Chimeric polypeptide assembly and methods of making and using the same

Abstract

The present invention relates to bispecific chimeric polypeptide assembly compositions comprising bulking moieties linked to binding domains by cleavable release segments that, when cleaved are capable of concurrently binding effector T cells with targeted tumor or cancer cells and effecting cytolysis of the tumor cells or cancer cells. The invention also provides compositions and methods of making and using the cleavable chimeric polypeptide assembly compositions.

1 - 61 . (canceled) 
     
     
         62 . A pharmaceutical composition comprising a chimeric polypeptide assembly comprising a first portion, a second portion, and a third portion wherein:
 a. said first portion comprises
 i. a first binding domain with binding specificity to a target cell marker; and 
 ii. a second binding domain with binding specificity to an effector cell antigen; 
   b. said second portion comprises a peptidyl release segment (RS) capable of being cleaved by one or more mammalian proteases; and   c. said third portion comprises a bulking moiety;   
       wherein said bulking moiety is capable of being released from said first portion by action of said mammalian protease on said second portion. 
     
     
         63 . The pharmaceutical composition of  claim 62 , wherein the composition is formulated for intradermal, subcutaneous, intravenous, intra-arterial, intraabdominal, intraperitoneal, intrathecal, or intramuscular administration. 
     
     
         64 . The pharmaceutical composition of  claim 62 , wherein the composition is in a liquid form. 
     
     
         65 . The pharmaceutical composition of  claim 64 , wherein the composition is in a pre-filled syringe for a single injection. 
     
     
         66 . The pharmaceutical composition of  claim 62 , wherein the composition is formulated as a lyophilized powder to be reconstituted prior to administration. 
     
     
         67 - 68 . (canceled) 
     
     
         69 . A method of treating a disease in a subject, comprising administering to the subject in need thereof a therapeutically effective dose of the chimeric polypeptide assembly or a pharmaceutical composition comprising a chimeric polypeptide assembly comprising a first portion, a second portion, and a third portion wherein:
 a. said first portion comprises
 i. a first binding domain with binding specificity to a target cell marker; and 
 ii. a second binding domain with binding specificity to an effector cell antigen; 
   b. said second portion comprises a peptidyl release segment (RS) capable of being cleaved by one or more mammalian proteases; and   c. said third portion comprises a bulking moiety;   
       wherein said bulking moiety is capable of being released from said first portion by action of said mammalian protease on said second portion. 
     
     
         70 . The method of  claim 69 , wherein the disease is selected from the group consisting of carcinomas, Hodgkin's lymphoma, non-Hodgkin's lymphoma, B cell lymphoma, T-cell lymphoma, follicular lymphoma, mantle cell lymphoma, blastoma, breast cancer, colon cancer, prostate cancer, head and neck cancer, any form of skin cancer, melanoma, genito-urinary tract cancer, ovarian cancer, ovarian cancer with malignant ascites, peritoneal carcinomatosis, uterine serous carcinoma, endometrial cancer, cervical cancer, colorectal cancer, an epithelia intraperitoneal malignancy with malignant ascites, uterine cancer, mesothelioma in the peritoneum kidney cancers, lung cancer, small-cell lung cancer, non-small cell lung cancer, gastric cancer, esophageal cancer, stomach cancer, small intestine cancer, liver cancer, hepatocarcinoma, hepatoblastoma, liposarcoma, pancreatic cancer, gall bladder cancer, cancers of the bile duct, salivary gland carcinoma, thyroid cancer, epithelial cancer, adenocarcinoma, sarcomas of any origin, primary hematologic malignancies including acute or chronic lymphocytic leukemias, acute or chronic myelogenous leukemias, myeloproliferative neoplastic disorders, or myelodysplastic disorders, myasthenia gravis, Morbus Basedow, Hashimoto thyroiditis, or Goodpasture syndrome. 
     
     
         71 . The method of  claim 69 , wherein the pharmaceutical composition is administered to the subject as one or more therapeutically effective doses administered twice weekly, once a week, every two weeks, every three weeks, or monthly. 
     
     
         72 . The method of  claim 69 , wherein the pharmaceutical composition is administered to the subject as one or more doses over a period of at least two weeks, or at least one month, or at least two months, or at least three months, or at least four months, or at least five months, or at least six months. 
     
