CN115814076A

CN115814076A - Application of anti-human VEGF antibody and chemical drug combination in preparation of drugs for treating ovarian cancer

Info

Publication number: CN115814076A
Application number: CN202211290780.9A
Authority: CN
Inventors: 薛黎婷; 杨文清; 冯晓飞; 张子婧; 任晋生
Original assignee: Jiangsu Simcere Pharmaceutical Co Ltd
Current assignee: Shandong Simcere Bio Pharmaceutical Co ltd
Priority date: 2021-07-01
Filing date: 2021-07-01
Publication date: 2023-03-21
Also published as: CN113244388A

Abstract

The invention relates to an application of a combination of an anti-human VEGF antibody and a chemical drug in preparation of a drug for treating ovarian cancer, in particular to a combination of the anti-human VEGF antibody and a chemotherapeutic drug, an ovarian cancer treatment method and an application of the combination in preparation of the drug for treating ovarian cancer. The invention is beneficial to increasing the selection of ovarian cancer patients, improving the treatment effect of ovarian cancer and benefiting more ovarian cancer patients.

Description

Application of anti-human VEGF antibody and chemical drug combination in preparation of drugs for treating ovarian cancer

Technical Field

The invention relates to the field of medicines, in particular to application of an anti-human VEGF antibody and a chemical drug in preparation of a drug for treating ovarian cancer.

Background

Ovarian cancer is a malignant tumor of epithelial origin, including epithelial ovarian cancer, fallopian tube cancer and primary peritoneal cancer, the second most common and most mortality gynecological malignancy. Ovarian cancer has no specific tumor markers, is difficult to diagnose early and is usually diagnosed at a later stage. Thus, the prognosis of ovarian cancer is poor, with a 5-year survival rate of only 37.6% in europe (Luigi Rossi et al, oncotarget,2016.8 p.12389-12405. Ovarian cancer is probably around 52100 new cases in china, and the rate of new cases increases by about 2% each year. Annual death cases are probably around 22500 (Chen W.et al, cancer statistics in China,2015.CA Cancer J Clin,2016.66 (2): p.115-32).

The accepted standard treatments for ovarian cancer are cytoreductive surgery and postoperative platinum-and paclitaxel-based chemotherapy. Despite the high response rate of first-line chemotherapy, up to 70-75% of patients still experience relapse (Luigi Rossi et al, oncotarget,2016.8 p.12389-12405. Ovarian cancer patients relapse within 6 months after treatment with platinum-based drugs is defined as platinum-based resistance. After the first relapse, approximately 25% of patients will develop platinum-based resistance, and eventually almost all relapsing patients will develop platinum-based resistance. In this case, the choice of effective treatment is very few and the prognosis is also poor. New therapeutic strategies are therefore needed to improve their clinical prognosis (Pujade-Lauraine, e., et al., J Clin Oncol,2014.32 (13): p.1302).

Vascular Endothelial Growth Factor (VEGF) is a multifunctional cytokine, is a currently known angiogenesis promoting factor with the strongest effect and the highest specificity, and participates in the generation and development of solid tumors by promoting tumor angiogenesis, increasing Vascular permeability, regulating host anti-tumor immune response and other mechanisms. Studies have shown that anti-human VEGF antibodies have the effect of inhibiting the growth of solid tumors.

The BD0801 antibody is a humanized rabbit anti-human VEGF monoclonal antibody, and the scheme of treating ovarian cancer by the BD0801 antibody is researched and discussed, so that the selection of patients is increased, the treatment effect of ovarian cancer is improved, and more ovarian cancer patients are benefited.

In view of the above, the present invention is particularly proposed.

Disclosure of Invention

The invention provides a combination of an anti-human VEGF antibody and a chemotherapeutic drug, a method for treating ovarian cancer by using the combination or application of the combination in preparing a drug for treating ovarian cancer.

In a first aspect, the present invention provides the use of an anti-human VEGF antibody in combination with a chemotherapeutic agent selected from a taxane compound or a camptothecin compound, in the manufacture of a medicament for the treatment of ovarian cancer, said antibody comprising a heavy chain variable region comprising HCDR1, HCDR2 and HCDR3 and a light chain variable region comprising LCDR1, LCDR2 and LCDR3, wherein,

the sequence of the HCDR1 is shown as SEQ ID NO. 5;

the sequence of the HCDR2 is shown as SEQ ID NO. 8;

the sequence of the HCDR3 is shown as SEQ ID NO. 11;

the sequence of the LCDR1 is shown as SEQ ID NO. 13;

the sequence of the LCDR2 is shown as SEQ ID NO. 15; and (c) and (d),

the sequence of the LCDR3 is shown as SEQ ID NO. 17.

In some embodiments, the heavy chain variable region has the sequence set forth in SEQ ID NO. 1 and the light chain variable region has the sequence set forth in SEQ ID NO. 3.

In some embodiments, the antibody further comprises a heavy chain constant region having the sequence set forth in SEQ ID NO. 2, and a light chain constant region having the sequence set forth in SEQ ID NO. 4.

In some embodiments, the ovarian cancer is epithelial ovarian cancer, fallopian tube cancer, or primary peritoneal cancer.

In some embodiments, the ovarian cancer is platinum-resistant recurrent ovarian cancer.

In some embodiments, the pathological type of ovarian cancer is serous adenocarcinoma, clear cell adenocarcinoma, or mixed epithelial carcinoma.

In some embodiments, the taxane compound is paclitaxel and the camptothecin compound is topotecan hydrochloride.

In some embodiments, the effective amount of the anti-human VEGF antibody administered is 0.5 to 2mg/kg.

In some embodiments, the effective amount of the anti-human VEGF antibody administered is 0.5mg/kg, 1mg/kg, 1.5mg/kg, or 2mg/kg.

In some embodiments, the dosage form of the anti-human VEGF antibody is a single dose dosage form, each dose containing an amount of the antibody effective for administration to a patient.

In some embodiments, the single dose dosage form contains 25 to 150mg of the anti-human VEGF antibody.

In some embodiments, the single dose dosage form contains 25mg, 50mg, 80mg, 100mg, or 150mg of the anti-human VEGF antibody.

In some embodiments, the effective amount of paclitaxel administered is 80mg/m2; the effective amount of topotecan hydrochloride administered is 4mg/m2.

In a second aspect, the present invention provides the use of an anti-human VEGF antibody in combination with a chemotherapeutic selected from a taxane compound or a camptothecin compound, for the manufacture of a medicament for the treatment of ovarian cancer, said antibody comprising a heavy chain variable region comprising HCDR1, HCDR2 and HCDR3 and a light chain variable region comprising LCDR1, LCDR2 and LCDR3, wherein:

the HCDR1 is selected from a sequence shown in any one of SEQ ID NO 5-7;

the HCDR2 is selected from a sequence shown in any one of SEQ ID NO 8-10;

the HCDR3 is selected from a sequence shown in any one of SEQ ID NO 11-12;

the LCDR1 is selected from a sequence shown in any one of SEQ ID NO 13-14;

the LCDR2 is selected from a sequence shown in any one of SEQ ID NO 15-16; and the combination of (a) and (b),

the LCDR3 is selected from a sequence shown in SEQ ID NO. 17.

In some embodiments, the heavy chain variable region has the sequence shown as SEQ ID No. 1, or a sequence that is at least 95%, 96%, 97%, 98%, or 99% identical to SEQ ID No. 1; the light chain variable region has the sequence shown as SEQ ID NO. 3, or a sequence with at least 95%, 96%, 97%, 98% or 99% identity compared to SEQ ID NO. 3.

In some embodiments, the antibody further comprises a heavy chain constant region having a sequence as set forth in SEQ ID No. 2, or a sequence at least 95%, 96%, 97%, 98%, or 99% identical to SEQ ID No. 2; the antibody also includes a light chain constant region having a sequence as set forth in SEQ ID NO. 4, or a sequence that is at least 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO. 4.

In some embodiments, the effective amount of the anti-human VEGF antibody administered is 0.5 to 2mg/kg, preferably 0.5mg/kg, 1mg/kg, 1.5mg/kg or 2mg/kg.

In some embodiments, the anti-human VEGF antibody is provided in a single dosage form, each containing an amount of antibody effective for administration to a patient, preferably, the single dosage form contains 25 to 150mg of the anti-VEGF antibody, more preferably 25mg, 50mg, 80mg, 100mg, or 150mg.

In some embodiments, the effective amount of paclitaxel administered is 80mg/m ² 。

In some embodiments, the effective amount of topotecan hydrochloride administered is 4mg/m ² 。

In a third aspect, the invention also provides a method of treating ovarian cancer, the method comprising administering to an ovarian cancer patient an effective amount of an anti-human VEGF antibody in combination with a chemotherapeutic selected from a taxane or a camptothecin, the antibody comprising a heavy chain variable region comprising HCDR1, HCDR2 and HCDR3 and a light chain variable region comprising LCDR1, LCDR2 and LCDR3, wherein:

the HCDR1 is selected from a sequence shown in any one of SEQ ID NO 5-7;

the HCDR2 is selected from a sequence shown in any one of SEQ ID NO 8-10;

the HCDR3 is selected from a sequence shown in any one of SEQ ID NO 11-12;

the LCDR1 is selected from a sequence shown in any one of SEQ ID NO 13-14;

the LCDR3 is selected from a sequence shown in SEQ ID NO. 17.

