US20160324944A1 - Methods and compositions relating to p62/SQSTM1 for the treatment and prevention of inflammation-associated diseases - Google Patents

Methods and compositions relating to p62/SQSTM1 for the treatment and prevention of inflammation-associated diseases Download PDF

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US20160324944A1
US20160324944A1 US15/108,653 US201415108653A US2016324944A1 US 20160324944 A1 US20160324944 A1 US 20160324944A1 US 201415108653 A US201415108653 A US 201415108653A US 2016324944 A1 US2016324944 A1 US 2016324944A1
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sqstm1
polypeptide
nucleic acid
disease
inflammatory
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Alexander Shneider
Franco Venanzi
Dimitrios Agas
Antonio Concetti
Maria Giovanna Sabbieti
Vladimir Gabai
Michael Sherman
Victor Shifrin
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CURELAB ONCOLOGY Inc
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Definitions

  • This invention relates generally to prevention and treatment of inflammatory diseases. More specifically, the invention relates to prevention and treatment of inflammatory diseases by administration of p62 compositions.
  • Inflammation is an essential immune response that enables survival during infection or injury and maintains tissue homeostasis under a variety of noxious conditions. It can be divided in acute and chronic inflammation. Acute inflammation is a protective response to pathogens like bacteria and viruses, or to tissue damage. In response to infection or tissue damage, macrophages induce production of inflammatory cytokines (e.g., TNF, IL-1, IL-6) and chemokines (e.g., CCL2 and CXCL8), as well as prostaglandins.
  • cytokines e.g., TNF, IL-1, IL-6
  • chemokines e.g., CCL2 and CXCL8
  • IL-1, TNF, and IL-6 can have systemic effects when secreted in sufficient amounts. They induce liver cells (hepatocytes) to produce acute phase proteins such as C-reactive protein and coagulation factors, and they activate brain endothelium to produce prostaglandins, including the major proinflammatory prostaglandin, PGE2. Locally produced PGE2, in turn, induces specific populations of neurons in the central nervous system to promote so-called sickness behavior: fever, anorexia, fatigue, sleepiness, and social withdrawal (Pecchi et al. 2009.
  • the resolution phase may not be appropriately induced and a chronic inflammatory state may ensue.
  • This state can be caused by chronic infections, unrepaired tissue damage, persistent allergens, undigestable foreign particles, or endogenous crystals, such as monosodium urate (Majno 2004. Cell, Tissues, and Disease; Kumar 2003. Robbins Basic Pathology.)
  • the chronic inflammatory response in these cases is typically localized to the site where the inflammatory inducer is present and often results in different types of local tissue remodeling.
  • Osteoporosis is the most common disease of the bone associated with bone loss and affecting mostly women after onset of menopause. Menopause leads to decrease in estrogen levels, thus ovariectomy in rodents leading to cessation of estrogen generation is the most common model for osteoporosis. Postmenopausal period is marked by elevation of cytokines such us IL-6, TNF-alpha and IL-1beta, and the same cytokines are elevated under ovariectomy.
  • TNF and IL-1 have potent antiapoptotic effects in OCs, prolonging OC lifespan, accelerating bone resorption and inhibiting bone formation, and blockade of TNF-alpha and IL-1beta prevents osteoporosis due to estrogen deficiency (Mundy 2007. Osteoporosis and Inflammation. Nutrition Reviews 65:S147-S151; Lencel and Magne 2011. Inflammaging: The driving force in osteoporosis? Medical Hypotheses 76:317-321).
  • ALS Amyotrophic lateral sclerosis
  • Lou Gehrig's disease is a progressive fatal neurodegenerative disease that affects motor neurons in the brainstem, spinal cord, and motor cortex.
  • ALS is universally fatal, with a median age of onset of 55 years and a survival of 2-5 years after the onset of symptoms.
  • Prominent neuroinflammation can be easily observed in pathologically affected areas of the CNS and in spinal cords from both human ALS patients and mouse models of the disease (Smith et al. 2012. Role of pro-inflammatory cytokines released from microglia in neurodegenerative diseases. Brain Res Bull 87:10-20).
  • inflammation in ALS is characterized by gliosis and the accumulation of large numbers of activated microglia and astrocytes.
  • Activation of glia in ALS has been extensively characterized and is marked by elevated production of potentially cytotoxic molecules such as ROS, inflammatory mediators such as COX-2, and proinflammatory cytokines such as IL-1 beta, TNF-alpha, and IL-6 (Smith et al. 2012. Role of pro-inflammatory cytokines released from microglia in neurodegenerative diseases. Brain Res Bull 87:10-20).
  • the most common mouse model of ALS is transgenic mouse expressing mutant form of superoxide dismutase, the same mutant form as seen in some ALS patients.
  • MS Multiple sclerosis
  • MS is a heterogeneous and complex autoimmune disease that is characterized by inflammation, demyelination, and axon degeneration in the CNS. This pathology results from a primary defect in the immune system that targets components of the myelin sheath, resulting in secondary effects on neurons.
  • MS is considered an immune-mediated disease characterized by the presence of inflammatory demyelinating lesions in the CNS. Infection by bacteria or viruses or other environmental stimuli trigger the activation of microglia and astrocytes in multiple sclerosis (MS), leading to the production of proinflammatory cytokines through activation of the transcription factors NF-kappa-B and AP-1 (Luessi et al. 2012.
  • EAE Experimental autoimmune encephalomyelitis
  • the first class includes such well-known drugs as aspirin, glucocorticoids, non-steroidal anti-inflammatory agents (celecoxib) and other agents (e.g., methotrexate, cyclosporine, rapamycin etc.).
  • the second class includes agents that reduce activity of specific cytokines or their receptors, e.g., antibodies to TNF (see scheme below).
  • p62 is a multifunctional protein that binds ubiquitin and regulates autophagy, activity of the nuclear factor kappa-B and some other signaling pathways.
  • the protein functions as a scaffolding/adaptor protein in concert with TNF receptor-associated factor 6 (TRF6) to mediate activation of NF-kappa-B in response to upstream signals.
  • TRF6 TNF receptor-associated factor 6
  • spliced transcript variants encoding either the same or different isoforms have been identified for this gene.
  • p62 was identified as 62-kDa protein that binds the src homology 2 (SH2) domain of tyrosine kinase Lckp56 in a phosphotyrosine-independent manner (Moscat et al. 2007. Signal integration and diversification through the p62 scaffold protein. Trends Biochem Sci 32:95-100). The primary sequence of p62 is known, and p62 was shown to bind ubiquitin. (Moscat et al. 2007. Signal integration and diversification through the p62 scaffold protein. Trends Biochem Sci 32:95-100).
  • FIG. 1 shows the nucleic acid sequence of the cDNA and FIG. 2 the amino acid sequence.
  • a proinflammatory cytokine in a subject by administering to the subject an agent that includes: (a) at least 30 amino acids of a p62/SQSTM1 polypeptide or a variant thereof; or, (b) a p62/SQSTM1 encoding nucleic acid, wherein said p62/SQSTM1 encoding nucleic acid encodes at least 30 amino acids of a p62/SQSTM1 polypeptide or a variant thereof.
  • the proinflammatory cytokine can be TNF ⁇ , IL-6, IL-1b, RANTES, IL-17, IL-23, CCL-1, MCP-5, or CXCL2.
  • the osteogenic transcription factor can be osterix or runx2.
  • the bone resorptive factor can be TNF ⁇ or RANKL.
  • a non-cancer-related chronic inflammatory disease in a subject by administering to the subject an agent comprising that includes: (a) at least 30 amino acids of a p62/SQSTM1 polypeptide or a variant thereof; or, (b) a p62/SQSTM1 encoding nucleic acid, wherein said p62/SQSTM1 encoding nucleic acid encodes at least 30 amino acids of a p62/SQSTM1 polypeptide or a variant thereof.
  • any of the above methods can include administration of a variant p62/SQSTM1, wherein the variant is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% homologous to any sequence selected from the group consisting of SEQ. ID. NO. 2-35 10.
  • the variant can have at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence homology to any sequence selected from the group consisting of SEQ. ID. NO. 2-35, or identical thereto.
  • Any of the above methods can include administration of p62-encoding nucleic acid comprises the sequence of SEQ ID NO:1.
  • Any of the above methods can include administration of p62 polypeptide or variant thereof having at least one domain deletion.
  • the deleted domain can be PB1, ZZ, NLS2, TB, NLS1, NES, LIR, KIR, and UBA.
  • any of the above methods can include administration of an agent including a p62 encoding nucleic acid, wherein said p62 encoding nucleic acid encodes a polypeptide, which is at least 95% identical to SEQ ID NO. 2, and wherein said p62 encoding nucleic acid further comprises a plasmid, RNA or a viral vector.
  • any of the above methods can include p62/SQSTM1 polypeptide or p62/SQSTM1 encoding nucleic acid further including a fusion polypeptide or nucleic acid encoding for a fusion polypeptide, respectively.
  • any of the above methods can include p62/SQSTM1 polypeptide or p62/SQSTM1 encoding nucleic acid in the form of a vaccine and further include administering an adjuvant to said subject.
  • the adjuvant can be gel-type, microbial, particulate, oil-emulsion, surfactant-based, and synthetic adjuvant.
  • the non-cancer-related chronic inflammatory disease can be obesity, metabolic syndrome, type 2 diabetes, fat liver, Crohn's Disease, pancreatitis, asthma, chronic obstructive pulmonary disease, arthritis, osteoporosis, osteoarthritis, multiple sclerosis, psoriasis, congestive heart failure atherosclerosis, neurodegenerative diseases, gout, asbestosis, and silicosis.
  • the neurodegenerative disease can be amyotrophic lateral sclerosis, Parkinson's disease, Huntington's disease, or Alzheimer's disease.
  • the methods to treat, alleviate, ameliorate, relieve, delay onset of, inhibit progression of, reduce severity of, or reduce incidence of one or more symptoms of a non-cancer-related chronic inflammatory disease in a subject can further include administering an anti-inflammatory therapy to said subject.
  • Any of the above methods can be applied to a subject that is a subject diagnosed with an inflammatory disease, a subject previously treated for an inflammatory disease, a subject with a family history of inflammatory disease, or a subject predisposed to an inflammatory disease.
  • the methods to treat, alleviate, ameliorate, relieve, delay onset of, inhibit progression of, reduce severity of, or reduce incidence of one or more symptoms of a non-cancer-related chronic inflammatory disease in a subject can further include a strategy for improving the efficiency of nucleic acid-based expression of p62 in subjects.
  • the strategy can include a self-replicating viral replicon, codon optimization, in vivo electroporation, incorporation of a CpG stimulatory motif, including a sequence for targeting of the endocytic or ubiquitin-processing pathways, including a Marek's disease virus type 1 VP22 sequence, a prime-boost regimen, a mucosal delivery vector, and a nucleic acid delivery system.
  • the nucleic acid delivery system can be a polymer gene delivery system, a liposomal delivery system, and a cell-penetrating peptide gene delivery system.
  • any of the above methods can further include administering an anti-inflammatory chemotherapeutic or biological agent.
  • the chemotherapeutic agent can be a nonsteroidal anti-inflammatory drug, a glucocorticoid, methotrexate, cyclosporine, or rapamycin.
  • the anti-inflammatory biological agent can be an anti-TNF antibody, an anti-IL1 antibody, an anti-IL6 antibody, an anti-IL6 receptor antibody, an anti-IL12/23 antibody, an anti-IL17 antibody, an anti-IL1R antibody, an anti-IL1 receptor antagonist, and a soluble IL-1 receptor.
