TGF- ⁇ receptor (e.g., a soluble TGF ⁇ RII receptor, e.g., a soluble human TGF ⁇ RII receptor).
  • the soluble human TGFR ⁇ RII includes a first sequence of soluble human TGFR ⁇ RII and a second sequence of soluble human TGFR ⁇ RII.
  • the soluble human TGFR ⁇ RII includes a linker disposed between the first sequence of soluble human TGFR ⁇ RII and the second sequence of soluble human TGFR ⁇ RII.
  • the linker includes the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the first sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 183).
  • the second sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 184).
  • the first sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCT
  • the second sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTCCTCCCCACGTGCAGAAGAGCGTGAATAATGACATGATCGTGACCGATA ACAATGGCGCCGTGAAATTTCCCCAGCTGTGCAAATTCTGCGATGTGAGGTTT TCCACCTGCGACAACCAGAAGTCCTGTATGAGCAACTGCTCCATCACCTCCAT CTGTGAGAAGCCTCAGGAGGTGTGCGTGGCTGTCTGGCGGAAGAATGACGAG AATATCACCCTGGAAACCGTCTGCCACGATCCCAAGCTGCCCTACCACGATTT CATCCTGGAAGACGCCGCCA
  • the soluble TGF- ⁇ receptor includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSD
  • the soluble TGF- ⁇ receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCAAATGCAT
  • the second target-binding domain includes a soluble CD137L protein (e.g., a soluble human CD137L protein).
  • a soluble human CD137L includes a sequence that is at least 80% identical (e.g., at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical) to: REGPELSPDDPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGGL SYKEDTKELVVAKAGVYYVFFQLELRRVVAGEGSGSVSLALHLQPLRSAAGAA ALALTVDLPPASSEARNSAFGFQGRLLHLSAGQRLGVHLHTEARARHAWQLTQ GATVLGLFRVTPEIPAGLPSPRSE (SEQ ID NO: 260).
  • a soluble human CD137L is encoded by a sequence that is at least 80% identical (e.g., at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical) to: CGCGAGGGTCCCGAGCTTTCGCCCGACGATCCCGCCGGCCTCTTGGACCTGC GGCAGGGCATGTTTGCGCAGCTGGTGGCCCAAAATGTTCTGCTGATCGATGG GCCCCTGAGCTGGTACAGTGACCCAGGCCTGGCAGGCGTGTCCCTGACGGGG GGCCTGAGCTACAAAGAGGACACGAAGGAGCTGGTGGTGGCCAAGGCTGGA GTCTACTATGTCTTCTTTCAACTAGAGCTGCGGCGCGTGGTGGCCGGCGAGGG CTCAGGCTCCGTTTCACTTGCGCTGCACCTGCAGCCACTGCTCTGCTGCTG GGGCCCTGGCTTTGACC
  • a soluble human CD137L includes a sequence that is at least 80% identical (e.g., at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical) to: DPAGLLDLRQGMFAQLVAQNVLLIDGPLSWYSDPGLAGVSLTGGLSYKEDTKEL VVAKAGVYYVFFQLELRRVVAGEGSGSVSLALHLQPLRSAAGAAALALTVDLP PASSEARNSAFGFQGRLLHLSAGQRLGVHLHTEARARHAWQLTQGATVLGLFR VTPEI (SEQ ID NO: 262).
  • a soluble human CD137L is encoded by a sequence that is at least 80% identical (e.g., at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical) to: GATCCCGCCGGCCTCTTGGACCTGCGGCAGGGCATGTTTGCGCAGCTGGTGG CCCAAAATGTTCTGCTGATCGATGGGCCCCTGAGCTGGTACAGTGACCCAGG CCTGGCAGGCGTGTCCCTGACGGGGGGCCTGAGCTACAAAGAGGACACGAA GGAGCTGGTGGTGGCCAAGGCTGGAGTCTACTATGTCTTCTTTCAACTAGAGC TGCGGCGCGTGGTGGCCGGCGAGGGCTCAGGCTCCGTTTCACTTGCGCTGCA CCTGCAGCCACTGCTCTGCTGCTGGGGCCGCCGCCCTGGCTTTGACCGTGG ACCTGCCTCCTCCGAGG
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRKN DENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGS
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGAAGAAGAAGAAG
  • a first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSIT SICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKP GETFFMC
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACT
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRKN DENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGS
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACTTC ATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGAAGAAGAAGAAG
  • a second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: MKWVTFISLLFLFSSAYSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFST CDNQKSCMSNCSITSICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILED AASPKCIMKEKKKPGETFFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSG GGGSIPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSIT SICEKPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKP GETFFMC
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATGAAGTGGGTGACCTTCATCAGCCTGCTGTTCCTGTTCTCCAGCGCCTACTCC ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACAA CAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTCA GCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCATCT GCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACGAGA ACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGACT
  • the first target-binding domain and the second targeting-binding domain each bind specifically to TGF- ⁇ .
  • the first chimeric polypeptide further includes the additional target- binding domain.
  • a non-limiting example of this type of multi-chain chimeric polypeptide is shown in Figures 209 and 210.
  • the first target- binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further comprises a linker sequence (e.g., any of the exemplary linkers described herein) between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the soluble tissue factor domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide.
  • the first chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the second domain of the pair of affinity domains and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
  • the second chimeric polypeptide further comprises one or more additional target-binding domains at the N-terminal end or the C-terminal end of the second chimeric polypeptide.
  • the additional target-binding domain and the second domain of the pair of affinity domains directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second domain of the pair of affinity domains and the additional target-binding domain in the second chimeric polypeptide.