In some embodiments, the heavy chain variable region has the sequence set forth as SEQ ID No. 1, or a sequence having at least 95%, 96%, 97%, 98%, or 99% identity compared to SEQ ID No. 1; the light chain variable region has a sequence as shown in SEQ ID NO 3, or a sequence that is at least 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO 3.

In some embodiments, the anti-human VEGF antibody is administered 1 time every 2 weeks.

In some embodiments, the paclitaxel is administered 1 time per week.

In some embodiments, the topotecan hydrochloride is administered 3 times every 4 weeks, preferably 1 time per week for the first 3 weeks.

In some embodiments, the combination is administered for a period of 28 days, the anti-human VEGF antibody is administered 1 time each on

days

1 and 15, the paclitaxel is administered 1 time each on

days

1, 8, 15 and 22, and the topotecan hydrochloride is administered 1 time each on

days

1, 8 and 15.

In a fourth aspect, the invention also provides a combination of an anti-human VEGF antibody and a chemotherapeutic selected from a taxane compound or a camptothecin compound for the treatment of ovarian cancer, the antibody comprising a heavy chain variable region comprising HCDR1, HCDR2 and HCDR3 and a light chain variable region comprising LCDR1, LCDR2 and LCDR3, wherein:

the HCDR1 is selected from a sequence shown in any one of SEQ ID NO 5-7;

the HCDR2 is selected from a sequence shown in any one of SEQ ID NO 8-10;

the HCDR3 is selected from a sequence shown in any one of SEQ ID NO 11-12;

the LCDR1 is selected from a sequence shown in any one of SEQ ID NO 13-14;

the LCDR2 is selected from a sequence shown in any one of SEQ ID NO 15-16; and (c) and (d),

the LCDR3 is selected from a sequence shown in SEQ ID NO. 17.

In some specific embodiments, the heavy chain variable region has the sequence shown as SEQ ID No. 1, or a sequence that is at least 95%, 96%, 97%, 98% or 99% identical to SEQ ID No. 1; the light chain variable region has a sequence as shown in SEQ ID NO 3, or a sequence that is at least 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO 3.

In some specific embodiments, the antibody further comprises a heavy chain constant region having the sequence shown as SEQ ID No. 2, or a sequence that is at least 95%, 96%, 97%, 98%, or 99% identical to SEQ ID No. 2; the antibody also includes a light chain constant region having a sequence as set forth in SEQ ID NO. 4, or a sequence that is at least 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO. 4.

In some specific embodiments, the dosage form of the anti-human VEGF antibody is a single dosage form, each containing an amount of the antibody effective for administration to a patient, preferably, the single dosage form contains 25 to 150mg of the anti-VEGF antibody, more preferably 25mg, 50mg, 80mg, 100mg, or 150mg.

In some specific embodiments, the effective amount of paclitaxel administered is 80mg/m ² 。

In some specific embodiments, the effective amount of topotecan hydrochloride to be administered is 4mg/m ² 。

Definition and description of terms

Unless defined otherwise herein, scientific and technical terms related to the present invention shall have the meanings that are understood by those of ordinary skill in the art.

Furthermore, unless otherwise indicated herein, singular terms herein shall include the plural and plural terms shall include the singular. More specifically, as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the content clearly dictates otherwise.

The terms "comprising," "including," and "having," used interchangeably herein, are intended to be inclusive and mean that there may be additional elements other than the listed elements. It should also be understood that the use of "including", "comprising" and "having" in this context also provides a "consisting of … …" solution.

The term "and/or" as used herein includes the meanings of "and", "or" and "all or any other combination of elements linked by the term.

The term "antibody" is used herein in the broadest sense and refers to a polypeptide or combination of polypeptides that comprises sufficient sequence from an immunoglobulin heavy chain variable region and/or sufficient sequence from an immunoglobulin light chain variable region to be capable of specifically binding to an antigen. The "antibodies" herein encompass various forms and structures as long as they exhibit the desired antigenBinding activity, illustratively, includes intact antibodies as well as antibody fragments having binding activity to an antigen. The term "desired antigen binding activity" as used herein means, for example, that an antibody typically binds specifically to an antigen and to substantially the same antigen with high affinity, but does not bind to an unrelated antigen with high affinity. Affinity is usually reflected in an equilibrium dissociation constant (KD), where a lower KD indicates a higher affinity. Illustratively, high affinity generally means having about 10 ^-7 M or less, about 10 ^-8 M or less, about 1X 10 ^-9 M or less, about 1X 10 ^-10 M or less, 1X 10 ^-11 M or less or 1X 10 ^-12 M or lower KD. KD is calculated as follows: KD = KD/Ka, where KD represents the dissociation rate and Ka represents the association rate. The equilibrium dissociation constant KD can be measured using methods well known in the art, such as surface plasmon resonance (e.g., biacore) or equilibrium dialysis assay.

The term "antibody" herein includes a typical "four-chain antibody" belonging to an immunoglobulin composed of two Heavy Chains (HC) and two Light Chains (LC); heavy chain refers to a polypeptide chain consisting of, in the direction from N-terminus to C-terminus, a heavy chain variable region (VH), a heavy chain constant region CH1 domain, a Hinge Region (HR), a heavy chain constant region CH2 domain, a heavy chain constant region CH3 domain; and, when the full length antibody is of IgE isotype, optionally further comprising a heavy chain constant region CH4 domain; a light chain is a polypeptide chain consisting of a light chain variable region (VL) and a light chain constant region (CL) in the N-terminal to C-terminal direction; the heavy chains are connected with each other through disulfide bonds, and the heavy chains are connected with the light chains through disulfide bonds to form a Y-shaped structure. The constant regions of immunoglobulin heavy chains differ in their antigenicity due to their differing amino acid composition and arrangement. Accordingly, the term "immunoglobulin" is used herein to refer to five classes, or isotypes called immunoglobulins, namely IgM, igD, igG, igA and IgE, with the corresponding heavy chains being the μ, δ, γ, α and ε chains, respectively. The same class of Ig can be divided into different subclasses according to the differences of amino acid composition of the hinge region and the number and position of heavy chain disulfide bonds, for example, igG can be divided into IgG1, igG2, igG3 and IgG4, and IgA can be divided into IgA1 and IgA2. Light chains are classified as either kappa or lambda chains by differences in the constant regions. Each of the five classes of Ig may have either a kappa chain or a lambda chain.

The "antibody" herein may be derived from any animal, including but not limited to humans and non-human animals which may be selected from primates, mammals, rodents and vertebrates, such as camelids, llamas, ostriches, alpacas, sheep, rabbits, mice, rats or chondroiidaes (e.g. shark).

The term "humanized antibody" herein refers to a non-human antibody that has been genetically engineered to have an amino acid sequence modified to increase homology to the sequence of a human antibody. Generally, all or a portion of the CDR regions of a humanized antibody are derived from a non-human antibody (donor antibody), and all or a portion of the non-CDR regions (e.g., variable region FR and/or constant regions) are derived from a human immunoglobulin (acceptor antibody). Humanized antibodies typically retain or partially retain the desired properties of the donor antibody, including, but not limited to, antigen specificity, affinity, reactivity, the ability to increase the activity of immune cells, the ability to enhance an immune response, and the like.

The term "variable region" herein refers to the region of an antibody heavy chain or light chain involved in binding an antibody to an antigen, the "heavy chain variable region" being used interchangeably with "VH", "HCVR" and the "light chain variable region" being used interchangeably with "VL", "LCVR". The variable domains of the heavy and light chains of natural antibodies (VH and VL, respectively) generally have similar structures, each domain comprising four conserved Framework Regions (FR) and three hypervariable regions (HVRs). See, e.g., kindt et al, kuby Immunology,6th ed, w.h.freeman and co., p.91 (2007). A single VH or VL domain may be sufficient to confer antigen binding specificity.

The terms "complementarity determining region" and "CDR" are used interchangeably herein and generally refer to the hypervariable region (HVR) of a heavy chain variable region (VH), which may be abbreviated as HCDR, or a light chain variable region (VL), which is also referred to as a complementarity determining region because it is sterically complementary to an epitope precisely. The terms "framework region" or "FR region" are used interchangeably herein to refer to those amino acid residues in an antibody heavy chain variable region or light chain variable region other than the CDRs. Generally, a typical antibody variable region consists of 4 FR regions and 3 CDR regions in the following order: FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4.