  • FIG. 1 shows a wild type nucleic acid sequence of human p62 (SEQ ID NO: 1);
  • FIG. 2 shows a wild type amino acid sequence of the human p62/SQSTM1 encoded by the nucleic acid sequence (SEQ ID NO: 2);
  • FIG. 3 shows a cartoon of the domain structure of human p62/SQSTM1
  • FIG. 4 shows the effect of p62/SQSTM1 DNA introduction on the prevention of osteoporosis in a mouse model for osteoporosis
  • FIG. 5 shows the effect of p62/SQSTM1 DNA introduction on osteogenic markers
  • FIG. 6 shows the effect of p62/SQSTM1 DNA introduction on bone mineral density (BMD) and content (BMC);
  • FIG. 7 Panel A shows the effect of p62/SQSTM1 DNA introduction on expression of osteogenic factors; Panel B shows the effect of p62/SQSTM1 DNA introduction on bone resorptive factors and NF-kappa-B expression;
  • FIG. 8 Panel A shows the effect of exogenous p62 on p62 expression; Panel B shows increase p62-immune labelling at the epiphyseal region of femurs of p62-OVX mice; Panel C Western Blots demonstrate that the increased level of p62 is from endogenous p62.
  • FIG. 9 shows the effect of p62/SQSTM1 DNA introduction on a mouse model of ALS
  • FIG. 10 shows the effect of p62/SQSTM1 DNA introduction on a mouse model of MS
  • FIG. 11 shows the effect of p62/SQSTM1 DNA introduction on growth of S37 sarcoma
  • FIG. 12 shows the effect of p62/SQSTM1 DNA introduction on survival of mice with breast carcinoma
  • FIG. 13 shows the effect of p62/SQSTM1 DNA introduction on metastasis in LLC.
  • FIG. 14 shows the effect of p62/SQSTM1 DNA introduction on metastasis in B16 melanoma.
  • p62 compositions and methods for treatment of chronic inflammation are provided herein.
  • the inventors have found that administering p62, such as a p62 encoding nucleic acid, to a subject suppresses generation of inflammatory cytokines. Consequently polynucleotides encoding a p62 polypeptide or, p62 polypeptides administered to a subject can be used to prevent and/or mitigate development of inflammation-associated diseases (the list of such diseases includes but is not limited to, osteoporosis, obesity, metabolic syndrome, type 2 diabetes, fat liver, inflammatory bowel disease, gastritis, chronic pancreatitis, asthma, chronic obstructive pulmonary disease (COPD), rheumatoid arthritis (RA), osteoarthritis, multiple sclerosis (MS), psoriasis, congestive heart failure (CHF), atherosclerosis, neurodegenerative diseases (ALS, Parkinson, Alzheimer's, Huntington disease), gout, asbestosis and silicosis.
  • p62 polypeptide means a polypeptide corresponding to the full length p62/SQSTM1 protein. The term also includes all homologs, analogs, fragments or derivatives of the p62/SQSTM1 protein.
  • the isolated p62 polypeptide has an amino acid sequence as shown in FIG. 2 (SEQ ID NO: 2).
  • a “p62 encoding nucleic acid” means a DNA or RNA that encodes at least a portion of a p62 polypeptide or variant.
  • the subject is a human. In other embodiments, the subject is a non-human mammal including, but not limited to, a horse, cow, sheep, pig, deer, dog, cat, rat, or a mouse.
  • polypeptides of the present invention may also include fragments or truncations, analogs, and homologs of the p62 polypeptide and truncations thereof as described herein. Fragments can include peptides (or encode peptides) of at least 5, at least 10, at least 15, at least 20, at least 25, at least 30, at least 50, at least 100, at least 200 or at least 300 amino acid residues of the full length polypeptide.
  • Deletions of one or more amino acids, or discrete portions from the amino acid sequence of the p62/SQSTM1 protein are also included.
  • the deleted amino acids may or may not be contiguous.
  • the lower limit length of the resulting analog with a deletion mutation is about 10, about 20, about 50, or about 100 amino acids.
  • the p62 polypeptide (or a nucleic acid encoding for the polypeptide) has one or more deleted domains. While not wishing to be held by theory, the inventors hold that the deletion of one or more domains of the p62 polypeptide provide a more compact and manipulable polypeptide for directing an immune response. For example, by disrupting or eliminating one or more of the domains of a p62 polypeptide, anti-inflammatory effect can be retained (or improved if the deleted or disrupted domain does not contribute to this effect) in a more compact molecule and potentially increase per weight basis.
  • the p62 polypeptide has a domain structure as provided in Table 2 below and as shown in FIG. 3 :
  • PB1-domain is conserved among eukaryotes domain (protista, plants, fungi and animals).
  • There domain) are 3 types of PB1-domains: type I domains contains acid OPCA-motif, type II domains contain conservative Lys residue in the first beta-sheet, and I/II type domains contain both of the above.
  • OPCA- motif can bind to basic amino acids (e.g., lysine) via salt bridges, enabling ability of PB1-domains to form heteromeric structures (Sumimoto et al., 2007).
  • PB1- domain of p62 is type I/II (Lamark et al., 2003).
  • PB1-domain is responsible for di- and multimerization of p62, as well as interaction with other proteins: MEKK3, MEK5, PKC ⁇ , PKC-lambda/ ⁇ (protein kinases containing PB1-domain), NBR1 (Next to BRCA1, contains PB1-domain) (Nezis, Stenmark, 2011).
  • ZZ Zn2+-finger 122-167 ZZ- domain is Zn 2+ -finger of C2H2 type.
  • ZZ-domain ZZ type of p62 binds to RIP1 (receptor interacting protein 1).
  • RIP1 is a regulatory protein kinase which integrates signaling pathways activated by bacterial or viral infection (via PAMP), death receptors, or genotoxins; it takes part in determination of cell fate (survival, apoptosis, or necrosis) (Festjens et al., 2007).
  • TRAF6- 228-233 p62 binds via TB domain to E3-ubiquitin protein binding ligase TRAF6.
  • TRAF6 activates kinase TAK1, domain polyubiqitinating it via K63).
  • TRAF6 participates in signaling from RANK-L, IL-1R, TCR, BCR and TGFbeta receptors (Landström, 2010). Interaction of p62 with TRAF6 stimulates autoubiqitination of TRAF6 nu E3- ligase activity. This process requires PB1- and UBA-domains (Moscat et al., 2006).
  • Keap1 (Kelch-like ECH- associated protein 1) is a regulator of activity of transcription factor Nrf2 (NF-E2-related factor 2). Nrf2 regulates expression of genes involved in glutathione synthesis, ROS detoxification, metabolism of xenobiotics and drug transport (Taguchi et al., 2011).
  • UBA-domains can be divided in four classes depending on their ability to bind polyubiquitin labels of different structures (K6, K29, K48, K63).
  • UBA-domain of p62 belongs to class 4, which consists of domains with equal affinity for binding to K6, K29, K48, K63 (Raasi et al., 2005).
  • UBA domain also participates in p62 dimerization (Garner et al., 2011). Most of the mutations associated with Paget disease are localized in UBA domain (Yan Jenny Chung, Van Hul, 2011). However, p62 mutations are not enough for osteoblasts to acquire the specific Paget phenotype: The expression of nucleocapsid protein of measles virus is also required (Singer, 2011). The structure of the UBA domain is known (Isogai et al., 2011).
  • one or more of the above domains are deleted from a human p62 polypeptide at corresponding codons for the nucleic acid regions of the p62 nucleic acid (in-frame deletions), as presented below.
  • any deletion of the encoding nucleic acid sequence that starts at nucleotide 102 up to nucleotide 122 and ends at 167 up to 183 is considered a ZZ deletion. Therefore, e.g. a deletion of nucleotides 110-175 is a ZZ deletion.
  • Techniques for creating in-frame deletions are well known to those skilled in the art.
  • biologically active refers to polypeptides according to the present invention having a similar structural function (but not necessarily to the same degree), and/or similar regulatory function (but not necessarily to the same degree), and/or similar biochemical function (but not necessarily to the same degree) as the individual wild type polypeptides.
  • a “deletion” is defined as a change in the nucleotide or amino acid sequence in which one or more nucleotide or amino acid residues are absent as compared to the wild-type polynucleotide or polypeptide, respectively.
  • an “insertion” or “addition” is a change in the nucleotide or amino acid sequence that has resulted in the addition of one or more nucleotide or amino acid residues as compared to the wild-type polynucleotide or polypeptide, respectively.
  • substitution results from the replacement of one or more nucleotides or amino acids by different nucleotides or amino acids, respectively, as compared to the wild-type polynucleotide or polypeptide, respectively.
  • the amino acid substitution mutation is C145R or Q418R.
  • the term “variant” means any polypeptide (including polypeptides encoded by the corresponding nucleic acid) having a substitution of, deletion of or addition of one (or more) amino acid from or to the sequence (or any combination of these), including allelic variations, as compared with the wild-type polypeptide.
  • the resultant polypeptide retains at least 75%, 80%, 85%, 90%, 95%, 99% or more of the biological activity as compared to the wild-type polypeptides as used in the present invention.
  • Variants of the p62 polypeptides can have at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to any of the amino acid sequences listed in Table 1.
  • Sequence identity or homology can be determined using standard techniques known in the art, such as the Best Fit sequence program described by Devereux et al., Nucl. Acid Res. 12:387-395 (1984) or the BLASTX program (Altschul et al., J Mol. Biol. 215:403-410).
  • the alignment may include the introduction of gaps in the sequences to be aligned.
  • the percentage of homology will be determined based on the number of homologous amino acids in relation to the total number of amino acids.
  • variants of the p62 polypeptides can have at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence homology to any of the polypeptide sequences listed in Table 1
  • variants or derivatives of the polypeptides of the present invention maintain the hydrophobicity/hydrophilicity of the amino acid sequence.
  • Conservative amino acid substitutions are known in the art and may be made, for example from 1, 2 or 3 to 10, 20 or 30 substitutions. Amino acid substitutions may include the use of non-naturally occurring analogues, for example to increase blood plasma half-life.
  • derivative as used herein in relation to the amino acid sequence means chemical modification of a polypeptide of the invention.
  • Non-limiting examples of such modifications may include but are not limited to aliphatic esters or amides of the carboxyl terminus or of residues containing carboxyl side chains, O-acyl derivatives of hydroxyl group-containing residues, and N-acyl derivatives of the amino-terminal amino acid or amino-group containing residues, e.g., lysine or arginine.
  • Additional modifications can include, for example, production of a polypeptide conjugated with a polymer such as polyethylene glycol (PEG), or addition of PEG during chemical synthesis of a polypeptide of the invention.
  • a polymer such as polyethylene glycol (PEG)
  • PEG polyethylene glycol
  • Modifications of polypeptides or portions thereof can also include reduction/alkylation, chemical coupling to an appropriate carrier, or mild formalin treatment.
  • post-translationally modified refers to any modification of an amino acid after its incorporation into a polypeptide chain.
  • the term encompasses, but is not limited to, co-translational in vivo modifications, post-translational in vivo modifications, and post-translational in vitro modifications.
  • polypeptides of the present invention include incorporation of unnatural amino acid residues, or phosphorylated amino acid residues such as phosphotyrosine, phosphoserine or phosphothreonine residues.
  • Other potential modifications include sulfonation, biotinylation, or the addition of other moieties, particularly those which have molecular shapes similar to phosphate groups.
  • Derivatives also include polypeptides modified by glycosylation. These can be made by modifying glycosylation patterns during synthesis and processing in various alternative eukaryotic host expression systems, or during further processing steps. Methods for producing glycosylation modifications include exposing the fusion proteins to glycosylating enzymes derived from cells that normally carry out such processing, such as mammalian glycosylation enzymes. Alternatively, deglycosylation enzymes can be used to remove carbohydrates attached during production in eukaryotic expression systems. Additionally, one can also modify the coding sequence so that glycosylations site(s) are added or glycosylation sites are deleted or disabled. Furthermore, if no glycosylation is desired, the proteins can be produced in a prokaryotic host expression system.
  • Variants and/or derivatives of the polypeptides of the invention can be prepared by chemical synthesis or by using site-directed mutagenesis (Gillman et al., Gene 8:81 (1979); Roberts et at, Nature 328:731 (1987) or Innis (Ed.), 1990, PCR Protocols: A Guide to Methods and Applications, Academic Press, New York, N.Y.) or the polymerase chain reaction (PCR) method (Saiki et al, Science 239:487 (1988)), as exemplified by Daugherty et at (Nucleic Acids Res. 19:2471 (1991)) to modify nucleic acids encoding the p62 polypeptides of invention.