     
         73 . The method of  claim 69 , wherein the dose is administered intradermally, subcutaneously, intravenously, intra-arterially, intra-abdominally, intraperitoneally, intrathecally, or intramuscularly. 
     
     
         74 . The method of  claim 69 , wherein the dose is administered as a bolus dose or by infusion of 5 minutes to 96 hours as tolerated for maximal safety and efficacy. 
     
     
         75 . The method of  claim 69 , wherein the dose is selected from the group consisting of at least about 0.005 mg/kg, at least about 0.01 mg/kg, at least about 0.02 mg/kg, at least about 0.04 mg/kg, at least about 0.08 mg/kg, at least about 0.1 mg/kg, at least about 0.12 mg/kg, at least about 0.14 mg/kg, at least about 0.16 mg/kg, at least about 0.18 mg/kg, at least about 0.20 mg/kg, at least about 0.22 mg/kg, at least about mg/kg, at least about 0.26 mg/kg, at least about 0.27 mg/kg, at least about 0.28 mg/kg, at least 0.3 mg/kg, at least 0.4. mg/kg, at least about 0.5 mg/kg, at least about 0.6 mg/kg, at least about 0.7 mg/kg, at least about 0.8 mg/kg, at least about 0.9 mg/kg, at least about 1.0 mg/kg, at least about 1.5 mg/kg, or at least about 2.0 mg/kg. 
     
     
         76 . The method of  claim 69 , wherein an initial dose is selected from the group consisting of at least about 0.005 mg/kg, at least about 0.01 mg/kg, at least about 0.02 mg/kg, at least about 0.04 mg/kg, at least about 0.08 mg/kg, at least about 0.1 mg/kg, and a subsequent dose is selected from the group consisting of at least about 0.1 mg/kg, at least about 0.12 mg/kg, at least about 0.14 mg/kg, at least about 0.16 mg/kg, at least about 0.18 mg/kg, at least about 0.20 mg/kg, at least about 0.22 mg/kg, at least about 0.24 mg/kg, at least about 0.26 mg/kg, at least about 0.27 mg/kg, at least about 0.28 mg/kg, at least 0.3 mg/kg, at least 0.4. mg/kg, at least about 0.5 mg/kg, at least about 0.6 mg/kg, at least about 0.7 mg/kg, at least about 0.8 mg/kg, at least about 0.9 mg/kg, at least about 1.0 mg/kg, at least about 1.5 mg/kg, or at least about 2.0 mg/kg. 
     
     
         77 . The method of  claim 69 , wherein the administration to the subject results in a plasma concentration of the chimeric polypeptide assembly of at least about 0.1 ng/mL to at least about 2 ng/mL or more in the subject for at least about 3 days, at least about 7 days, at least about 10 days, at least about 14 days, or at least about 21 days. 
     
     
         78 . The method of  claim 69 , wherein the subject is selected from the group consisting of mouse, rat, monkey, and human. 
     
     
         79 - 84 . (canceled) 
     
     
         85 . A kit comprising the pharmaceutical composition of  claim 62 , a container and a label or package insert on or associated with the container. 
     
     
         86 - 99 . (canceled) 
     
     
         100 . An isolated nucleic acid comprising a polynucleotide sequence selected from
 (a) a polynucleotide encoding a chimeric polypeptide assembly comprising a first portion, a second portion, and a third portion wherein:
 a. said first portion comprises
 i. a first binding domain with binding specificity to a target cell marker; and 
 ii. a second binding domain with binding specificity to an effector cell antigen; 
 
 b. said second portion comprises a peptidyl release segment (RS) capable of being cleaved by one or more mammalian proteases; and 
 c. said third portion comprises a bulking moiety; 
   
       wherein said bulking moiety is capable of being released from said first portion by action of said mammalian protease on said second portion; or
 (b) the complement of the polynucleotide of (a). 
 
     
     
         101 . (canceled) 
     
     
         102 . An expression vector comprising the polynucleotide sequence of  claim 100  and a recombinant regulatory sequence operably linked to the polynucleotide sequence. 
     
     
         103 . An isolated host cell, comprising the expression vector of  claim 102 . 
     
     
         104 - 138 . (canceled)