For further description of the CDRs, reference is made to Kabat et al, j.biol.chem., 252; kabat et al, united states department of health and public service, "Sequences of proteins of immunological interest" (1991); chothia et al, J.mol.biol.196:901-917 (1987); al-Lazikani b. Et Al, j.mol.biol., 273; macCallum et al, J.mol.biol.262:732-745 (1996); abhinandan and Martin, mol.immunol., 45; lefranc m.p. et al, dev.comp.immunol., 27; and honeyger and pluckthun, j.mol.biol.,309 (2001). The "CDRs" herein may be labeled and defined by means known in the art, including but not limited to Kabat numbering system, chothia numbering system, or IMGT numbering system, using tool sites including but not limited to AbRSA sites (http:// cao. Lab share. Cn/AbRSA/cds. Php), abYsis sites (www.abysis.org/analysis/sequence _ input/key _ indication. Cgi), and IMGT sites (http:// www.imgt.org/3Dstructure-DB/cgi/DomainGapAlign. Cgi # results).

Illustratively, SEQ ID NO:1 or 3 is as shown in the following table: wherein Kabat and Chothia use the abYsis website and IMGT uses the IMGT website.

TABLE 1 amino acid sequence listing of heavy and light chain CDR regions of anti-human VEGF antibody

The term "heavy chain constant region" herein refers to the carboxy-terminal portion of an antibody heavy chain that is not directly involved in binding of the antibody to an antigen, but exhibits effector functions, such as interaction with an Fc receptor, which has a more conserved amino acid sequence relative to the variable domain of the antibody. The "heavy chain constant region" comprises at least: a CH1 domain, a hinge region, a CH2 domain, a CH3 domain, or a variant or fragment thereof. The "heavy chain constant region" includes a "full-length heavy chain constant region" having a structure substantially similar to that of a natural antibody constant region, and a "heavy chain constant region fragment" including only a portion of the full-length heavy chain constant region. Illustratively, a typical "full-length antibody heavy chain constant region" consists of a CH1 domain-hinge region-CH 2 domain-CH 3 domain; when the antibody is IgE, it further comprises a CH4 domain; when the antibody is a heavy chain antibody, it does not include a CH1 domain. Illustratively, a typical "heavy chain constant region fragment" may be selected from the CH1, fc or CH3 domains.

The term "light chain constant region" herein refers to the carboxy-terminal portion of an antibody light chain that is not directly involved in binding of the antibody to an antigen, which light chain constant region may be selected from a constant kappa domain or a constant lambda domain.

The term "Fc" herein refers to the carboxy-terminal portion of an antibody that is papain-hydrolyzed from an intact antibody, typically comprising the CH3 and CH2 domains of the antibody. Fc regions include, for example, native sequence Fc regions, recombinant Fc regions, and variant Fc regions. Although the boundaries of the Fc region of an immunoglobulin heavy chain may vary slightly, the Fc region of a human IgG heavy chain is generally defined as extending from the amino acid residue at position Cys226 or from Pro230 to its carboxy terminus. The C-terminal lysine of the Fc region (residue 447 according to the Kabat numbering system) may be removed, for example, during production or purification of the antibody, or by recombinant engineering of the nucleic acid encoding the heavy chain of the antibody, and thus, the Fc region may or may not include Lys447.

The term "identity" herein can be calculated by: to determine the percent "identity" of two amino acid sequences or two nucleic acid sequences, the sequences are aligned for optimal comparison purposes (e.g., gaps can be introduced in one or both of the first and second amino acid sequences or nucleic acid sequences for optimal alignment or non-homologous sequences can be discarded for comparison purposes). The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, then the molecules are identical at that position.

The percent identity between two sequences varies with the identity position shared by the sequences, taking into account the number of gaps that need to be introduced and the length of each gap for optimal alignment of the two sequences.

The term "drug" herein refers to a substance used for the prevention, treatment and diagnosis of a disease, including "pharmaceutical compositions" or "drug combinations", illustratively, the drug also includes a cell therapeutic agent, such as a therapeutic CAR-T cell or CAR-NK cell. By "pharmaceutical composition" is meant a formulation that is present in a form that allows the biological activity of the active ingredient contained therein to be effective and that does not contain additional ingredients that have unacceptable toxicity to the subject to which the pharmaceutical composition is administered. The term "combination" or "pharmaceutical combination" as used herein refers to an unfixed combination, wherein the active agent and the at least one further active agent may be administered separately, either simultaneously or within time intervals, in particular where these time intervals allow the combination partners to show a cooperative (e.g. synergistic) effect. The term "non-fixed combination" means that the active ingredients (e.g., one active agent and at least one additional active agent) are both administered to a patient as separate entities either simultaneously or sequentially without specific time constraints, wherein such administration provides therapeutically effective levels of both compounds in the patient.

The term "treatment" as used herein refers to surgery or drug treatment (therapeutic or therapeutic treatment) for the purpose of preventing, slowing (reducing) the progression of an undesired physiological change or pathology, such as cancer or a tumor, in a subject being treated. Beneficial or desired clinical results include, but are not limited to, alleviation of symptoms, diminishment of extent of disease, stabilized (i.e., not worsening) state of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, and remission (whether partial or complete), whether detectable or undetectable. Subjects in need of treatment include subjects already suffering from a condition or disease as well as subjects susceptible to a condition or disease or subjects for whom prevention of a condition or disease is intended. When terms such as slow, moderate, weak, moderate, etc. are referred to, their meanings also include elimination, disappearance, nonoccurrence, etc.

The term "patient" or "subject" herein refers to an organism that is receiving treatment for a particular disease or disorder as described herein. Examples of "patients" or "subjects" include mammals, such as humans, primates (e.g., monkeys), or non-primate mammals, that are being treated for a disease or disorder.

The term "effective amount" herein refers to an amount of a therapeutic agent that is effective to prevent or ameliorate a disease condition or the progression of the disease when administered alone or in combination with another therapeutic agent to a cell, tissue, or subject. An "effective amount" also refers to an amount of a compound or cell having a therapeutic effect (e.g., CAR-T cells, CAR-NK cells) sufficient to alleviate symptoms, e.g., to treat, cure, prevent, or alleviate a related medical condition, or to treat, cure, prevent, or alleviate an increased rate of such a condition. When the active ingredient is administered alone to an individual, a therapeutically effective dose refers to the ingredient alone.

The term "cancer" herein refers to or describes a physiological condition in mammals that is typically characterized by unregulated cell growth. Included in this definition are benign and malignant cancers. The term "tumor" or "neoplasm" herein refers to all neoplastic (neoplastic) cell growth and proliferation, whether malignant or benign, and all pre-cancerous (pre-cancerous) and cancerous cells and tissues. The terms "cancer" and "tumor" are not mutually exclusive when referred to herein.

Drawings

FIG. 1. Anti-tumor effects of BD0801, avastin and Paclitaxel in the human ovarian cancer OVCAR-8 nude mouse transplant tumor model.

FIG. 2. Anti-tumor effects of BD0801, avastin and Paclitaxel in combination in human ovarian cancer OVCAR-8 nude mouse transplant tumor model.

FIG. 3 anti-tumor effects of BD0801, avastin and Paclitaxel single drug and combination in human ovarian cancer SK-OV-3 nude mouse transplantation tumor model.

Figure 4.Bd0801 in combination with PTX group subjects Objective Remission Rate (ORR).

FIG. 5 is a graph of the Objective Remission Rate (ORR) of BD0801 combined PTX/TOPO group ovarian cancer patients versus BD0801 AUC for a hierarchical analysis.

Figure 6, median Progression Free Survival (PFS) profile in subjects overall.

Figure 7 bd0801 in combination with PTX exposure as a function of percentage reduction of the sum of the longest diameters (SLD) of the tumors.

Detailed Description

The invention will be further described with reference to specific embodiments, and the advantages and features of the invention will become apparent as the description proceeds. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products available commercially.

The examples are exemplary only and do not limit the scope of the invention in any way. It will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention, and that such changes and modifications may be made without departing from the spirit and scope of the invention.

The BD0801 antibody used in the following examples was produced and supplied by Shandong Mitsui Bio-pharmaceuticals, inc. and stored at 2-8 ℃. The sequence information of the BD0801 antibody is shown below.

Heavy chain variable region (SEQ ID NO: 1):

EVQLVESGGGLVKPGGSLRLSCAASGFSFSNNDVMCWVRQAPGKGLEWIGCIMTTDVVTEYANWAKSRFTVSRDSAKNSVYLQMNSLRAEDTAVYFCARDSVGSPLMSFDLWGPGTLVTVSS；

heavy chain constant region (SEQ ID NO: 2):

ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK；

light chain variable region (SEQ ID NO: 3):

DIQMTQSPSSLSASVGDRVTINCQASQSIYNNNELSWYQQKPGKPPKLLIYRASTLASGVPSRFSGSGSGTDFTLTISSLQPEDVATYYCGGYKSYSNDGNGFGGGTKVEIK；

light chain constant region (SEQ ID NO: 4):

RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC。

example 1 evaluation of the therapeutic Effect of BD0801 on the model of human ovarian carcinoma nude mouse transplanted tumor

1.1 test materials and tumor evaluation methods (unless otherwise specified, this example uses the test materials and tumor evaluation methods described in 1.1)

The tested medicine is as follows:

BD0801 is manufactured and provided by tsukusan shou bio-pharmaceuticals, inc;

bevacizumab (Avastin) was purchased from Roche (Roche);

paclitaxel (Paclitaxel) was purchased from Beijing synergia.