  • site-directed mutagenesis Gillman et al., Gene 8:81 (1979); Roberts et at, Nature 328:731 (1987) or Innis (Ed.), 1990, PCR Protocols: A Guide to Methods and Applications, Academic Press, New York,
  • polypeptides of the present invention may contain a heterologous signal sequence at its N-terminus.
  • expression and/or secretion of the fusion protein can be increased through use of a heterologous signal sequence.
  • Signal sequences are typically characterized by a core of hydrophobic amino acids, which are generally cleaved from the mature protein during secretion in one or more cleavage events.
  • Such signal peptides contain processing sites that allow cleavage of the signal sequence from the mature proteins as they pass through the secretory pathway.
  • the invention pertains to the described polypeptides having a signal sequence, as well as to polypeptides from which the signal sequence has been proteolytically cleaved (i.e., the cleavage products).
  • the polypeptides of the present invention can also be modified to incorporate one or more polymorphisms in the amino acid sequence resulting from natural allelic variation.
  • D-amino acids, non-natural amino acids or non-amino acid analogues can be substituted or added to produce a modified p62 polypeptide within the scope of this invention.
  • polypeptides of the present invention may be produced by expression of a nucleotide sequence coding for same in a suitable expression system.
  • polypeptides can be produced using chemical methods to synthesize the desired amino acid sequence, in whole or in part.
  • polypeptides can be synthesized by solid phase techniques, cleaved from the resin, and purified by preparative high performance liquid chromatography (e.g., Creighton (1983) Proteins Structures And Molecular Principles, WH Freeman and Co, New York N.Y.).
  • the composition of the synthetic polypeptides may be confirmed by amino acid analysis or sequencing (e.g., the Edman degradation procedure).
  • the amino acid sequence of a p62 polypeptide, or any part thereof may be altered during direct synthesis and/or combined using chemical methods with a sequence from other subunits, or any part thereof, to produce a variant polypeptide.
  • fusion proteins refers to chimeric proteins comprising amino acid sequences of two or more different proteins. Typically, fusion proteins result from in vitro recombinant techniques well known in the art.
  • the fusion proteins of the present invention may further comprise one or more additional polypeptide domains added to facilitate protein purification, to increase expression of the recombinant protein, or to increase the solubility of the recombinant protein.
  • additional polypeptide domains include, but are not limited to, metal chelating peptides such as histidine-tryptophan modules that allow purification on immobilized metals (Porath J (1992) Protein Expr Purif 3-.26328 1), protein A domains that allow purification on immobilized immunoglobulin, and the domain utilized in the FLAGS extension/affinity purification system (Immunex Corp, Seattle, Wash.).
  • the inclusion of a cleavable linker sequence such as Factor Xa or enterokinase (Invitrogen, San Diego, Calif.) between the purification domain and the protein is useful to facilitate purification.
  • Additional fusion expression vectors include pGEX (Pharmacia, a Piscataway, N.J.), pMAL (New England Biolabs, Beverly, Mass.) and pRITS (GE Healthcare Biosciences, Piscataway, N.J.) which fuse glutathione S transferase (GST), maltose B binding protein, or protein A, respectively, to the target recombinant protein.
  • GST glutathione S transferase
  • EBV, BKV, and other episomal expression vectors (ThermoFisher Scientific) can also be used.
  • p62 polypeptides can be non-covalently linked to a transport moiety or transfection agent.
  • a non-covalently linked peptide transfection agent is the Chariot protein delivery system (See U.S. Pat. No. 6,841,535; Morris et al. (1999) J. Biol. Chem. 274(35):24941-24946; and Morris et al. (2001) Nature Biotech. 19:1173-1176).
  • nucleic acid molecule encoding p62 polypeptide is utilized.
  • the nucleic acid molecule may comprise or consist of a nucleotide sequence encoding one or more p62 polypeptides, or fragments (including fragments that code for domains in any order or polypeptides wherein one or more domains are deleted or disrupted) or derivatives thereof, such as that contained in a DNA insert in an ATCC Deposit.
  • Variants of the p62 a nucleic acid molecules encoding p62 polypeptide can have at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 1.
  • nucleic acid sequence refers to a DNA or RNA sequence.
  • the term encompasses molecules formed from any of the known base analogs of DNA and RNA such as, but not limited- to 4-acetylcytosine, 8-hydroxy-N6-methyladenosine, aziridinyl-cytosine, pseudoisocytosine, 5-(carboxyhydroxylmethyl) uracil, 5-fluorouracil, 5-bromouracil, 5-carboxymethylaminomethyl-2-thiouracil, 5-carboxy-methylaminomethyluracil, dihydrouracil, inosine, N6-iso-pentenyladenine, 1-methyladenine, 1-methylpseudouracil, 1-methylguanine, 1-methylinosine, 2,2-dimethyl-guanine, 2-methyladenine, 2-methylguanine, 3-methylcytosine, 5-methylcytosine, N6-methyladenine, 7-methylgu
  • vectors are used to transfer a polynucleotide encoding a polypeptide to a cell.
  • a vector is any molecule used to transfer a nucleic acid sequence to a host cell.
  • an expression vector is utilized.
  • An expression vector is a nucleic acid molecule that is suitable for introduction to and/or propagation in a host cell and contains nucleic acid sequences that direct and/or control the expression of the transferred nucleic acid sequences. Expression includes, but is not limited to, processes such as transcription, translation, and splicing, if introns are present.
  • Expression vectors typically comprise one or more flanking sequences operably linked to a heterologous nucleic acid sequence encoding a polypeptide.
  • Flanking sequences may be homologous (i.e., from the same species and/or strain as the host cell), heterologous (i.e., from a species other than the host cell species or strain), hybrid (i.e., a combination of flanking sequences from more than one source), or synthetic, for example.
  • flanking sequence is capable of effecting the replication, transcription and/or translation of the coding sequence and is operably linked to a coding sequence.
  • operably linked refers to a linkage of polynucleotide elements in a functional relationship.
  • a promoter or enhancer is operably linked to a coding sequence if it affects the transcription of the coding sequence.
  • a flanking sequence need not necessarily be contiguous with the coding sequence, so long as it functions correctly. Thus, for example, intervening untranslated yet transcribed sequences can be present between a promoter sequence and the coding sequence and the promoter sequence may still be considered operably linked to the coding sequence.
  • an enhancer sequence can be located upstream or downstream from the coding sequence and affect transcription of the sequence.
  • the flanking sequence is a transcriptional regulatory region that drives high-level gene expression in the target cell.
  • the transcriptional regulatory region can include, for example, a promoter, enhancer, silencer, repressor element, or combinations thereof.
  • the transcriptional regulatory region can be constitutive, tissue-specific, cell-type specific (i.e., the region is drives higher levels of transcription in a one type of tissue or cell as compared to another), or regulatable (i.e., responsive to interaction with a molecule).
  • the source of a transcriptional regulatory region may be any prokaryotic or eukaryotic organism, any vertebrate or invertebrate organism, or any plant, provided that the flanking sequence functions in a cell by causing transcription of a nucleic acid within that cell.
  • a wide variety of transcriptional regulatory regions can be utilized.
  • Suitable transcriptional regulatory regions include, for example, the CMV promoter (i.e., the CMV-immediate early promoter); promoters from eukaryotic genes (i.e., the estrogen-inducible chicken ovalbumin gene, the interferon genes, the gluco-corticoid-inducible tyrosine aminotransferase gene, and the thymidine kinase gene); and the major early and late adenovirus gene promoters; the SV40 early promoter region (Bernoist and Chambon, 1981, Nature 290:304-10); the promoter contained in the 3′ long terminal repeat (LTR) of Rous sarcoma virus (RSV) (Yamamoto, et al., 1980, Cell 22:787-97); the herpes simplex virus thymidine kinase (HSV-TK) promoter (Wagner et al., 1981, Proc.
  • CMV promoter i.e., the C
  • Tissue- and/or cell-type specific transcriptional control regions include, for example, the elastase I gene control region which is active in pancreatic acinar cells (Swift et al., 1984, Cell 38:639-46; Ornitz et al., 1986, Cold Spring Harbor Symp. Quant. Biol.
  • the beta-globin gene control region in myeloid cells (Mogram et al., 1985, Nature 315:338-40; Kollias et al., 1986, Cell 46:89-94); the myelin basic protein gene control region in oligodendrocyte cells in the brain (Readhead et al., 1987, Cell 48:703-12); the myosin light chain-2 gene control region in skeletal muscle (Sani, 1985, Nature 314:283-86); the gonadotropic releasing hormone gene control region in the hypothalamus (Mason et al., 1986, Science 234:1372-78), and the tyrosinase promoter in melanoma cells (Hart, I.
  • Inducible promoters that are activated in the presence of a certain molecule or condition such as light, heat, radiation, tetracycline, or heat shock proteins, for example, can also be used (see, for example, WO 00/10612).
  • Other suitable promoters are known in the art.
  • enhancers may also be suitable flanking sequences.
  • Enhancers are cis-acting elements of DNA, usually about 10-300 bp in length, that act on the promoter to increase transcription. Enhancers are typically orientation- and position-independent, having been identified both 5′ and 3′ to controlled coding sequences.
  • enhancer sequences available from mammalian genes are known (i.e., globin, elastase, albumin, alpha-feto-protein and insulin).
  • the SV40 enhancer, the cytomegalovirus early promoter enhancer, the polyoma enhancer, and adenovirus enhancers are useful with eukaryotic promoter sequences.
  • an enhancer may be spliced into the vector at a position 5′ or 3′ to nucleic acid coding sequence, it is typically located at a site 5′ from the promoter.
  • Other suitable enhancers are known in the art, and would be applicable to the present invention.
  • vaccines that include a p62 polynucleotide or p62 polypeptide.
  • Such vaccines can further include an adjuvant.
  • Any of a variety of adjuvants can be employed in the vaccines of this invention to enhance the immune response.
  • Most adjuvants contain a substance designed to protect the antigen from rapid catabolism, such as aluminum hydroxide or mineral oil, and a nonspecific stimulator of immune responses.
  • Suitable adjuvants are commercially available and include, for example, Freund's Incomplete Adjuvant and Freund's Complete Adjuvant (Difco Laboratories) and Merck Adjuvant 65 (Merck and Company, Inc., Rahway, N.J.).
  • Suitable adjuvant types include, but are not limited to, gel-type, microbial, particulate, oil-emulsion, surfactant-based, and synthetic adjuvants.
  • a p62 polypeptide or polynucleotide encoding a p62 polypeptide, or derivative thereof with one or more co-stimulatory component(s) such as cell surface proteins, cytokines, chemokines, or signaling molecules in a composition of the present invention.
  • the co-stimulatory component may be included in the composition as a polypeptide or as a nucleic acid encoding the polypeptide, for example.
  • Suitable co-stimulatory molecules include, for instance, polypeptides that bind members of the CD28 family (i.e., CD28, ICOS; Hutloff, et al. Nature 1999, 397: 263-265; Peach, et al.
  • CD28 binding polypeptides B7.1 CD80; Schwartz, 1992; Chen et al, 1992; Ellis, et al. J. Immunol., 156(8): 2700-9) and B7.2 (CD86; Ellis, et al. J. Immunol., 156(8): 2700-9); polypeptides which bind members of the integrin family (i.e., LFA-1 (CD11a/CD18); Sedwick, et al. J Immunol 1999, 162: 1367-1375; Wulfing, et al. Science 1998, 282: 2266-2269; Lub, et al.
  • CD2 family members i.e., CD2, signaling lymphocyte activation molecule (CDw150 or “SLAM”; Aversa, et al. J Immunol 1997, 158: 4036-4044)
  • CD58 LFA-3; CD2 ligand; Davis, et al. Immunol Today 1996, 17: 177-187) or SLAM ligands (Sayos, et al. Nature 1998, 395: 462-469); polypeptides which bind heat stable antigen (HSA or CD24; Zhou, et al.
  • polypeptides which bind to members of the TNF receptor (TNFR) family i.e., 4-1BB (CD137; Vinay, et al. Semin Immunol 1998, 10: 481-489), OX40 (CD134; Weinberg, et al. Semin Immunol 1998, 10: 471-480; Higgins, et al. J Immunol 1999, 162: 486-493), and CD27 (Lens, et al. Semin Immunol 1998, 10: 491-499)
  • 4-1 BBL (4-1 BB ligand; Vinay, et al. Semin Immunol 1998, 10: 481-48; DeBenedette, et al.