  • a linker sequence e.g., any of the exemplary linkers described herein
  • the additional target-binding domain and the second target-binding domain directly abut each other in the second chimeric polypeptide.
  • the second chimeric polypeptide further includes a linker sequence (e.g., any of the exemplary linkers described herein) between the second target-binding domain and the additional target-binding domain in the second chimeric polypeptide.
  • the soluble tissue factor domain can be any of the exemplary soluble tissue factor domains described herein.
  • the pair of affinity domains can be any of the exemplary pairs of affinity domains described herein.
  • the first target- binding domain binds specifically to TGF- ⁇
  • the second target-binding domain binds specifically to TGF- ⁇
  • the first target-binding domain and/or the second target-binding domain is a soluble TGF- ⁇ receptor (e.g., a soluble TGF ⁇ RII receptor, e.g., a soluble human TGF ⁇ RII receptor).
  • the soluble human TGFR ⁇ RII includes a first sequence of soluble human TGFR ⁇ RII and a second sequence of soluble human TGFR ⁇ RII.
  • the soluble human TGFR ⁇ RII includes a linker disposed between the first sequence of soluble human TGFR ⁇ RII and the second sequence of soluble human TGFR ⁇ RII.
  • the linker includes the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 102).
  • the first sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 183).
  • the second sequence of soluble human TGFR ⁇ RII receptor comprises a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPD (SEQ ID NO: 184).
  • the first sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCT
  • the second sequence of soluble human TGFR ⁇ RII receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATTCCTCCCCACGTGCAGAAGAGCGTGAATAATGACATGATCGTGACCGATA ACAATGGCGCCGTGAAATTTCCCCAGCTGTGCAAATTCTGCGATGTGAGGTTT TCCACCTGCGACAACCAGAAGTCCTGTATGAGCAACTGCTCCATCACCTCCAT CTGTGAGAAGCCTCAGGAGGTGTGCGTGGCTGTCTGGCGGAAGAATGACGAG AATATCACCCTGGAAACCGTCTGCCACGATCCCAAGCTGCCCTACCACGATTT CATCCTGGAAGACGCCGCCA
  • the soluble TGF- ⁇ receptor includes a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGET FFMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDM IVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRK NDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETFFMCSCSSD
  • the soluble TGF- ⁇ receptor is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCAAATGCAT
  • the first chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRKN DENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGS
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCACCGCACGTTCAGAAGTCGGTGAATAACGACATGATAGTCACTGACA ACAACGGTGCAGTCAAGTTTCCACAACTGTGTAAATTTTGTGATGTGAGATTT TCCACCTGTGACAACCAGAAATCCTGCATGAGCAACTGCAGCATCACCTCCA TCTGTGAGAAGCCACAGGAAGTCTGTGTGGCTGTATGGAGAAAGAATGACGA GAACATAACACTAGAGACAGTTTGCCATGACCCCAAGCTCCCCTACCATGAC TTTATTCTGGAAGATGCTGCTTCTCCAAAGTGCATTATGAAGGAAAAAAAAAAAAAAAAAAAAAAAAAAAAA
  • a first chimeric polypeptide can include a soluble IL-15 including a D8N amino acid substitution and have a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: (Signal peptide) MGVKVLFALICIAVAEA (Single chain Human TGF-beta Receptor II homodimer) IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLC
  • a first chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: (Signal peptide) ATGGGAGTGAAAGTTCTTTTTTTGCCCTTATTTGTATTGCTGTGGCCGAGGCC (Single chain Human TGF-beta Receptor II homodimer) ATCCCACCGCACGTTCAGAAGTCGGTGAATAACGACATGATAGTCACTGACA ACAACGGTGCAGTCAAGTTTCCACAACTGTGTAAATTTTGTGATGTGAGATTT TCCACCTGTGACAACCAGAAATCCTGCATGAGCAACTGCAGCATCACCTCCA TCTGTGAGAAGCCACAGGAAGTCTGTGTGGCTGTATGGAGAAAGAATGACGA GA GA
  • the second chimeric polypeptide can include a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: IPPHVQKSVNNDMIVTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICE KPQEVCVAVWRKNDENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGSGGGGSGGGGSIPPHVQKSVNNDMI VTDNNGAVKFPQLCKFCDVRFSTCDNQKSCMSNCSITSICEKPQEVCVAVWRKN DENITLETVCHDPKLPYHDFILEDAASPKCIMKEKKKPGETF FMCSCSSDECNDNIIFSEEYNTSNPDGGGGS
  • a second chimeric polypeptide is encoded by a sequence that is at least 80% identical (e.g., at least 82% identical, at least 84% identical, at least 86% identical, at least 88% identical, at least 90% identical, at least 92% identical, at least 94% identical, at least 96% identical, at least 98% identical, at least 99% identical, or 100% identical) to: ATCCCCCCCCATGTGCAAAAGAGCGTGAACAACGATATGATCGTGACCGACA ACAACGGCGCCGTGAAGTTTCCCCAGCTCTGCAAGTTCTGCGATGTCAGGTTC AGCACCTGCGATAATCAGAAGTCCTGCATGTCCAACTGCAGCATCACCTCCA TCTGCGAGAAGCCCCAAGAAGTGTGCGTGGCCGTGTGGCGGAAAAATGACG AGAACATCACCCTGGAGACCGTGTGTCACGACCCCAAGCTCCCTTATCACGA CTTCATTCTGGAGGACGCTGCCTCCCCCAAATGCATCATGAAGGAAGAAG AAGC

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