The preparation method of the medicine comprises the following steps: the drugs were prepared and diluted with sterile normal saline (purchased from Jiangsu Huai' an double crane, pharmaceutical Co., ltd.).

OVCAR-8 cells: national Institute of Health (NIH).

SK-OV-3 cells: shanghai national institute of Life sciences (SIBS), chinese academy of sciences.

Reagent: RPMI1640: gibco, cat number: c22400500BT;

fetal bovine serum: gibco, cat number: 10099-141C;

PBS: hyclone, cat # s: SH30256.01;

matrigel: corning, cargo number: 354234.

the tumor evaluation method comprises the following steps:

tumor volume V =0.5 × a × b ² Wherein a is the major diameter of the tumor and b is the minor diameter of the tumor;

relative tumor volume RTV = V _t /V ₀ In which V is ₀ Tumor volume of the animal at the time of grouping; v _t Is the tumor volume of the animal after treatment.

Relative tumor proliferation rate T/C (%) = T _RTV /C _RTV X 100% where T _RTV Mean RTV for treatment groups; c _RTV The negative control group mean RTV.

Tumor growth inhibition ratio TGI (%) = (1-T/C) × 100%.

1.2 BD0801 curative effect evaluation on human ovarian cancer OVCAR-8 nude mouse transplantation tumor model-single medicine

Experimental animals: BALB/c Nude mouse, female, 6-8 weeks, weight 18.5-25.5g, purchased from Jiangsu Jiejiegaokang Biotech, inc., production license number: SCXK (su) 2018-0008, animal certification No.: 201911201. a breeding environment: SPF grade.

OVCAR-8 cells were cultured in RPMI1640 medium containing 10% fetal bovine serum. Collecting cells in exponential growth phase, 1X 10 ⁷ OVCAR-8 cells were resuspended in 1: PBS 1 in matrigel (0.1 ml/mouse) was inoculated subcutaneously into experimental mice. The tumor is grown to the average volume of 158.62mm ³ Group administration, the day of group administration being defined as day 0 (D0), mice were randomized into 8 groups (n = 8), each group of mice being administered saline (vehicle), 0.8mg/kg,2.5mg/kg or 7.5mg/kg of the test drug BD0801,0.8mg/kg,2.5mg/kg or 7.5mg/kg of the control drug Avastin, or 15mg/kg of the positive control drug Paclitaxel, respectively, via the tail vein. BD0801 and Avastin, administered 2 times per week for about 6 weeks, paclitaxel administered 1 time every four days for 4 times. Tumor volume was measured 2 times per week, mice were weighed, data was recorded, and tumor size change and mouse body weight change were counted over about 6 weeks. Therefore, in the human ovarian cancer OVCAR-8 nude mouse transplantation tumor model, the BD0801 can obviously inhibit the tumor growth at 2.5mg/kg and 7.5mg/kg, and the tumor inhibition effect is better than Avastin.

On day 38 after administration, the tumor volumes of the 0.8mg/kg BD0801 group were not statistically different from those of the Vehicle control group, and the tumor volumes of the 2.5mg/kg and 7.5mg/kg BD0801 groups showed significant tumor suppression effects compared with those of the control group. The tumor volumes of the 0.8mg/kg,2.5mg/kg and 7.5mg/kg Avastin groups were not statistically different compared to the Vehicle control group. The anti-tumor effect of BD0801 is better than that of Avastin under the same dose, and the two doses of BD0801 are statistically different from each other under the doses of 0.8mg/kg and 2.5 mg/kg. No weight loss or other significant toxic response was seen in the animals for the different doses of BD0801 and Avastin. 15mg/kg of Paclitaxel also produced significant tumor suppression, but the drug had a significant toxic response, with 2 of 8 mice dying. See table 2 and figure 1 for details. The result shows that in the human ovarian cancer OVCAR-8 nude mouse transplantation tumor model, BD0801 can obviously inhibit the tumor growth at 2.5mg/kg and 7.5mg/kg, and the tumor inhibition effect is superior to Avastin.

TABLE 2 tumor inhibiting Effect of test substances on OVCAR-8 xenograft tumor model

Note: a. data are shown as mean ± Standard Deviation (SD).

b.P values the relative tumor volumes were analyzed using two-way ANOVA, showing P values for each group compared to vehicle controls.

The 2 tumor-bearing mice of the Paclixel 15mg/kg group were found dead on

days

5 and 12, respectively, and none of these mice had been found for tumor volume on day 38 and corresponding calculations.

1.3 Evaluation of curative effect of BD0801 on human ovarian cancer OVCAR-8 nude mouse transplantation tumor-combined chemotherapy

Experimental animals: BALB/c Nude mouse, female, 6-8 weeks, body weight 16.2-21.5g, purchased from Jiangsu Jiejiegaokang Biotech limited, production license number: SCXK (su) 2018-0008, animal certification No.: 202002168. a breeding environment: SPF grade.

OVCAR-8 cells were cultured in RPMI1640 medium containing 10% fetal bovine serum. Collecting cells in exponential growth phase, 1X 10 ⁷ OVCAR-8 cells were resuspended in 1:1 (0.1 ml/mouse) in a matrix gel to inoculate the cells subcutaneously in the experimental mice. The tumor is grown to the average volume of 159.42mm ³ Group administration, the day of group administration being defined as day 0 (D0), mice were randomly divided into 6 groups (n = 8), and each group of mice was administered physiological saline (Vehicle) via the tail vein, 2.5mg/kg of the test drug BD0801,2.5mg/kg of the control drug Avastin,10mg/kg of the positive control drug Paclitaxel,2.5mg/kg BD0801 in combination with 10mg/kg Pacliaxel, or 2.5mg/kg Avastin in combination with 10mg/kg Pacliaxel. BD0801 and Avastin, administered 2 times per week for 4 weeks, paclitaxel administered 1 time every four days for 4 times. Tumor volume was measured 2 times per week, mice were weighed, data was recorded, and tumor size change and mouse body weight change were counted over about 4 weeks.

On the 24 th day after administration, 2.5mg/kg BD0801 or 2.5mg/kg Avastin can remarkably inhibit the growth of tumors, and the tumor inhibition effect is equivalent. The 2.5mg/kg Avastin and 10mg/kg Paclitaxel combination group did not show significantly enhanced anti-tumor effects relative to the Avastin monotherapy group; the combination of 2.5mg/kg BD0801 and 10mg/kg Paclixel showed significantly enhanced tumor suppression compared with the two single drug groups, and the tumor suppression effect of the combined group tended to be superior to that of the Avastin and Paclixel combined group. The tumor-bearing mice were tolerated by treatment with BD0801 alone at 2.5mg/kg, avastin at 2.5mg/kg, paclixel at 10mg/kg, and BD0801 at 2.5mg/kg in combination with Paclixel at 10mg/kg, and Avastin at 2.5mg/kg in combination with Paclixel at 10 mg/kg. See table 3 and figure 2 for details. The result shows that in a human ovarian cancer OVCAR-8 nude mouse transplantation tumor model, 2.5mg/kg BD0801 single drug treatment has a remarkable tumor inhibition effect, and when the drug is combined with 10mg/kg Paclitaxel, the drug shows a remarkably enhanced tumor inhibition effect relative to two single drug groups, and the trend of the antitumor effect is remarkably superior to that of the combined group of the same dose of Avastin and Paclitaxel.

TABLE 3 tumor-inhibiting Effect of test substances on OVCAR-8 xenograft tumor model

Note: a. data are shown as mean ± SD.

1.4 BD0801 curative effect evaluation-combined chemotherapy of human ovarian cancer SK-OV-3 nude mouse transplantation tumor

Experimental animals: BALB/c Nude mice, female, 6-8 weeks, and 18.0-23.5g in weight, purchased from Jiangsu Jiejiaokang Biotech, inc., production license number: SCXK (su) 2018-0008, animal certification number: 202002168. a breeding environment: SPF grade.