  • CD154 CD40 ligand or “CD40L”; Gurunathan, et al. J. Immunol., 1998, 161: 4563-4571; Sine, et al. Hum. Gene Ther., 2001, 12: 1091-1102) may also be suitable.
  • Additional strategies for improving the efficiency of nucleic acid-based immunization may also be used including, for example, the use of self-replicating viral replicons (Caley, et al. 1999. Vaccine, 17: 3124-2135; Dubensky, et al. 2000. Mol. Med. 6: 723-732; Leitner, et al. 2000. Cancer Res. 60: 51-55), codon optimization (Liu, et al. 2000. Mol. Ther., 1: 497-500; Dubensky, supra; Huang, et al. 2001. J. Virol. 75: 4947-4951), in vivo electroporation (Widera, et al. 2000. J. Immunol.
  • Nucleic acids encoding p62 polypeptides can be administered to subjects by any of several available techniques.
  • Various viral vectors that have been successfully utilized for introducing a nucleic acid to a host include retrovirus, adenovirus, adeno-associated virus (AAV), herpes virus, and poxvirus, among others. It is understood in the art that many such viral vectors are available in the art.
  • the vectors can be constructed using standard recombinant techniques widely available to one skilled in the art. Many such techniques are disclosed in Molecular Cloning: A Laboratory Manual (Sambrook, et al., 1989, Cold Spring Harbor Laboratory Press), Gene Expression Technology (Methods in Enzymology, Vol. 185, edited by D. Goeddel, 1991. Academic Press, San Diego, Calif.), and PCR Protocols: A Guide to Methods and Applications (Innis, et al. 1990. Academic Press, San Diego, Calif.).
  • Suitable retroviral vectors include derivatives of lentivirus as well as derivatives of murine or avian retroviruses.
  • suitable retroviral vectors include, for example, Moloney murine leukemia virus (MoMuLV), Harvey murine sarcoma virus (HaMuSV), murine mammary tumor virus (MuMTV), SIV, BIV, HIV and Rous Sarcoma Virus (RSV).
  • MoMuLV Moloney murine leukemia virus
  • HaMuSV Harvey murine sarcoma virus
  • MuMTV murine mammary tumor virus
  • SIV BIV
  • HIV Rous Sarcoma Virus
  • retroviral vectors can incorporate multiple exogenous nucleic acid sequences.
  • helper cell lines encoding retrovirus structural genes.
  • Suitable helper cell lines include.
  • Retroviral vectors may be administered by traditional methods (i.e., injection) or by implantation of a “producer cell line” in proximity to the target cell population (Culver, K., et al., 1994, Hum. Gene Ther., 5 (3): 343-79; Culver, K., et al., Cold Spring Harb. Symp; Quant. Biol., 59: 685-90); Oldfield, E., 1993, Hum. Gene Ther., 4 (1): 39-69).
  • the producer cell line is engineered to produce a viral vector and releases viral particles in the vicinity of the target cell. A portion of the released viral particles contact the target cells and infect those cells, thus delivering a nucleic acid of the present invention to the target cell. Following infection of the target cell, expression of the nucleic acid of the vector occurs.
  • Adenoviral vectors are useful for gene transfer into eukaryotic cells (Rosenfeld, M., et al., 1991, Science, 252 (5004): 431-4; Crystal, R., et al., 1994, Nat. Genet., 8 (1): 42-51). Routes for administrating recombinant adenovirus to different tissues in vivo have included intratracheal instillation (Rosenfeld, M., et al., 1992, Cell, 68 (1): 143-55) injection into muscle (Quantin, B., et al., 1992, Proc. Natl. Acad. Sci.
  • Adeno-associated virus demonstrates high-level infectivity, broad host range and specificity in integrating into the host cell genome (Hermonat, P., et al., 1984, Proc. Natl. Acad. Sci. U.S.A., 81 (20): 6466-70).
  • Herpes Simplex Virus type-1 (HSV-1) is yet another attractive vector system, especially for use in the nervous system because of its neurotropic property (Geller, A., et al., 1991, Trends Neurosci., 14 (10): 428-32; Glorioso, et al., 1995, Mol. Biotechnol., 4 (1): 87-99; Glorioso, et al., 1995, Annu. Rev. Microbiol., 49: 675-710).
  • Poxvirus is another useful expression vector (Smith, et al. 1983, Gene, 25 (1): 21-8; Moss, et al, 1992, Biotechnology, 20: 345-62; Moss, et al, 1992, Curr. Top. Microbiol. Immunol., 158: 25-38; Moss, et al. 1991. Science, 252: 1662-1667).
  • Poxviruses shown to be useful include vaccinia, NYVAC, avipox, fowlpox, canarypox, ALVAC, and ALVAC(2), among others.
  • NYVAC (vP866) was derived from the Copenhagen vaccine strain of vaccinia virus by deleting six nonessential regions of the genome encoding known or potential virulence factors (see, for example, U.S. Pat. Nos. 5,364,773 and 5,494,807). The deletion loci were also engineered as recipient loci for the insertion of foreign genes.
  • NYVAC (vP866), vP994, vCP205, vCP1433, placZH6H4L reverse, pMPC6H6K3E3 and pC3H6FHVB were also deposited with the ATCC under the terms of the Budapest Treaty, accession numbers VR-2559, VR-2558, VR-2557, VR-2556, ATCC-97913, ATCC-97912, and ATCC-97914, respectively
  • ALVAC-based recombinant viruses are also suitable vectors (see, for example, U.S. Pat. No. 5,756,103).
  • ALVAC(2) is identical to ALVAC(1) except that ALVAC(2) genome comprises the vaccinia E3L and K3L genes under the control of vaccinia promoters (U.S. Pat. No. 6,130,066; Beattie et al., 1995a, 1995b, 1991; Chang et al., 1992; Davies et al., 1993).
  • ALVAC was deposited under the terms of the Budapest Treaty with the American Type Culture Collection (ATCC), 10801 University Boulevard, Manassas, Va. 20110-2209, USA, ATCC accession number VR-2547.
  • TROVAC refers to an attenuated fowlpox that was a plaque-cloned isolate derived from the FP-1 vaccine strain of fowlpoxvirus which is licensed for vaccination of 1 day old chicks. TROVAC was likewise deposited under the terms of the Budapest Treaty with the ATCC, accession number 2553.
  • Non-viral plasmid vectors may also be suitable in practicing the present invention. Suitable plasmid vectors are compatible with bacterial, insect, and/or mammalian host cells. Such vectors include, for example, PCR-II, pCR3, and pcDNA3.1 (ThermoFisher), pBSII (Agilent Technology Santa Clara CApET15 (EMD Millipore, Bullerica, Mass.) pGEX (GE HealthCare Bioscience, Piscataway, N.J.), pEGFP-N2 (Clontech, Palo Alto, Calif.), pETL (BlueBacII, Thermofisher), pDSR-alpha (PCT pub. No.
  • WO 90/14363 and pFastBacDual (ThermoFisher)) as well as Bluescript® plasmid derivatives (a high copy number COLE1-based phagemid, (Agilent Technology Santa Clara, Calif.), PCR cloning plasmids designed for cloning Taq-amplified PCR products (e.g., TOPOTM TA Cloning® kit, PCR2.1® plasmid derivatives, Thermofisher Bacterial vectors may also be used with the current invention.
  • Bluescript® plasmid derivatives a high copy number COLE1-based phagemid, (Agilent Technology Santa Clara, Calif.
  • PCR cloning plasmids designed for cloning Taq-amplified PCR products e.g., TOPOTM TA Cloning® kit, PCR2.1® plasmid derivatives, Thermofisher Bacterial vectors may also be used with
  • vectors include, for example, Shigella, Salmonella, Vibrio cholerae, Laclobacillus, Bacille calmette guerin (BCG), and Streptococcus (see for example, WO 88/6626; WO 90/0594; WO 91/13157; WO 92/1796; and WO 92/21376).
  • BCG Bacille calmette guerin
  • Streptococcus see for example, WO 88/6626; WO 90/0594; WO 91/13157; WO 92/1796; and WO 92/21376).
  • Many other non-viral plasmid expression vectors and systems are known in the art and could be used with the current invention.
  • Suitable nucleic acid delivery techniques include DNA-ligand complexes, adenovirus-ligand-DNA complexes, direct injection of DNA, CaPO4 precipitation, gene gun techniques, electroporation polymer gene delivery system, cell-penetrating peptide gene delivery system, and colloidal dispersion systems, among others.
  • Polymer gene delivery system includes polyetherimide- and pluronic-based delivery systems.
  • Cell-penetrating peptide-based systems include 9-35 mer cationic and/or amphipathic peptides capable of mediating translocation of DNA across plasma membrane.
  • Colloidal dispersion systems include macromolecule complexes, nanocapsules, microspheres, beads, and lipid-based systems including oil-in-water emulsions, micelles, mixed micelles, and liposomes.
  • Liposomes are artificial membrane vesicles useful as delivery vehicles in vitro and in vivo. RNA, DNA and intact virions can be encapsulated within the aqueous interior and be delivered to cells in a biologically active form (Fraley, R., et al., 1981, Trends Biochem. Sci., 6: 77).
  • the composition of the liposome is usually a combination of phospholipids, particularly high-phase-transition-temperature phospholipids, usually in combination with steroids, especially cholesterol.
  • phospholipids or other lipids may also be used.
  • the physical characteristics of liposomes depend on pH, ionic strength, and the presence of divalent cations.
  • lipids useful in liposome production include phosphatidyl compounds, such as phosphatidylglycerol, phosphatidylcholine, phosphatidylserine, phosphatidylethanolamine, sphingolipids, cerebrosides, and gangliosides.
  • the lipid can be a diacylphosphatidylglycerol.
  • The, lipid moiety of diacylphosphatidylglycerol can have 14-18 carbon atoms.
  • the lipid moiety of diacylphosphatidylglycerol can have 16-18 carbon atoms.
  • the lipid moiety of diacylphosphatidylglycerol can be saturated.
  • Illustrative phospholipids include egg phosphatidylcholine, phosphatidylethanolamine, sphingolipids, cerebrosides, and gangliosides.
  • a method for the treatment of inflammatory diseases is provided.
  • the method for the treatment of inflammatory diseases that are not related to cancer is provided.
  • the treatment of inflammatory diseases comprises administering a therapeutically effective amount of p62 polypeptides or p62 encoding nucleic acids to a subject in need thereof, in such amounts and for such time as is necessary to achieve the desired result.
  • a “therapeutically effective amount” of an inventive targeted particle is that amount effective for treating, alleviating, ameliorating, relieving, delaying onset of, inhibiting progression of, reducing severity of, and/or reducing incidence of one or more symptoms or features of inflammatory diseases.
  • a method for administering p62 polypeptides or p62 encoding nucleic acids to a subject suffering from inflammatory disease or relapse is provided.
  • p62 polypeptides or p62 encoding nucleic acids are administered to a subject in such amounts and for such time as is necessary to achieve the desired result (i.e. treatment of inflammatory disease).
  • a “therapeutically effective amount” of p62 polypeptides and p62 encoding nucleic acids is that amount effective for treating, alleviating, ameliorating, relieving, delaying onset of, inhibiting progression of, reducing severity of, and/or reducing incidence of one or more symptoms or features of inflammatory disease.
  • the p62 polypeptides or p62 encoding nucleic acids of the invention are administered to a subject previously treated for inflammatory disease. In some embodiments, the p62 polypeptides or p62 encoding nucleic acids of the invention are administered to a subject with a family history of inflammatory disease. In some embodiments, the p62 polypeptides or p62 encoding nucleic acids of the invention are administered to a subject with a predisposition for inflammatory diseases. For example, a subject who is genetically predisposed to inflammatory diseases (ALS, Parkinson disease, Huntington disease), or subjected to environmental factors provoking inflammatory disease (e.g., tobacco smoke, asbestos, silica particles).