SK-OV-3 cells were cultured in McCoy's 5a medium (McCoy's 5a, cat # 12330-031) containing 10% fetal bovine serum (Isuase, cat # FND 500). Collecting cells in exponential growth phase, 1X 10 ⁷ SK-OV-3 cells were resuspended in 1:1 (0.1 ml/mouse) in a matrix gel to inoculate the cells subcutaneously in the experimental mice. The tumor is grown to the average volume of 135.48mm ³ Group administration, the day of group administration being defined as day 0 (D0), the mice were randomized into 8 groups (n = 8), each group of mice being administered saline (vehicle) via the tail vein, 2.5mg/kg or 7.5mg/kg of the test drug BD0801,2.5mg/kg or 7.5mg/kg of the control drug Avastin,10mg/kg of the positive control drug Paclitaxel,10mg/kg of Paclitaxel in combination with 2.5mg/kg BD0801, or 10mg/kg of Paclitaxel in combination with 2.5mg/kg Avastin, respectively. BD0801 and Avastin, administered 2 times per week for 4 weeks, paclitaxel administered 1 time every four days for 4 times. Tumor volume was measured 2 times per week, mice were weighed, data was recorded, and tumor size change and mouse weight change were counted over about 4 weeks.

On day 24 after the administration, all the administration groups showed significant tumor growth inhibitory activity compared to the control group. The test drug BD0801 showed very good dose dependence, whereas the control drug Avastin showed a slightly weaker dose dependence. The tumor inhibition effect of BD0801 is equivalent to that of Avastin at the dose of 2.5mg/kg, and the tumor inhibition effect of BD0801 is obviously superior to that of Avastin at the dose of 7.5 mg/kg. The 2.5mg/kg Avastin and 10mg/kg Paclitaxel combination group did not show significantly enhanced anti-tumor effects relative to the Avastin monotherapy group; while the combination of 2.5mg/kg BD0801 and 10mg/kg Paclixel showed significantly enhanced tumor suppression relative to both single drug groups. The tumor-bearing mice were tolerated by treatment with BD0801 at 2.5mg/kg and 7.5mg/kg, respectively, with Avastin administered alone at 2.5mg/kg and 7.5mg/kg, respectively, and with Paclitaxel at a dose of 10mg/kg, and with BD0801 at 2.5mg/kg in combination with Paclitaxel at a dose of 10mg/kg, and with Avastin at 2.5mg/kg in combination with Paclitaxel at a dose of 10 mg/kg. See table 4 and fig. 3 for details. The result shows that in a human ovarian cancer SK-OV-3 nude mouse transplantation tumor model, BD0801 and Avastin treatment have obvious tumor inhibition effect on an SK-OV-3 mouse ovarian cancer model, BD0801 shows a very good dose response relationship, and the Avastin dose response relationship is slightly weak. BD0801 showed significantly better tumor suppression at high doses of 7.5mg/kg than Avastin at the same dose, and significantly enhanced tumor suppression at 2.5mg/kg in combination with Paclitaxel relative to both single drug groups.

TABLE 4 tumor-inhibiting Effect of test substances on SK-OV-3 xenograft tumor model

Note: a. data are shown as mean ± SD.

Example 2 clinical trials of BD0801 combination chemotherapy in ovarian cancer patients

Object of the study

BD0801 monoclonal antibody was evaluated for safety and tolerability, pharmacokinetic profile and anti-tumor efficacy in combination with chemotherapy for platinum-resistant relapsed epithelial ovarian cancer, fallopian tube cancer and primary peritoneal carcinoma patients.

Design of research

Adopts a multicenter, open and BD0801 monoclonal antibody dosage increasing design and combines chemotherapy to treat patients with platinum drug-resistant recurrent epithelial ovarian cancer, salpingemphraxis and primary peritoneal carcinoma.

The test is divided into two parts: the first phase is a dose escalation phase, wherein the BD0801 monoclonal antibody dose escalation is designed to be 0.5mg/kg, 1mg/kg, 1.5mg/kg and 2mg/kg and is administered once every 2 weeks, and the dose escalation is based on dose-limiting toxicities (DLTs) occurring in 2 administration cycles (two BD0801 monoclonal antibody combination chemotherapies, generally for 28 days, and the maximum dose does not exceed 56 days) during which blood samples are collected for determination of drug concentration. The chemotherapeutic drug selected for combination was paclitaxel (once weekly) or topotecan hydrochloride (3 times every 4 weeks). After treatment is complete, efficacy is assessed and subjects with clinical benefit can continue to take the drug at the original dose and frequency until disease progression or intolerance of adverse reactions occurs or informed consent/loss is withdrawn or a new anti-tumor treatment is initiated.

The second stage is an extension into the group stage to further investigate the safety and effectiveness of the product. If no DLT was observed in the first 3 subjects at the end of the first 2 treatment cycles (week 4) or only 1 subject observed DLT at the end of the first 2 treatment cycles (week 4) in the first 6 subjects per dose group, then group entry was allowed to continue to 8-12. In addition, a particular dose group can be selected to extend up to 40 subjects, discussed collectively by investigators and sponsors. The administration method and blood sampling method are the same as those of the subjects in the dose group.

Dose escalation and termination criteria:

no dose escalation was performed in the subjects. Dose escalation between groups will be guided by the occurrence of dose-limiting toxicity (DLT) over 2 dosing cycles (two administrations of BD0801 mab, typically 28 days, up to 56 days). The traditional mode of 3+3 is adopted for dose increasing, each dose group firstly enters 1 subject, and after 1 BD0801 monoclonal antibody administration (d 14) is completed, if no DLT occurs, 2 subjects enter the group. At least 3 subjects completed treatment and evaluation on

cycles

1 and 2 per dose group and were available for DLT assessment, and were escalated to the next dose in the absence of DLT; 1 patient presented with DLT as in the previous 3, and 3 additional patients were enrolled into the dose group, and if ≧ 2/6 subjects presented with DLT, dose escalation was discontinued; as in the previous 3 patients with 2 patients who developed DLT, dose escalation was stopped.

MTD (maximum tolerated dose) is defined as the highest dose at which less than one-third of the subjects develop DLT within a specified time. The determination of MTD is based on the occurrence of DLT within 2 administration cycles (BD 0801 monoclonal antibody is administered twice, generally for 28 days, and the maximum duration is not more than 56 days).

Population of subjects

Patients with platinum-based drug-resistant recurrent epithelial ovarian cancer, fallopian tube cancer, and primary peritoneal cancer.

And (3) inclusion standard:

1. age: is more than or equal to 18 years old;

2. histologically or cytologically confirmed platinum-based drug-resistant epithelial ovarian cancer, fallopian tube cancer or primary peritoneal cancer of the types: non-specific adenocarcinoma (NOS), clear cell adenocarcinoma, endometrial adenocarcinoma, malignant brenner's tumor, mixed epithelial carcinoma, mucinous adenocarcinoma, serous adenocarcinoma, transitional cell carcinoma, and undifferentiated carcinoma;

3. at least one measurable tumor lesion (according to RECIST 1.1);

ECOG PS score 0-1;

5. subjects must have proper organ function and meet all the following laboratory test results before enrollment:

a) Bone marrow stores were essentially normal (no transfusions and blood products, no correction with G-CSF and other hematopoietic stimulators within 14 days prior to screening): ANC ≥ 1.5 × 10 ⁹ /L，HB≥90g/L，PLT≥100×10 ⁹ /L；

b) Liver function is essentially normal: TBIL is less than or equal to 1.5 × ULN, AST is less than or equal to 2.5 × ULN, and alkaline phosphatase is less than or equal to 2.5 × ULN;

c) The renal function is basically normal: the serum creatinine is less than or equal to ULN or the GFR (glomerular clearance) is more than or equal to 60mL/min, and the serum creatinine is calculated according to a Cockroft-Gault formula: male GFR = (140-age) BW/(Scr × (72), female GFR = male 0.85;

d) The blood coagulation function is basically normal: INR < 1.5 (if the patient receives a stable dose of warfarin to manage venous thrombosis, INR should range from 2-3), APTT < 1.2 × ULN;

6. disease progression occurs within 6 months after completion of at least 4 cycles of platinum-containing therapy;

7. the estimated survival time is more than or equal to 12 weeks;

8. at least 4 weeks from the last chemotherapy, and if the anti-tumor biological product is received, at least 4 half-lives of the elution phase are required;

9. the toxic and side effects caused by the prior treatment need to be recovered to the grade less than or equal to 1 (NCI CTC 4.03);

10. subjects had to give informed consent to the study prior to the trial and voluntarily signed a written informed consent form;

11. the subjects were able to communicate well with the investigator and completed the study in accordance with the study prescription.

Exclusion criteria:

1. has previously received >2 anti-tumor treatment regimens;

2. patients were refractory to platinum, defined as progression of disease during previous platinum-containing therapy;

3. ovarian tumors with low malignant potential, such as borderline tumors;

4. patients with aggressive malignancies (except for non-melanoma skin cancers) or previous treatments for malignancies are contraindicated by current treatment regimens;

5. the patient received any pelvic or abdominal radiotherapy;

6. patients are associated with severe, non-healing wounds, ulcers or fractures;

7. patients with a history of intestinal obstruction (including occlusive disease) had a history of peritoneal fistula, gastrointestinal perforation, and peritoneal abscess. In the screening period, pelvic cavity examination and rectum sigmoid colon invasion are carried out, or a patient with intestinal invasion is found by CT examination, or intestinal obstruction with clinical symptoms is carried out;

severe infections require intravenous infusion of antibiotics or hospitalization;

9. the disease history of thromboembolism or hemorrhagic diseases is within 6 months before the treatment;

10. CNS diseases unrelated to tumors, symptomatic brain metastases;

11. cardiovascular diseases with clinical significance:

a) Uncontrollable hypertension (systolic pressure is more than or equal to 150mmHg and/or diastolic pressure is more than or equal to 90 mmHg);

b) A history of myocardial infarction or unstable angina within 6 months prior to group entry;

c) Congestive heart failure or new york heart disease society (NYHA) grade ii heart failure;

d) Cardiac arrhythmias in severe need of drug therapy; asymptomatic atrial fibrillation without a controllable heart rate;

12. left ventricular ejection fraction below the lower limit of normal;

13. neuropathy more than or equal to CTCAE 2 level exists in the past (only limited to a taxol + BD0801 monoclonal antibody group);

14. known to be severely allergic to the therapeutic drugs or adjuvants used in the regimen;

15. pregnant or lactating women;

16. patients with proteinuria (proteinuria >1+ found in screening phase exam, or proteinuria of 1+, not returning to normal within 24 hours);

17. those who have been treated with anti-VEGF protein drugs such as bevacizumab and BD0801 monoclonal antibody;

18. the patient undergoes major surgery or is expected to undergo surgical treatment:

a) Large surgical operations or obvious trauma were performed within 28 days before group entry;

b) Major surgical procedures are contemplated during the study, including but not limited to abdominal surgery (open or laparoscopic surgery) prior to disease progression;

c) Open biopsy was performed within 7 days before group entry;

19. treatment with other study drugs is currently or within 30 days prior to group entry.

Test drug

The BD0801 antibody was produced and supplied by Shandong Mitsui Bio-pharmaceuticals, inc., and stored at 2-8 ℃.

Paclitaxel injection: the manufacturer: beijing collaborates with the pharmaceutical factory.

Topotecan hydrochloride for injection: the manufacturer: jiangsu Osekang pharmaceutical Co., ltd.

Mode of administration

BD0801 mab dosing regimen: the specified dose (0.5 mg/kg, 1mg/kg, 1.5mg/kg, 2 mg/kg) was administered once every 2 weeks (14 days).

Paclitaxel (paclitaxel): 80mg/m ² And the administration is carried out once on the 1 st, 8 th, 15 th and 22 th days in the 28-day administration cycle. Applied as per the instructions.

Topotecan hydrochloride: 4mg/m ² Intravenous infusion was given 1 time each on

days

1, 8, and 15 of the 28-day dosing cycle. Applied as per the instructions.

The administration sequence is as follows: the administration was carried out according to the respective administration cycles. If the BD0801 monoclonal antibody and the chemotherapeutic agent are administered on the same day, the BD0801 monoclonal antibody is infused first.

Test grouping

BD0801 mab + paclitaxel: BD0801 mAb is administered every 2 weeks (14 days) as a dosing cycle, and paclitaxel is administered once every 1, 8, 15, 22 days in a 28-day dosing cycle, if the two are administered on the same day, BD0801 mAb is infused first.

BD0801 mab + topotecan hydrochloride group: BD0801 mAb is administered every 2 weeks (14 days) for a single administration cycle, and topotecan hydrochloride is administered 1 time each on

days

1, 8, and 15 in a 28-day administration cycle, such as on the same day, and BD0801 mAb is first infused.

Treatment continues until disease progression or intolerance of adverse effects occurs or other end-of-treatment criteria are met.

Research index and evaluation standard thereof

The safety index is as follows: DLT, adverse events, clinical safety laboratory examinations, electrocardiogram, vital signs, echocardiogram, etc.

The therapeutic end point is as follows: objective Remission Rate (ORR), disease Control Rate (DCR), progression Free Survival (PFS), and Overall Survival (OS).

Pharmacokinetic indexes are as follows: cmax, tmax, AUC _(TAU) 、Accumulation Ratio(AR)、C _trough 。

Statistics and analysis

Analyzing the data set:

total analysis set (FAS): all subjects enrolled and received study medication and had at least one tumor assessment after the baseline period.

Security data set (SS): all subjects were enrolled and received study medication. DLT assessment analysis set: the DLT assessment set included subjects who completed at least one treatment with BD0801 mab during the dose escalation phase and completed the DLT assessment observation period (e.g., 28 days from first dose) or withdrawal from DLT during the observation period.

PK concentration set (PKCS): all subjects were enrolled and received study medication with at least 1 effective blood level data during the trial.

PK parameters analysis set (PKPS): all subjects enrolled and received study medication with at least 1 effective PK parameter during the trial. Those not included in PKPS also include: a) Those who have entered a serious violation scenario, affect the PK parameter outcome, or cannot estimate the parameters; b) Concomitant medication occurred during the course of the trial and had an effect on PK parameters.

And (3) safety analysis:

analysis was performed according to the SS/DLT assessment analysis set. DLT and MTD were determined. Descriptive statistics are used to analyze adverse events, laboratory examination indices, vital signs, electrocardiograms, and the like. Adverse events, adverse reactions, major adverse events, severe adverse events, adverse events leading to discontinuation of medication, and the like were summarized according to dose groups.

Pharmacokinetic analysis:

performing descriptive statistical summary on the blood concentration of the BD0801 monoclonal antibody according to the dose level and the planning time; PK parameters were analyzed by dose level descriptive statistics. The drug-time curve and the mean drug-time curve for each subject were plotted against the blood concentration-time data for each subject measured in the experiment.

Immunogenicity analysis:

the frequency and rate of immunogenicity positive reactions were summarized and the corresponding titer values were given as tabulated by subject and study date. And the influence of immunogenicity on the pharmacokinetics and safety of the BD0801 monoclonal antibody is discussed.

And (3) analyzing the curative effect:

objective Remission Rate (ORR), disease Control Rate (DCR) and its 95% confidence interval were calculated from the total analysis set (FAS). For progression-free survival (PFS) and Overall Survival (OS), a Kaplan-Meier method was used to estimate survival curves and plot survival curves. Descriptive statistics were performed for tumor size changes.

Objective Remission Rate (ORR): ORR and its 95% confidence interval (Clopper-Pearson exact method) were calculated.

ORR＝(CR+PR)/(CR+PR+SD+PD+NE)×100％

The confirmed (confirmed) and unconfirmed (unconfirmed) CR and PR results were analyzed simultaneously.

Disease Control Rate (DCR): the DCR and its 95% confidence interval were calculated (Clopper-Pearson exact method).

DCR＝(CR+PR+SD)/(CR+PR+SD+PD+NE)×100％

Both confirmed (confirmed) and unconfirmed (unconfirmed) results were analyzed.

Progression Free Survival (PFS): the time between the start of study medication until the first recorded PD or death date, depending on which occurred first; if no disease progression is observed, the truncation date should be the date of the last tumor measurement before receiving a new antineoplastic agent. The PFS was summarized using the Kaplan-Meier test plot, listing the deletion rate, quartile and 95% CI (Brookmeyer-Crowley method), respectively, and estimating the time-to-time progression-free survival rate as 95% CI (loglog conversion).

PFS (month) = (first disease progression or death date or last tumor measurement date-first dosing date + 1)/30.4375

Overall Survival (OS): time from study medication initiation to death date (for any reason); if no mortality is observed, the last survival date is known as the truncation date. The OS was summarized using the Kaplan-Meier test plot, listing the deletion rates, quartile and 95% CI (Brookmeyer-Crowley method), and estimating the survival rates at different times and 95% CI (loglog conversion), respectively.

OS (month) = (death date or last follow-up date-first dosing date + 1)/30.4375

Tumor size change: descriptive statistics were performed on the measurable tumor Sum of Longest Diameter (SLD) versus baseline changes. And the maximum percentage of each patient's SLD reduction from the screening period was waterfall. In addition, the change in SLD from baseline at each tumor assessment after each patient dose will also be listed.

Results

1. Population of subjects

A total of 29 subjects participated in the trial and received at least 1 study drug treatment. Of 29 subjects, the mean course was about 28.19 months, most of which were epithelial ovarian cancer (25, 86.2%), primary peritoneal and fallopian tube cancer (2 each (6.9%), pathological diagnosis was mainly serous adenocarcinoma (25, 86.2%), clear cell adenocarcinoma 2 (6.9%), mixed epithelial cancer 1 (3.4%), and others 1 (3.4%). See table 5 for details.

TABLE 5 history of tumor

The 29 subjects all had target lesions, the maximum sum of which was 195.54mm, the minimum 12mm, and the average 71.342mm.

The subjects in the group had received chemotherapy, had received no radiation therapy, and 1 subject (3.4%) had received PARP inhibitor treatment and 8 subjects (27.6%) had received additional treatment.