  • ALS genetic predisposition to ALS is associated with mutations in genes of superoxidismutase or TAR DNA-binding protein 43 (TDP-43). Genetic predisposition to Parkinson disease is associated with the parkin and synuclein genes. Genetic predisposition to Huntington disease is associated with a mutation in huntingtin gene (Glass, C. K., K. Saijo, et al. (2010). “Mechanisms Underlying Inflammation in Neurodegeneration.” Cell 140(6): 918-934). Tobacco smoke, asbestos, silica particles are all well-known inducers of chronic inflammation as persistent allergens and undigestable foreign particles; they lead to chronic obstructive pulmonary disease, asbestosis and silicosis (Medzhitov 2010. Inflammation 2010: New Adventures of an Old Flame. Cell 140:771-776)
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of an inflammatory disease, disorder, and/or condition.
  • the inflammatory disease, disorder, and/or condition is non-cancer-related.
  • Such diseases include, but not limited to, osteoporosis, obesity, metabolic syndrome, type 2 diabetes, fat liver, inflammatory bowel disease, gastritis, chronic pancreatitis, asthma, chronic obstructive pulmonary disease (COPD), rheumatoid arthritis (RA), osteoarthritis, multiple sclerosis (MS), psoriasis, congestive heart failure (CHF), atherosclerosis, neurodegenerative diseases (ALS, Parkinson, Alzheimer's, Huntington disease), gout, asbestosis and silicosis.
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of osteoporosis.
  • Osteoporosis is the most common disease of the bone associated with bone loss and affecting mostly women after onset of menopause. Menopause leads to decrease in estrogen levels, thus ovariectomy in rodents leading to cessation of estrogen generation is the most common model for osteoporosis.
  • Postmenopausal period is marked by elevation of cytokines such us IL-6, TNF-alpha and IL-1 beta, and the same cytokines are elevated under ovariectomy.
  • TNF and IL-1 have potent antiapoptotic effects in osteoclasts prolonging OC lifespan, accelerating bone resorption and inhibiting bone formation, and blockade of TNF-alpha and IL-1beta prevents osteoporosis due to estrogendeficiency. (Mundy 2007. Osteoporosis and Inflammation. Nutrition Reviews 65:S147-S151; Lencel and Magne 2011. Inflammaging: The driving force in osteoporosis? Medical Hypotheses 76:317-321).
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of obesity, type 2 diabetes and fat liver disease.
  • Obesity is increasingly prevalent in the population and strongly associated with the development of insulin resistance, an underlying feature of both type 2 diabetes (T2D) and metabolic syndrome.
  • Insulin resistance has been recognized as the integral feature of metabolic syndrome, which includes glucose intolerance, insulin resistance, obesity, hypertriglyceridemia, low HDL cholesterol, hypertension, and accelerated atherosclerosis.
  • TNF- ⁇ The proinflammatory cytokine TNF- ⁇ has been demonstrated to mediate insulin resistance as a result of obesity in many rodent obesity models.
  • various other inflammatory mediators and cytokines are also overexpressed in adipose and other tissues in experimental mouse models of obesity and in humans.
  • Proinflammatory cytokines can cause insulin resistance and anti-inflammatory medications may reverse it suggesting that inflammation may be directly involved in its (Hotamisligil 2006. Inflammation and metabolic disorders. Nature 444:860-867).
  • Non-alcoholic fatty liver disease is recognized as the hepatic manifestation of metabolic syndrome, and is characterized by the accumulation of fatty infiltrations affecting >5% of the liver.
  • the clinical implications of NAFLD are derived by its potential to progress to steatohepatitis (NASH), fibrosis, cirrhosis, and in some cases, hepatocellular carcinoma.
  • NASH steatohepatitis
  • fibrosis fibrosis
  • cirrhosis cirrhosis
  • hepatocellular carcinoma hepatocellular carcinoma.
  • the prevalence of NAFLD is rising in parallel with the increasing rate of obesity, and this global trend is attributed to the diet of the Western lifestyle.
  • the pathogenesis of NAFLD is not well understood, but is proposed to be a “two-hit” process. The first “hit” leads to lipid accumulation and steatosis.
  • lipid accumulation Mechanisms of this lipid accumulation are unclear, but likely involve dysregulated lipid homeostasis including beta-oxidation, very low density lipoprotein secretion, de novo lipogenesis, and lipid trafficking and storage.
  • This hepatic steatosis sensitizes the liver to a “second hit” leading to inflammation, a key pathophysiologic feature of steatohepatitis, and progressive liver disease (Renaud et al. 2014. Effect of Diet on Expression of Genes Involved in Lipid Metabolism, Oxidative Stress, and Inflammation in Mouse Liver-Insights into Mechanisms of Hepatic Steatosis. PLoS ONE 9:e88584)
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of IBD, including, but not limited to ulcerative colitis and Crohn's disease.
  • IBD Inflammatory Bowel Disease
  • IBD is a chronic inflammatory disorder of gastrointestinal tract. Examples of IBD include ulcerative colitis and Crohn's disease.
  • Substantial evidence from human genetic studies and from preclinical IBD models suggests that failures by either the mucosal epithelial layer or the immune system to properly interact with the lumen microbial community may underlie the pathogenic processes.
  • IBD is also an important risk factor for development of colon carcinoma, where proinflammatory cytokines TNFa, interleukin-1, IL-6 plays a significant role role (Danese and Mantovani 2010. Inflammatory bowel disease and intestinal cancer: a paradigm of the Yin-Yang interplay between inflammation and cancer. Oncogene 29:3313-3323)
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of asthma.
  • Asthma is an inflammatory disease in the airway, leading to airway hyperresponsiveness, obstruction, mucus hyper-production and airway wall remodeling.
  • Asthma is classically recognized as the typical Th2 disease, with increased IgE levels and eosinophilic inflammation in the airway. Emerging Th2 cytokines modulates the airway inflammation, which induces airway remodeling.
  • the relatively simple paradigm has been doubted because of the realization that strategies designed to suppress Th2 function are not effective enough for all patients in the clinical trials. (Kudo et al. 2013. Pathology of asthma. Frontiers in Microbiology 4:263).
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of COPD (chronic obstructive pulmonary disease).
  • COPD chronic inflammation affecting predominantly lung parenchyma and peripheral airways and results in largely irreversible and progressive airflow limitation. This inflammation is characterized by increased numbers of alveolar macrophages, neutrophils, and T lymphocytes, which are recruited from the circulation. Oxidative stress plays a key role in driving this inflammation. The pulmonary inflammation may enhance the development and growth of lung cancer.
  • peripheral inflammation extends into the circulation, resulting in systemic inflammation with the same inflammatory proteins.
  • Systemic inflammation may worsen comorbidities. Treatment of pulmonary inflammation may therefore have beneficial effects. (Barnes 2014. Cellular and Molecular Mechanisms of Chronic Obstructive Pulmonary Disease. Clinics in Chest Medicine 35:71-86).
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of Rheumatoid Arthritis (RA).
  • RA is a chronic, inflammatory, systemic autoimmune disease that affects about 1% of the general population in Western countries and is two to three times more common in women than in men.
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of osteoarthritis (OA) is a disease of the joints that affects several million individuals worldwide. This disease, which involves mainly the diarthrodial joints, is chronic and develops slowly over decades. The role of synovial inflammation in the pathophysiology of OA is now widely accepted. Synovitis has been considered secondary to the cartilage changes yet findings indicate that synovial inflammation could be a component of the early events leading to the clinical stage of OA.
  • OA osteoarthritis
  • Synovial inflammation leads to the production and release of pro-inflammatory cytokines and several other inflammatory mediators. Some of these factors, including the pro-inflammatory cytokines, diffuse through the synovial fluid into the cartilage, where they activate chondrocyte production of the catabolic factors through auto- and paracrine mechanisms.
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of Multiple Sclerosis (MS).
  • MS is characterized by inflammation, demyelination, and axon degeneration in the CNS. Individuals are plagued by MS-associated comorbidities, such as chronic pain, fatigue, depression, sleep disorders, spasticity, gait and coordination imbalances, migraines, sensory organ dysfunctions, and overall cognitive impairment (Damal et al. 2013.
  • MS is considered an immune-mediated disease characterized by the presence of inflammatory demyelinating lesions in the CNS. Infection by bacteria or viruses or other environmental stimuli trigger the activation of microglia and astrocytes in multiple sclerosis (MS), leading to the production of proinflammatory cytokines through activation of the transcription factors NF-kappa-B and AP-1 (Luessi et al. 2012. Neurodegeneration in multiple sclerosis: novel treatment strategies. Expert Rev Neurother 12:1061-1076).
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of atherosclerosis.
  • Atherosclerosis shares features with traditional inflammatory diseases including rheumatoid arthritis. Atherosclerosis is the main cause of coronary artery and cerebrovascular disease, which are the leading cause of death worldwide. Atherogenesis is thought to begin with the development of endothelial dysfunction caused by the exposure of the vessel wall to systemic risk factors and local hemodynamics. The ensuing endothelial activation promotes the accumulation of inflammatory cells in the vessel wall.
  • inflammatory cells produce cytokines and growth factors, which evoke smooth muscle cell migration into the intima.
  • the architecture of the intima changes profoundly leading to the formation of two compartment lesions, the fibrous cap and the necrotic core.
  • Inflammatory cells may also produce matrix degrading enzymes that disrupt the integrity of the fibrous cap or procoagulant molecules such as tissue factor, ultimately leading to plaque rupture and thrombosis (Cole et al. 2011. Toll-like receptors in atherosclerosis: a “Pandora's box” of advances and controversies. Trends in Pharmacological Sciences 34:629-636).
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of gastritis.
  • a cause of gastritis is Helicobacter pylon colonizing the gastric mucosa causing chronic inflammation, which is characterized by enhanced expression of many inflammatory genes.
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of chronic pancreatitis (CP).
  • Chronic pancreatitis (CP) is a fibro-inflammatory disease involving the pancreatic parenchyma which is progressively destroyed and replaced by fibrotic tissues. Histologically, acinar cell damage, mononuclear cell infiltration, and fibrosis are observed. There are various causes that may lead to CP, but the exact pathophysiology of the disease is still unclear.
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of psoriasis.
  • Psoriasis is a chronic inflammatory skin disease, most commonly resulting in the occurrence of red and silver scaly plaques; it affects approximately 2-3% of the general population. Although its pathogenesis is not fully understood, there is an underlying interaction between numerous immune effector cells and aberrant hyperproliferation and differentiation of epidermal keratinocytes.
  • NF-kappB An essential transcription factor in psoriasis. Journal of Dermatological Science 69:89-94).
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of congestive heart failure (CHF).
  • CHF congestive heart failure
  • the heart failure syndrome is characterized by impaired systolic and/or diastolic function and various clinical signs such as fatigue, dyspnea, fluid retention, and cachexia. An inflammatory activation in CHF patients has long been recognized.
  • TNF-alpha tumor necrosis factor-alpha
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of neurodegenerative disease.
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of Alzheimer's disease (AD).
  • AD Alzheimer's disease
  • AD Alzheimer's disease
  • AD is the most common form of dementia in the elderly resulting in a progressive decline in a number of cognitive functions including short-term memory.
  • AD is characterized by the formation of two characteristic lesions: extracellular beta-amyloid deposits forming senile plaques and intracellular neurofibrillary tangles made up of the microtubule associated protein tau.
  • a strong link between inflammation, primarily mediated by pro-inflammatory cytokines, and AD has been established both in clinical data and bench research. Recent findings also suggest that AD may be associated with a more widespread inflammatory state characterized by increased peripheral blood levels of IL-1, IL-6, TNF-alpha, TGF-beta, and IL-18 (Smith et al. 2012. Role of pro-inflammatory cytokines released from microglia in neurodegenerative diseases. Brain Res Bull 87:10-20).
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of Parkinson's disease (PD).
  • PD is the most common neurodegenerative movement disorder and is caused by the progressive loss of dopaminergic neurons from the substantia nigra pars compacta that normally innervate the striatum.
  • the pathological hallmark of PD is intracellular accumulation of alpha-synuclein leading to the formation of Lewy bodies.
  • PD may result in a number of different presenting symptoms including resting tremor, bradykinesia, cogwheel rigidity, and postural instability.