The majority of 29 subjects (26, 89.7%) scored 0 for ECOG and 3 subjects (10.3%) scored 1 for inclusion.

29 subjects had a significant surgical history.

The most common concomitant diseases are hypertension (5 cases, 17.2%) and diabetes (5 cases, 17.2%), and the most common concomitant medication is systemic antibacterial (2 cases, 6.9%).

In 29 subjects, 22 (75.9%) were treated with the subsequent chemotherapeutic anti-tumor treatment after the end of the study treatment, 2 in the 0.5mg/kg dose group, 11 in the 1mg/kg dose group, and 9 in the 1.5mg/kg dose group.

2. Safety feature

No DLT event occurred in this study, MTD did not reach, and the recommended dose was 1.5mg/kg Q2W.

By the end of the study, 29 subjects (100%) had adverse events, and the frequency of adverse events was not different between each dose group and the different chemotherapy regimen groups, as hematological toxicity was more common. Grade 3 and above adverse events with incidence > 5% were decreased neutrophil count (51.72%), decreased leukocyte count (41.38%), decreased platelet count (17.24%), anemia (17.24%), increased gamma-glutamyltransferase (10.34%), emesis (10.34%), hypertension (10.34%), hypertriglyceridemia (6.90%).

A total of 27 subjects (93.10%) experienced adverse events associated with BD0801 mab, with similar rates of drug-related adverse events in the three dose groups, and the highest incidence of drug-related adverse events, classified according to PT, was a decrease in white blood cell count (24.14%).

A total of 1 death-causing SAE occurred, with intracranial bleeding occurring in the 1.5mg/kg dose group (in combination with topotecan hydrochloride), and the investigator believes that this SAE may not be associated with the test drug, nor with the combination of chemotherapeutic drugs. In a total of 11 subjects, after 15 treatments, SAE, the highest incidence of SAE was a decrease in neutrophil count (6.9%) according to PT classification.

A total of 26 subjects (89.66%) experienced adverse events that resulted in a decrement or suspension of BD0801 mab, with the highest incidence of decreased neutrophil count (44.83%); in total, 3 subjects (10.34%) had adverse events leading to permanent withdrawal of BD0801 mAb after administration, 2 in the 1.5mg/kg dose group, decreased platelet count and proteinuria, and 1 in the 1.0mg/kg dose group, respectively, as abdominal pain.

Adverse events of particular interest for this study included: gastrointestinal perforation or fistulae, hypertension, proteinuria, embolism and thrombotic events, bleeding of various types, etc., occur as detailed in table 6. According to SOC classification, the highest incidence of AESI was renal and urological disorders (11/37.93%), followed by various examinations (9/31.03%), vascular and lymphatic disorders (7/24.14%), and respiratory, thoracic and mediastinal disorders (3/10.34%). According to the PT classification, the highest incidence of AESI was proteinuria (11 cases/37.93%), followed by detection of urine protein (7 cases/24.14%), hypertension (6 cases/20.69%).

TABLE 6 analysis of adverse events with specific attention after drug administration in a subsystem (SS)

Grade 3 and above adverse event analysis

In total, 25 subjects (86.21%) had grade 3 or greater severity after drug administration, with the 0.5mg/kg, 1.0mg/kg and 1.5mg/kg groups being 100%, 91.67% and 78.57%, respectively. Grade 3 and above adverse events with incidence rates greater than or equal to 5% included decreased neutrophil count (51.72%), decreased leukocyte count (41.38%), decreased platelet count (17.24%), anemia (17.24%), increased gamma-glutamyltransferase (10.34%), hypertension (10.34%), decreased neutrophil percentage (6.90%), emesis (10.34%), hypertriglyceridemia (6.90%). See table 7 for details.

TABLE 7 adverse event subsystem analysis (SS) grade no less than 3 after dosing

Grade 3 or more adverse event incidence associated with BD0801 mAb was 37.93%, with a 1.5mg/kg group incidence of 57.14%, slightly higher than 1.0mg/kg group (25.0%), and paclitaxel subgroups slightly higher than the topotecan hydrochloride subgroup (45.45% vs. v33.33%). Grade 3 adverse events associated with BD0801 with an incidence of 5% or greater included decreased leukocyte count (10.34%), decreased neutrophil count (10.34%), hypertension (10.34%), increased gamma-glutamyltransferase (6.90%), and decreased platelet count (6.90%). See table 8 for details. Adverse events associated with BD0801 monoclonal antibody and not associated with chemotherapy at grade 3 or more after no drug administration.

TABLE 8 grade 3 sub-System analysis (SS) of adverse events associated with BD0801 monoclonal antibody after drug administration

3. Pharmacokinetics/pharmacodynamics

The pharmacokinetic parameters of BD0801 monoclonal antibody at different doses after administration are detailed in tables 9-1, 9-2 and 9-3. Wherein, after a single administration, the dose range of T is 0.5-1.5mg/kg _1/2 56.41-93.4h, CL 0.35-0.38 ml/h/kg, V28.52E48.63ml/kg; major exposure parameter (C) _max 、AUC _0-t 、AUC _0-inf ) Increases with increasing dose and exhibits a better linear relationship. After multiple administrations, T is in the dose range of 0.5-1.5mg/kg _1/2 63.5 to 101.97h, 0.35 to 0.49ml/h/kg of CL and 32.27 to 61.84ml/kg of V; major exposure parameter (C) _max 、AUC _0-t 、AUC _0-inf ) Increases with increasing dose and exhibits a better linear relationship. The AUC under the steady state is similar to the AUC value of the first administration, and the BD0801 monoclonal antibody has no obvious accumulation after multiple administrations.

Remarking: * T is _max Described by mean (range); AR, accumulation coefficient, AR = AUC _0-t (Steady State)/AUC _0-t (first time).

TABLE 9-2 BD0801 post-administration pharmacokinetic parameters (Mean SD)

TABLE 9-3 BD0801 Mb pharmacokinetic parameters (Mean SD)

Remarking: * T is a unit of _max Using mean (range) description; AR, accumulation coefficient, AR = AUC _0-t (Steady State)/AUC _0-t (first time).

Immunogenicity

BD0801 mab total 29 subjects included immunogenicity analysis in a platinum-resistant recurrent epithelial ovarian cancer, fallopian tube cancer and primary peritoneal carcinoma patient study. Compared with the baseline level, after the BD0801 monoclonal antibody is administered, the sera of 9 patients detect positive ADA (anti-drug antibody), and the positive detection rate of ADA individuals is 31.0% (9/29).

4. Effectiveness of

(1) Analysis (confirmation) of overall best curative effect of tumor focus

The overall Objective Remission Rate (ORR) was 31%, the clinical benefit was found in a total of 20 subjects, the clinical benefit rate (DCR) was 69.0%, and the overall optimal efficacy was CR in 1 (3.4%) of the subjects, which was a subset of paclitaxel in the 1.0mg/kg dose group. The best efficacy was 8 (27.6%) subjects with PR, with 1 in the 0.5mg/kg dose group (33.3%), 3 in the 1mg/kg dose group (25.0%), and 4 in the 1.5mg/kg dose group (28.6%). The optimal therapeutic effect was SD in a total of 11 subjects (37.9%), 6 in the 1mg/kg dose group (50.0%), and 5 in the 1.5mg/kg dose group (35.7%). A detailed overall best-fit analysis of each dose group is shown in table 10.

Remission occurred in 7 of 11 subjects with BD0801 mab in combination with paclitaxel subgroup, with an overall objective remission rate of 63.6%,8 clinical benefits, and a DCR of 72.7%. ORRs of the 0.5mg/kg, 1.0mg/kg, and 1.5mg/kg dose groups were 50.0%, 66.7%, and 66.7%, respectively.

In 18 subjects in the group of BD0801 monoclonal antibody and topotecan hydrochloride, 2 subjects showed remission, the overall objective remission rate was 11.1%,12 subjects gained clinical benefit, and the DCR was 66.7%

. ORRs of the 0.5mg/kg dose group, 1.0mg/kg dose group, and 1.5mg/kg dose group were 0, 22.2%, and 0, respectively.

TABLE 10 Total best efficacy (confirmation) analysis of tumor lesions (FAS)

Note: optimal efficacy refers to the best efficacy recorded from the start of treatment to before disease progression/recurrence.

Objective Remission Rate (ORR) = N1/N, N1= CR + PR;

clinical benefit rate (DCR) = N2/N, N2= CR + PR + SD;

conditions that could not be classified as CR, PR, SD and PD were classified as NE (not evaluable).

And (3) confirming that: CR or PR patients must be reviewed 4 weeks after initial assessment as CR or PR to confirm efficacy.