  • Epidemiologic findings from a number of studies suggest that inflammation may be involved in the pathogenesis of PD. This is also supported, in part, by post-mortem analysis of cerebrospinal fluid and brain demonstrating elevated protein levels of pro-inflammatory cytokines in PD patients (Smith et al. 2012. Role of pro-inflammatory cytokines released from microglia in neurodegenerative diseases. Brain Res Bull 87:10-20).
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of Amyotrophic Lateral Sclerosis (ALS), or Lou Gehrig's disease.
  • ALS is a progressive fatal neurodegenerative disease that affects motor neurons in the brainstem, spinal cord, and motor cortex. ALS is universally fatal, with a median age of onset of 55 years and a survival of 2-5 years after the onset of symptoms.
  • Prominent neuroinflammation can be easily observed in pathologically affected areas of the CNS and in spinal cords from both human ALS patients and mouse models of the disease.
  • inflammation in ALS is characterized by gliosis and the accumulation of large numbers of activated microglia and astrocytes.
  • Activation of glia in ALS has been extensively characterized and is marked by elevated production of potentially cytotoxic molecules such as ROS, inflammatory mediators such as COX-2, and proinflammatory cytokines such as IL-1beta, TNF-alpha, and IL-6 (Smith et al. 2012. Role of pro-inflammatory cytokines released from microglia in neurodegenerative diseases. Brain Res Bull 87:10-20).
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of Huntington disease.
  • Huntington's disease is characterized by a progressive course of disease until death 15-20 years after the first symptoms occur and is caused by a mutation with expanded CAG repeats in the huntingtin (htt) protein. Mutant htt (mhtt) in the striatum is assumed to be the main reason for neurodegeneration. Both innate and adaptive immune systems may play an important role in HD.
  • Activation of microglia with expression of proinflammatory cytokines, impaired migration of macrophages, and deposition of complement factors in the striatum indicate an activation of the innate immune system (Ellrichmann et al. 2013. The Role of the Immune System in Huntington's Disease. Clinical and Developmental Immunology 2013:11). Enhancement of the NFkB-mediated inflammatory response in astrocytes contributes to HD pathogenesis (Hsiao et al. 2013. A critical role of astrocyte-mediated nuclear factor-kB-dependent inflammation in Huntington's disease. Human Molecular Genetics 22:1826-1842).
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of depression or schizophrenia.
  • Depression is associated with both a chronic low-grade inflammatory response, activation of cell-mediated immunity and activation of the compensatory anti-inflammatory reflex system (CIRS), characterized by negative immunoregulatory processes.
  • CIRS compensatory anti-inflammatory reflex system
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of asbestosis.
  • Asbestosis is a chronic inflammatory and fibrotic medical condition affecting the parenchymal tissue of the lungs caused by the inhalation and retention of asbestos fibers. It usually occurs after high intensity and/or long-term exposure to asbestos (particularly in those individuals working on the production or end-use of products containing asbestos) and is therefore regarded as an occupational lung disease.
  • Sufferers may experience severe dyspnea (shortness of breath) and are at an increased risk for certain malignancies, including lung cancer but especially mesothelioma.
  • Asbestosis specifically refers to interstitial (parenchymal) fibrosis from asbestos, and not pleural fibrosis or plaquing.
  • the primary symptom of asbestosis is generally the slow onset of dyspnea, especially on exertion.
  • Clinically advanced cases of asbestosis may lead to respiratory failure.
  • the characteristic pulmonary function finding in asbestosis is a restrictive ventilatory defect. In the more severe cases, the drastic reduction in lung function due to the stiffening of the lungs and reduced TLC may induce right-sided heart failure (cor pulmonale).
  • asbestosis may produce reduction in diffusion capacity and arterial hypoxemia.
  • p62 polypeptides or p62 encoding nucleic acids in accordance with the present invention may be used to treat, alleviate, ameliorate, relieve, delay onset of (prophylaxis), inhibit progression of, reduce severity of, and/or reduce incidence of one or more symptoms or features of silicosis.
  • Silicosis (previously miner's phthisis, grinder's asthma, potter's rot and other occupation-related names) is a form of occupational lung disease caused by inhalation of crystalline silica dust, and is marked by inflammation and scarring in the form of nodular lesions in the upper lobes of the lungs. It is a type of pneumoconiosis.
  • Silicosis (particularly the acute form) is characterized by shortness of breath, cough, fever, and cyanosis (bluish skin). Signs and symptoms include: dyspnea, cough, fatigue, tachypnea, loss of appetite and weight loss, chest pain, and fever. In advanced cases, symptoms may include cyanosis, cor pulmonale, and respiratory insufficiency. Subjects with silicosis are particularly susceptible to tuberculosis (TB) infection—known as silicotuberculosis.
  • TB tuberculosis
  • Pulmonary complications of silicosis also include Chronic Bronchitis and airflow limitation (indistinguishable from that caused by smoking), non-tuberculous Mycobacterium infection, fungal lung infection, compensatory emphysema, and pneumothorax.
  • silicosis and certain autoimmune diseases, including nephritis, Scleroderma, and Systemic Lupus Erythematosus, especially in acute or accelerated silicosis.
  • Anti-inflammatory chemotherapeutic agents are any chemical entity or drug that reduces inflammation.
  • Anti-inflammatory chemotherapeutic agents include, but are not limited to, a nonsteroidal anti-inflammatory drug (NSAID), a glucocorticoid, methotrexate, cyclosporine, and rapamycin.
  • NSAIDs are cyclooxygenase inhibitors. Examples of NSAIDs include aspirin, ibuprofen, naproxen sodium, diclofenac, etodolac, fenoprofen, flurbiprofen, oxaprozin.
  • Glucocorticoids are a class of steroid hormones that bind to the glucocorticoid receptor. Methotrexate is a chemical analog of folic acid which inhibits the metabolism of folic acid. Cyclosporine and rapamycin, anti-rejection drugs, have anti-inflammatory properties.
  • Anti-inflammatory biologic agents are any naturally-occurring biologic entity that reduces inflammation.
  • Anti-inflammatory biologic agents include, but are not limited to, an anti-TNF antibody, an anti-IL1 antibody, an anti-IL6 antibody, an anti-IL6 receptor antibody, an anti-IL12/23 antibody, an anti-IL17 antibody, an anti-IL1R antibody, an anti-IL1 receptor antagonist, and a soluble IL-1 receptor.
  • compositions of the invention can be administered as a pharmaceutical or medicament formulated with a pharmaceutically acceptable carrier. Accordingly, the compounds and compositions may be used in the manufacture of a medicament or pharmaceutical composition.
  • Pharmaceutical compositions of the invention may be formulated as solutions or lyophilized powders for parenteral administration. Powders may be reconstituted by addition of a suitable diluent or other pharmaceutically acceptable carrier prior to use. Liquid formulations may be buffered, isotonic, aqueous solutions. Powders also may be sprayed in dry form. Examples of suitable diluents are normal isotonic saline solution, standard 5% dextrose in water, or buffered sodium or ammonium acetate solution.
  • Such formulations are especially suitable for parenteral administration, but may also be used for oral administration or contained in a metered dose inhaler or nebulizer for insufflation. It may be desirable to add excipients such as polyvinylpyrrolidone, gelatin, hydroxy cellulose, acacia, polyethylene glycol, mannitol, sodium chloride, sodium citrate, and the like.
  • compounds may be encapsulated, tableted or prepared in an emulsion or syrup for oral administration.
  • Pharmaceutically acceptable solid or liquid carriers may be added to enhance or stabilize the composition, or to facilitate preparation of the composition.
  • Solid carriers include starch, lactose, calcium sulfate dihydrate, terra alba, magnesium stearate or stearic acid, talc, pectin, acacia, agar or gelatin.
  • Liquid carriers include syrup, peanut oil, olive oil, saline and water.
  • the carrier may also include a sustained release material such as glyceryl monostearate or glyceryl distearate, alone or with a wax.
  • the amount of solid carrier varies but, preferably, will be between about 20 mg to about 1 g per dosage unit.
  • the pharmaceutical preparations are made following the conventional techniques of pharmacy involving milling, mixing, granulating, and compressing, when necessary, for tablet forms; or milling, mixing and filling for hard gelatin capsule forms.
  • the preparation may be in the form of a syrup, elixir, emulsion, or an aqueous or non-aqueous suspension.
  • the invention compounds may be combined with excipients such as cocoa butter, glycerin, gelatin or polyethylene glycols and molded into a suppository.
  • Compounds or compositions may be formulated to include other medically useful drugs or biological agents.
  • the compounds or compositions also may be administered in conjunction with the administration of other drugs or biological agents useful for the disease or condition to which the invention compounds or compositions are directed.
  • an effective amount refers to a dose sufficient to provide concentrations high enough to impart a beneficial effect on the recipient thereof.
  • the specific therapeutically effective dose level for any particular subject will depend upon a variety of factors including the disorder being treated, the severity of the disorder, the activity of the specific compound or composition, the route of administration, the rate of clearance of the compound or composition, the duration of treatment, the drugs used in combination or coincident with the compound or composition, the age, body weight, sex, diet, and general health of the subject, and like factors well known in the medical arts and sciences.
  • a compound can be administered parenterally, such as intravascularly, intravenously, intraarterially, intramuscularly, subcutaneously, or the like. Administration can also be orally, nasally, rectally, transdermally, intravaginally or inhalationally via an aerosol.
  • a compound or composition can be administered to the inflammation-related lesions. The compound or composition may be administered as a bolus, or slowly infused, or be administered as an intradermal, subcutaneous or intramuscular or intraperitoneal injection.
  • a therapeutically effective dose can be estimated initially from cell culture assays by determining p62 expression levels upon introduction of the DNA or RNA encoding p62 into the cells. A dose can then be confirmed in animal models to achieve suppression of generation of inflammatory cytokines and/or alleviation of inflammation. Such information can be used to more accurately determine useful initial doses in humans. The exact formulation, route of administration and dosage can be chosen by medical professional in view of the subject's presentation.
  • Full length cDNA encoding the longer isoform of p62 (Transcript Variant 1, Gen Bank reference No. NP_003891) was amplified by PCR (HotStar HiFidelity Polymerase Kit Qiagen) using the following primers: FW: 5-CCCGCTAGCATGGCGTCGCTCACCGTG-3 and REV: 5′-CCCAAGCTTTCACAACGGCGGGGGATGCTTTG-3′.
  • PCR products were purified and Nhe I-Hind III digested fragments cloned into a DNA vector with nucleic acid sequence corresponding to pcDNA3.1 resulting in p62 plasmid.
  • a full length ovalbumin gene (pOVA) cloned in pcDNA3.1 was taken as reference plasmid.
  • Large scale preparations of the endotoxin-free plasmids were routinely performed by alkaline lysis using either Endo Free Plasmid Kit (Qiagen) or Gen Elute HPSelect Plasmid Giga Prep columns (SIGMA # NA0800). The DNA constructs were confirmed by sequencing.
  • SO sham operated
  • mice were sacrificed by CO2 narcosis according to the recommendation of the Italian Ethical Committee.
  • mice were ovariectomized (OVX) and left untreated for 2 months. Afterwards, mice were randomized in 4 subgroups, and injected with plasmid as described above. After 2 months mice were sacrificed for analysis.
  • OVX ovariectomized
  • Bone mineral density (BMD) and bone mineral content (BMC) were measured using a PIXImus DEXA.
  • BMSC Bone Marrow Cell
  • cytokine/chemokine profiles of BMSCs supernatants were assessed by ELISA-based cytokine array by using Mouse Cytokine Array Panel A kit (R&D Systems, Milano, Italy) accordingly to the manufacturer's instructions.
  • Immunoreactive dots were visualized using LiteAblot Turbo luminol reagents (Euroclone, Milano, Italy) and Hyperfilm-ECL film (Euroclone, Milano, Italy) and quantitated densitometrically.
  • Proteins from total bone marrow cells population were extracted in Cell Lysis Buffer (Cell Signaling Euroclone, Milano, Italy) immediately after flushing the bone marrow cavity, and the concentration was determined by the BCA protein assay reagent (Pierce, Euroclone Milano, Italy). Western blotting was performed by standard methods.
  • mice were first injected either with p62DNA or reference plasmids (pcDNA 3.1, pOVA) and then ovariectomized (OVX).
  • pcDNA 3.1, pOVA p62DNA or reference plasmids
  • OVX ovariectomized mice
  • SO sham operated mice
  • mice Two months after surgery mice were sacrificed, and the collected long bones subjected to histological examination.
  • the metaphyseal regions of the distal femurs from pcDNA3.1-OVX (control) mice displayed classic osteoporotic features characterized by significant bone loss and thinned disconnected trabecular structure.
  • p62-OVX bones (treated mice) showed a micro-architecture essentially indistinguishable to that seen in SO mice. Moreover, examination of cross sections femur diaphysis from p62DNA-OVX mice revealed (at variance of those obtained from reference plasmids treated mice) an enhanced anabolic-osteoblastic activity as evidenced by new cortical bone apposition suggesting an anabolic action of p62 treatment.
  • BMCs were flushed from the bone cavities, and cultured for 3 days. Afterwards, both supernatants and cells were collected and analyzed respectively either for the release of inflammatory cytokines, or for expression of osteogenic markers. As shown in FIG. 4 the marked up-regulation and release of pro-inflammatory cytokines by BMCs from OVX compare to SO operated mice was drastically suppressed by p62-DNA pre-treatment. The inhibitory effect of p62 DNA extended to an array of cytokines such as TNF ⁇ , IL-6, IL-1b IL-17, all known to be essential inducers of inflammatory diseases and bone loss.
  • cytokines such as TNF ⁇ , IL-6, IL-1b IL-17
  • mice were ovariectomized and, after two months, injected either with p62-DNA or reference plasmids (see M&M for details). Two months after last plasmids injections, bones were collected and histologically evaluated.
  • OVX-p62 treated mice group (in contrast to control groups) showed a restored trabecular microarchitecture at metaphyseal regions of the distal femurs and a decreased porosity in cortical bone.
  • p62-DNA treatment proved to increase both bone mineral density (BMD) and content (BMC) as judged by DXA analysis ( FIG. 6 ). Finally, coupled with marked up-regulation of osteoblastogenic Runx2 and Osterix ( FIG.
  • RANKL is a key mediator of inflammation that, by binding to its receptor RANK on osteoclast precursors, fosters osteoclastogenesis via intracellular NF-kB signaling.
  • Down-regulation of NF-kappa-B in OVX-p62 BMCs was also observed ( FIG. 7 , panel B). Consequently, p62 administration reversed osteoporosis in a mouse model.
  • ALS Amyotrophic Lateral Sclerosis
  • FIG. 9 shows the effect of p62 on a mouse model of ALS.
  • Neuroinflammation is a prominent pathologic feature in the spinal cord of patients with ALS, and is characterized by glial activation and infiltrating T cells.
  • a similar inflammatory response is present in spinal cords of ALS mice.
  • This is the most common mouse model of ALS expressing G93A mutant form of SOD, the same mutation which occurs in a humans with ALS, and this model is widely used for testing of drugs against ALS.
  • mice were treated with p62 plasmid or pcDNA 3.1 plasmids (6 times weekly with 150 ug/mouse i.m.) from day 75 after birth (6 mice per group), and a standard test for ALS (hind limb extension reflex) was applied. This reflex was evaluated as the ability to perform complete extension of the hind limbs when the animal was suspended by the tail. Without ALS, both control and p62 treated mice demonstrate strong reflex, which started to decline in ALS mice after 60 days of age, and it decreased from 100% to zero on 100 days. Treatment with p62, however, markedly delayed decline in the reflex. Consequently, administration of p62 plasmid reduced symptoms related to ALS in a mouse model.
  • ALS hind limb extension reflex
  • MS Multiple Sclerosis
  • MS is a chronic inflammatory disease resulting in demyelination and axonal loss throughout the central nervous system (CNS), with unknown cause and only limited treatment options (Noseworthy, J. H., C. Lucchinetti, et al. (2000)“Multiple sclerosis.” N Engl J Med 343(13): 938-52; Lassmann et al., 2001“Heterogeneity of multiple sclerosis pathogenesis: implications for diagnosis and therapy.” Trends Mol Med 7(3): 115-21).
  • the most commonly used animal model for MS research is murine experimental allergic encephalomyelitis (EAE) induced by administration of MOG polypeptides.
  • EAE murine experimental allergic encephalomyelitis
  • FIG. 10 demonstrates clinical signs of rMOG-induced EAE showing mean clinical scores.
  • mice (4 per group) were examined daily for clinical signs of EAE and were scored as followed: 0, no disease; 1, limp tail; 2, hind limb weakness; 3, complete hind limb paralysis; 4, hind limb paralysis plus forelimb paralysis; and 5, moribund or dead.
  • MOG immunization On day 0 MOG immunization.
  • p62 DNA 100 ⁇ g/50 ⁇ l administered at days 16 and 22 after MOG immunization as indicated by red arrows.
  • red arrows As one can see, whereas control mice had higher EAE score till the end of observation (day 31), in mice treated with p62 plasmid this score progressively declined (to about 0.5 at day 31). Consequently, administration of p62 plasmid alleviated MS-like symptoms in a mouse model of MS.
  • LLC is a widely used model of metastatic lung cancer: after subcutaneous inoculation of tumor cells in flanks of mice, within 3 weeks they form metastases in lungs which can be easily counted. Treatment of mice with p62 polynucleotide markedly suppresses formation of both small and big metastases thus effectively blocking metastatic process ( FIG. 13 ). Consequently, administration of p62 plasmid suppresses lung cancer in a mouse model.
  • B16 melanoma cells were injected intravenously (in tail vein), thereby imitating a clinical situation wherein cancer is diagnosed when tumor cells are already in blood stream. Tumor cells forms metastases in the lung, which can be detected and counted.
  • p62 polynucleotide was administered 1, 8, 15 days after tumor cell inoculation. As seen in FIG. 14 , treatment with p62 polynucleotide significantly reduced number and size of metastatic tumors, thereby demonstrating ant-metastatic effect.
  • mice Three groups of newborn male mice received subcutaneous injections of i) placebo or ii) and iii) 3 mg/kg of monosodium glutamate (MSG) daily for 10 days. Group 3 then received 5 weekly injections of 200 ug of p62-encoding plasmid intramuscularly. Body weight of animas in all 3 groups was measured and compared. MSG markedly increased body weight in both groups compared to the control group (p ⁇ 0.01 and p ⁇ 0.05). However, increase of body weight in the group receiving p62 plasmid was significantly less than in a group receiving MSG alone. Consequently, administration of the p62-encoding plasmid reduced obesity in the mouse model.
  • MSG monosodium glutamate
  • ZDF Diabetic Fatty
  • mice Two groups of mice were fed with MCD diet (10% fat, 40% sucrose, no choline, no methionine). Each group contained 15 mice. Serum level of alanine aminotransferase (ALT) was monitored starting at 10 weeks on the diet.
  • An experimental group received 5 weekly intramuscular injections of p62-encoding plasmid, 200 ug/injection.
  • the placebo group received MCD diet with no plasmid intervention.
  • Control group received normal diet and no plasmid through the entire research period. Both groups dieting on MCD have demonstrated elevated level of ALT compared to the control group (p ⁇ 0.01 for both groups). However, p62 treatment reduced ALT level compared to placebo group (p ⁇ 0.05). Consequently, administration of the p62 plasmid reduced the incidence of fat liver disease in the mouse model.
  • mice received drinking water containing 8% dextran sulfate sodium (DSS) for 2 weeks. Contemporarily, one group of mice received 5 weekly in injections of 200 ug of p62-encoding plasmid while the other group constituted the control (10 animals in each group). p62-treated animals demonstrated reduction in bloody stool and diarrhea. Also, the weight of each animal was monitored individually once a week. p62 inhibited weight loss (p ⁇ 0.05) although it did not entirely stop it. Consequently, administration of the p62 plasmid reduced symptoms related to Crohn's Disease in a mouse model.
  • DSS dextran sulfate sodium
  • mice were treated with 4 mg/animal LPS and 1 ug per animal caerulein for 10 weeks, twice a week, to induce pancreatitis.
  • 3 mice were used as a control group, and 3 received 5 weekly injections of p62 vaccine 200 ug/injection intramuscularly. After 15 weeks mice were sacrificed and subjected to histological analysis.
  • p62 plasmid treatment reduced the extent of observed chronic pancreatitis. Consequently, administration of the p62 plasmid reduced pancreatitis in a mouse model.
  • mice were subjected to three intraperitoneal injections of 1% ovalbumin solution. Injections were made with 14 days intervals. A week after the 3 d injection animals were exposed to a 1% ovalbumin aerosol for 30 min each day for 3 days to induce asthma-like disease. During this 5 weeks period, one group of mice received 5 weekly intramuscular injections of 200 ug of p62 plasmid, while another was kept as a control. Two days after the final exposure, airway hyperresponsiveness (AHR) was measured and compared in control and treatment group. AHR was induced either by cold air or by hyperventilation. AHR manifestation was significantly reduced in p62-treated group. Consequently, administration of the p62 plasmid reduced asthma symptoms in a mouse model.
  • AHR airway hyperresponsiveness
  • mice Two groups of mice (10 animals per group) were administered collagen II (CII) with complete Freund adjuvant to induce collagen-induced arthritis. Animals were 6 weeks of age to avoid spontaneous arthritis, which can be observed in older animals.
  • An experimental group also received 5 weekly injections of 200 of of p62-encoding plasmid. The control group manifested first signs on arthritis 30-33 weeks after the CII challenge.
  • the p62-treated group demonstrated collagen-induced arthritis 38-43 weeks after CII injections. Consequently, administration of the p62 plasmid delayed collagen-induced arthritis in a mouse model.
  • ApoE( ⁇ / ⁇ ) mice were maintained on a high-fat diet for 8 wks.
  • the control group was not vaccinated while the experimental group received 5 weekly injections of 200 ug of p62-encoding plasmid.
  • the plasmid treatment reduced the atherosclerotic plaque area and plaque neovessel density. It increased the plaque collagen and elastin contents, and reduced plasma angiotensin II levels and plaque macrophage infiltration and cathepsin S (CatS) protein.
  • CatS cathepsin S
  • p62 administration also decreased the levels of AT1R, gp91phox, TLR2, monocyte chemotactic protein-1 in the aortic roots. Consequently, administration of the p62 plasmid reduced atherosclerosis in a mouse model.
  • Ovariectomy induces neurodegenerative changes analogous to Huntington and Alzheimer's diseases in the mouse. Mice were subjected to sham operation or ovariectomy. Both operated groups were divided in 2 sub-groups: one subgroup received p62-plasmid vaccination, and another did not. A battery of behavioral tests were conducted as described below. In each test we have observed that p62 vaccination significantly alleviated the neurodegenerative effect of ovariectomy, which indicates it may have strong potential in treatment and/or prevention of Alzheimer's and Huntington's diseases.
  • Behavioral testing took place during four consecutive days. On the first day, animals were subjected to the whole battery of sensorimotor tests. On the second day, open field was performed. On the third day, animals were subjected to the Porsolt-test. Finally, the plus-maze test or light/dark test was performed on the last day.
  • the visual placing reflex was tested in order to evaluate the function of the visual system.
  • the mouse was suspended by the tail and lowered toward a solid black surface. Complete extension of the forelimbs was considered a positive response.
  • the mean response was rated in three trials (Baeza et al., 2010).
  • This reflex was evaluated during the previous test as the ability to perform complete extension of the hindlimbs when the animal was suspended by the tail. Such response was considered positive.
  • the mean response was rated in three trials (Baeza et al., 2010).
  • This method is used to evaluate the vitality loss in aging mice by testing their muscular vigor, motor coordination and traction in two training trials of 5 s and a test trial of 60 s (Miguel and Blasco, Ex p Gerontol. 1978; 13(6):389-96 and Baeza et al., 2010). Mice were suspended by their forelimbs in the middle of an elevated horizontal tightrope (40 cm height, 60 cm length and divided in segments of 10 cm). Muscular vigor was assessed as the percentage of mice falling off the rope and the latency to fall (in seconds). Motor coordination included the percentage of mice that walk at least 1 segment (criteria 1) and the percentage of mice that complete the test (criteria 2).