(2) Comprehensive analysis of overall best treatment of tumor lesions (not confirmed)

The overall objective remission rate was 37.9% for the unidentified subjects, the clinical benefit rate was 69.0%, and the overall best efficacy was CR for 1 (3.4%) of the subjects, which was the 1.0mg/kg dose group. The overall best effect was found in 10 subjects (34.5%) with PR, with 1 in the 0.5mg/kg dose group (33.3%), 3 in the 1.0mg/kg dose group (25.0%), and 6 in the 1.5mg/kg dose group (42.9%). A detailed overall best-fit analysis of each dose group is shown in table 11.

Objective remission with ORR of 72.7% was observed in 8 out of 11 subjects in the BD0801 mab and paclitaxel treatment subgroup, with ORRs in the 0.5mg/kg, 1.0mg/kg, and 1.5mg/kg dose groups being 50.0%, 66.7%, and 83.3%, respectively. In total, 8 subjects had clinical benefit, with DCR of 72.7%, DCR of 50.0%, 66.7% and 83.3% in the 0.5mg/k, 1.0mg/kg and 1.5mg/kg dose groups, respectively.

BD0801 mab in combination with topotecan hydrochloride treatment subgroup of 18 subjects showed objective remission with an ORR of 16.7%, with 0.5mg/kg, 1.0mg/kg and 1.5mg/kg dose groups having ORRs of 0, 22.2% and 12.5%, respectively. A total of 12 subjects had clinical benefit with DCR of 66.7%,0.5mg/kg, 1.0mg/kg and 1.5mg/kg dose groups with DCR of 0, 88.9%, 50.0%, respectively.

TABLE 11 Overall best efficacy (not confirmed) of tumor lesions general analysis (FAS)

Objective Remission Rate (ORR) = N1/N, N1= CR + PR;

clinical benefit rate (DCR) = N2/N, N2= CR + PR + SD;

the disease conditions that could not be classified as CR, PR, SD and PD were classified as NE (not evaluable).

Not confirmed: unconfirmed CR or PR (calculated without consideration of the next tumor assessment confirmation).

(3) Disease progression free survival (monthly) analysis

Of the 29 subjects, 27 (93.1%) subjects developed PFS events and 2 subjects in the 1.5mg/kg dose group were deleted and were a subset of paclitaxel. Median PFS was 5.4 months, with median PFS for the 0.5mg/kg, 1.0mg/kg and 1.5mg/kg groups being 1.9 months, 5.9 months and 4.6 months, respectively. The survival rate of no progress is 65.5% in 3 months, 40.3% in 6 months and 4.0% in 1 year. Detailed progression-free survival analysis is shown in tables 12-13.

PFS events occurred in 9 (81.8%) of 11 subjects of the BD0801 mAb and paclitaxel subgroup with median PFS of 5.3 months, 5.2 months, 5.3 months and 5.7 months in the 0.5mg/kg, 1.0mg/kg and 1.5mg/kg groups, respectively. The survival rate of no progress is 72.7% in 3 months, 43.6% in 6 months and 14.5% in 1 year.

A subset of 18 subjects (100%) with BD0801 mab combined with topotecan hydrochloride had PFS events with a median PFS of 5.5 months, 1.0mg/kg and 1.5mg/kg groups for 1.0 month, 6.1 months and 4.0 months, respectively. The survival rate of no progress at 3 months is 61.1%, the survival rate of no progress at 6 months is 38.9%, and the survival rate of no progress at 1 year is 0.

TABLE 12 disease progression free survival analysis (FAS)

Note: 95% CI was calculated using the Brookmeyer-Crowley method.

TABLE 13 progression free survival (%) at different times (month) (FAS)

Note: the 95% CI was calculated using a loglog transformation.

(4) Total Life time (month) analysis

By 20 days 10 months after 2020, median follow-up time was 12.8 months, 15 (51.7%) OS events were followed, 14 were missed (survived 10, missed 4), median OS was 14.9 months, 0.5mg/kg dose group mOS did not reach, 1.0mg/kg dose group, 1.5mg/kg dose group mean OS was 13.5 months, 18.9 months, respectively. The total survival rate in 6 months is 86.2%, the total survival rate in 1 year is 71.0%, and the total survival rate in 2 years is 31.2%. The detailed overall survival and survival analysis are shown in tables 14-15.

While 4 (36.4%) OS events were co-visited in 11 subjects treated with BD0801 mAb in combination with paclitaxel, 2 subjects in the 0.5mg/kg dose group did not reach OS, and median OS was 14.2 months and 18.9 months in the 1.0mg/kg dose group and 1.5mg/kg dose group, respectively. The 6-month survival rates of the 0.5mg/kg dose group, the 1.0mg/kg dose group, and the 1.5mg/kg dose group were 100%, and 83.3%, respectively, and the 1-year overall survival rates were 100%, 66.7%, and 83.3%, respectively.

BD0801 mab in combination with topotecan hydrochloride treatment group of 18 subjects out of 11 (61.1%) had OS events with a median OS of 12.8 months. The median OS was 3.5 months, 12.8 months and 15.1 months in the 0.5mg/kg, 1.0mg/kg and 1.5mg/kg dose groups, respectively. The total survival rate in 6 months is 83.3%, the total survival rate in 1 year is 65.7%, and the total survival rate in 2 years is 24.6%.

TABLE 14 Total Life time (month) analysis (FAS)

TABLE 15 Total survival (%) at different times (months) (%) (FAS)

(4) Analysis of relationship between exposure and curative effect

In 11 patients with BD0801 monoclonal antibody in combination with Paclitaxel (PTX) chemotherapy regimen, ORR was 7/11 (63.6%), the 11 PTX groups of subjects were further analyzed by dose-grouping, and dose-effect relationship analysis was performed between Objective Remission Rate (ORR) for optimal overall efficacy and the dose administered to the subjects as shown in FIG. 4.

In the BD0801 mab with Paclitaxel (PTX) or topotecan hydrochloride chemotherapy regimen, subjects were ranked from low to high AUC, layered 3 total. The dose-effect relationship between the ORR of the best overall curative effect of the subjects and the drug exposure (AUC) was analyzed in a layered manner, and the ORR and the BD0801 monoclonal antibody exposure showed a positive correlation, compared with the BD0801 monoclonal antibody combined with topotecan hydrochloride (TOPO), which showed no significant correlation between the ORR and the exposure, as shown in FIG. 5.

There was no significant correlation between Progression Free Survival (PFS) and dose at different doses in the population as a whole, as detailed in figure 6.

The relationship between exposure of BD0801 mab in combination with Paclitaxel (PTX) and the percentage of tumor longest diameter Sum (SLD) shrinkage, as shown in fig. 7, shows that exposure of BD0801 mab is positively correlated with tumor shrinkage.

Claims

1. Use of an anti-human VEGF antibody in combination with a chemotherapeutic agent, wherein the chemotherapeutic agent is selected from paclitaxel or topotecan hydrochloride, for the manufacture of a medicament for the treatment of platinum-resistant recurrent ovarian cancer, the antibody comprising a heavy chain variable region comprising HCDR1, HCDR2 and HCDR3 and a light chain variable region comprising LCDR1, LCDR2 and LCDR3, wherein,

the sequence of the HCDR1 is shown as SEQ ID NO. 5;

the sequence of the HCDR2 is shown as SEQ ID NO. 8;

the sequence of the HCDR3 is shown as SEQ ID NO. 11;

the sequence of the LCDR1 is shown as SEQ ID NO. 13;

the sequence of the LCDR2 is shown as SEQ ID NO. 15; and the combination of (a) and (b),

the sequence of the LCDR3 is shown as SEQ ID NO. 17;

the ovarian cancer is epithelial ovarian cancer, fallopian tube cancer or primary peritoneal cancer, and the pathological type of the ovarian cancer is serous adenocarcinoma, clear cell adenocarcinoma or mixed epithelial cancer.

2. The use of claim 1, wherein the heavy chain variable region has the sequence shown in SEQ ID NO. 1 and the light chain variable region has the sequence shown in SEQ ID NO. 3.

3. The use of claim 2, wherein the antibody further comprises a heavy chain constant region having the sequence set forth in SEQ ID NO. 2, and a light chain constant region having the sequence set forth in SEQ ID NO. 4.

4. The use according to any one of claims 1 to 3, wherein the effective amount of anti-human VEGF antibody administered is between 0.5 and 2mg/kg.

5. The use of claim 5, wherein said effective amount of anti-human VEGF antibody administered is 0.5mg/kg, 1mg/kg, 1.5mg/kg or 2mg/kg.

6. The use of claim 6, wherein said anti-human VEGF antibody is administered in a single dose, each dose containing an amount of antibody effective for administration to a patient.

7. The use of claim 7, wherein said single dosage form contains 25-150 mg of said anti-human VEGF antibody.

8. The use of claim 8, wherein said single dose dosage form contains 25mg, 50mg, 80mg, 100mg or 150mg of said anti-human VEGF antibody.

9. The use of claim 9, wherein the effective amount of paclitaxel administered is 80mg/m ² (ii) a The effective application amount of the topotecan hydrochloride is 4mg/m ² 。