  • Traction was evaluated by analyzing the different parts of the body that mice used to remain suspended (forelimbs, hindlimbs and tail) and, subsequently, the percentages of mice displaying the maximum (forelimbs, hindlimbs and tail) and minimum (forelimbs only) traction capacities were assessed within each group.
  • This group includes different tests that study the depressive-like and anxiety-like behaviors in the animals: the FST, the open field, the light/dark and the EPM tests.
  • the forced-swimming test is the best recognized pharmacological model for assessing antidepressant-like activity in rodents (Porsolt et al., 1977a Arch Int Pharmacodyn Ther 229: 327-336, Porsolt et al., 1977b Nature 266: 730-732; Willner, 1990 Pharmacol Ther 45: 425-455; Al-Rahbi et al., Biomed Res Int. 2013; 2013:493643.doi: 10.1155/2013/493643).
  • mice were placed individually in a glass cylinder (20 cm in height, 14 cm in diameter) filled to a 15 cm depth with water (23 ⁇ 1° C.). At this water depth, the mice could touch the bottom of the jar with their tail, but they could not support themselves with their hind limbs.
  • Each mouse was given a 6 min swimming test, and the duration of immobility was noted during the final 4 min interval of the test, since the first 2 min were used to allow the animals to familiarize them-selves with the surroundings. All the swim-test sessions were recorded by a video camera positioned directly above the cylinder. Two experienced observers, who were blind to the treatment conditions, scored the videotapes. An immobility period was regarded as the time spent by the mouse floating in the water without struggling and while making only the very slight movements that are necessary to keep its head above the water. Following these swimming sessions, the mice were towel dried and returned to their housing. Each animal was tested only once.
  • Locomotor activity was measured by an open-field apparatus consisting of a square arena (43.2 cm ⁇ 43.2 cm) equipped with two lines of 16 photocells to measure horizontal and vertical activity. The arena was lit by one red light lamp (25 W), and a white noise generator in the room produced an ambient background noise of ⁇ 70 dB. All data were recorded on a personal computer (MED-PC Open-Field Activity Software) in an adjacent control room. Mice were placed in the centre of the apparatus and the test was carried out for 5 min. A number of conventional and ethological parameters (Choleris et al., 2001 Neurosci Biobehav Rev 25: 353-360; Perfumi and Mattioli, Phytother Res. 2007 January; 21(1):37-43) were collected during the session.
  • the horizontal activity (i.e. distance travelled, ambulation time, resting time) and the vertical activity (i.e. rearing) in the central and peripheral zone were recorded automatically.
  • the time spent in the central area, the ambulation time and vertical activity in this zone, and the latency to leave the starting central point and that to reach the periphery (“freezing behavior”) were measured as indicators of the emotional reactivity of the mouse (Baeza et al., 2010).
  • freezing behavior were measured as indicators of the emotional reactivity of the mouse (Baeza et al., 2010).
  • aging involves a decrease in the defecatory behaviour and an increase of urine incontinence. Therefore, the number of fecal boli and presence of urine were also considered in the different groups of age (in an attempt to study whether ovariectomy in mature animals caused these behaviors to be more similar to those observed in aged animals).
  • a relevant test system to detect anxiety-related behaviour in mice is the light/dark exploration test, which uses the aversion of rodents for brightly lit large spaces (Hascoet et al., 2001 Prog Neuropsychopharmacol Biol Psychiatry 25: 141-166; Bourin and Hascoet, 2003 Eur J Pharmacol 463: 55-65).
  • the light-dark apparatus consisted of an open-topped rectangular Plexiglas box (45 ⁇ 30 ⁇ 30 cm; I ⁇ b ⁇ h) that was divided into a small (18 ⁇ 30 cm) area and a large (27 ⁇ 30 cm) area with an opening door (7.5 ⁇ 7.5 cm) located in the center of the partition at floor level.
  • the small compartment was painted black and stayed dark, whereas the large compartment was painted white and was brightly illuminated with a 60 W (400 l ⁇ ) light source. Briefly, each animal was placed at the center of the illuminated compartment, facing one of the dark areas. The latency time for their first passage from the light compartment to the dark one, the number of entries into each compartment, the time spent in the illuminated area, and the number of times that the mouse reared on its hindpaws in the light space (rearing), were recorded for 5 min (Waif and Frye., 2010).
  • the elevated plus maze assesses anxiety-like behavior and consisted of black Plexiglas with two open arms (30 ⁇ 3.5 cm) and two enclosed arms of the same size (14 cm high walls). The four arms were connected by a central square (6 ⁇ 6 cm square) and were elevated approximately 74 cm from the ground. Briefly, mice were placed in the central square facing one of the closed arms and its behavior was scored for 5 min. The number of entries with all four paws within the arms and the time spent in the arms were scored separately for open and closed arms. A greater amount of raw time and a greater proportion of time (%) spent on the brightly-lit open arms of the elevated plus maze was considered an index of anxiety-like behavior (Kolosova et al., Aging (Albany N.Y.). 2013 June; 5(6):474-84; Walf and Frye., 2010).
  • a or “an” can refer to one of or a plurality of the elements it modifies (e.g., “a reagent” can mean one or more reagents) unless it is contextually clear either one of the elements or more than one of the elements is described.
  • the term “about” as used herein refers to a value within 10% of the underlying parameter (i.e., plus or minus 10%), and use of the term “about” at the beginning of a string of values modifies each of the values (i.e., “about 1, 2 and 3” refers to about 1, about 2 and about 3).
  • a weight of “about 100 grams” can include weights between 90 grams and 110 grams.

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2020028923A3 (en) * 2018-08-02 2020-03-12 Curelab Oncology, Inc. Compositions and methods relating to p62 for the treatment and prophylaxis of age-related macular degeneration

Families Citing this family (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
SG11201605316VA (en) 2013-12-29 2016-07-28 Curelab Oncology Inc Methods and compositions relating to p62/sqstm1 for the treatment and prevention of inflammation-associated diseases
US20220065875A1 (en) * 2019-02-08 2022-03-03 Ajou University Industry-Academic Cooperation Foundation Biomarker composition for diagnosis of degenerative brain disease
WO2021231652A1 (en) * 2020-05-14 2021-11-18 Curelab Oncology, Inc. Using the p62 plasmid to treat or reduce the severity of coronavirus infections
WO2022129180A1 (en) * 2020-12-15 2022-06-23 Centre National De La Recherche Scientifique Sqstm1 and its use in cancer therapy
WO2023156807A2 (en) * 2021-09-27 2023-08-24 Curelab Oncology, Inc. Prevention and treatment of diseases by modulating activity of mesenchymal stem cells with p62 (sqstm1)-encoding vector and pharmaceutical formulations containing p62 (sqstm1) proteins
CN117603982B (zh) * 2024-01-22 2024-04-19 中国医学科学院北京协和医院 肌萎缩侧索硬化症的SQSTM1的p.P374TfsTer18突变致病基因及其应用

Family Cites Families (30)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5364773A (en) 1991-03-07 1994-11-15 Virogenetics Corporation Genetically engineered vaccine strain
EP0681026A1 (en) 1987-03-02 1995-11-08 Whitehead Institute For Biomedical Research Recombinant mycobacterial vaccine
US5504005A (en) 1987-03-02 1996-04-02 Albert Einstein College Of Medicine Of Yeshiva University Recombinant mycobacterial vaccine
AU648505B2 (en) 1989-05-19 1994-04-28 Amgen, Inc. Metalloproteinase inhibitor
EP0517751A4 (en) 1990-02-26 1993-03-31 Commonwealth Scientific & Industrial Research Organisation ( C.S.I.R.O. ) Shuttle plasmid for escherichia coli and mycobacteria
GB9015888D0 (en) 1990-07-19 1990-09-05 Smithkline Biolog Vectors
WO1992015672A1 (en) 1991-03-07 1992-09-17 Virogenetics Corporation Genetically engineered vaccine strain
AU2185592A (en) 1991-06-06 1993-01-08 Med Immune, Inc. Induction of ctl responses to foreign antigens expressed in mycobacteria
US5962224A (en) * 1995-12-19 1999-10-05 Dana-Farber Cancer Institute Isolated DNA encoding p62 polypeptides and uses therefor
US5990091A (en) 1997-03-12 1999-11-23 Virogenetics Corporation Vectors having enhanced expression, and methods of making and uses thereof
US5977311A (en) * 1997-09-23 1999-11-02 Curagen Corporation 53BP2 complexes
NZ509966A (en) 1998-08-18 2003-07-25 Res Dev Foundation Recombinant vector comprising a cassette with TET-ON under the control of a heat shock promoter, a cassette with a cloning site for a therapeutic gene, a cassette comprising antisense TET-ON and cassette with a dominative negative TET-ON and its use for metastatic prostate, breast and ovarian cancer
CN1322730A (zh) * 2000-05-09 2001-11-21 上海博德基因开发有限公司 一种新的多肽——p62cdc6蛋白24和编码这种多肽的多核苷酸
AU2001280767A1 (en) 2000-07-31 2002-02-13 Active Motif Peptide-mediated delivery of molecules into cells
US20060035823A1 (en) * 2000-08-17 2006-02-16 Seth Lederman Isolated fragments of p62 nucleoporin and uses thereof
EP1372720A4 (en) * 2001-03-02 2006-07-26 Medimmune Inc PREVENTING OR TREATING INFLAMMATORY OR AUTOIMMUNE DISORDERS BY ADMINISTERING ANTAGONISTS OF INTEGRIN ALPHA V BETA 3
ATE389015T1 (de) 2001-07-30 2008-03-15 G R M O Groupe De Rech En Mala Paget knochenkrankheit
ATE421532T1 (de) 2002-05-14 2009-02-15 Jaekyoon Shin Die rolle von p62 bei altersbedingten erkrankungen
EP1578367A4 (en) * 2002-11-01 2012-05-02 Genentech Inc COMPOSITIONS AND METHODS FOR THE TREATMENT OF IMMUNE DISEASES
GB2443819B (en) 2003-11-17 2009-09-02 Univ Auburn Methods and compositions for inhibiting intracellular aggregate formation
WO2005073164A1 (en) * 2004-01-30 2005-08-11 Peplin Biolipids Pty Ltd Therapeutic and carrier molecules
GB0412301D0 (en) * 2004-06-02 2004-07-07 Diagenic As Product and method
WO2007044522A1 (en) * 2005-10-05 2007-04-19 Auburn University P62 as a risk determinant for metabolic syndrome
WO2011039734A2 (en) * 2009-10-02 2011-04-07 Enzo Medico Use of genes involved in anchorage independence for the optimization of diagnosis and treatment of human cancer
US8945847B2 (en) 2010-05-24 2015-02-03 Yissum Research Development Company Of The Hebrew University Of Jerusalem Ltd. Methods and kits for ascertaining biosafety of an agent
JP6193231B2 (ja) * 2011-08-08 2017-09-06 チュアラボ オンコロジー, インコーポレーテッドCurelab Oncology, Inc. がんの治療および予防のための、ｐ６２に関する方法および組成物
KR20150038915A (ko) 2013-10-01 2015-04-09 코닝정밀소재 주식회사 전기 이중층 캐패시터의 전극 소재용 그래핀 플레이크 제조방법, 이에 의해 제조된 그래핀 플레이크 및 이를 전극 소재로 포함하는 전기 이중층 캐패시터
KR101594168B1 (ko) 2013-10-02 2016-02-15 서울대학교산학협력단 p62의 ZZ 영역에 의해 매개되는 자가포식 조절 방법 및 그 용도
SG11201605316VA (en) 2013-12-29 2016-07-28 Curelab Oncology Inc Methods and compositions relating to p62/sqstm1 for the treatment and prevention of inflammation-associated diseases
WO2020028923A2 (en) * 2018-08-02 2020-02-06 Curelab Oncology, Inc. Compositions and methods relating to p62 for the treatment and prophylaxis of age-related macular degeneration

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2020028923A3 (en) * 2018-08-02 2020-03-12 Curelab Oncology, Inc. Compositions and methods relating to p62 for the treatment and prophylaxis of age-related macular degeneration

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