Dnase fusion polypeptides and related compositions and methods

Abstract

Compositions and methods relating to DNase fusion polypeptides are disclosed. The fusion polypeptides include a biologically active DNase joined to the amino-terminus of an immunoglobulin Fc region via a flexible polypeptide linker (e.g., a linker containing at least 26 amino acid residues). Typically, the DNase is a hyperactive and/or actin-resistant DNase1 variant (e.g., a variant of human DNase1 having one or more amino acid substitutions selected from substitutions at Asp-53, Tyr-65, Glu-69, Arg-74, Gly-105, and Ala-114 according to amino acid position numbering of mature wild-type human DNase1) or a DNase1L3 variant (e.g., a variant of human DNase1L3 in which the native nuclear localization signals are removed). In some embodiments, the fusion polypeptide includes a polypeptide segment located carboxyl-terminal to the Fc region and which may be, e.g., a biologically active paraoxonase. Also disclosed are dimeric proteins comprising first and second DNase fusion polypeptides as disclosed herein. The fusion polypeptides and dimeric proteins are useful in methods for therapy, including methods for treating diseases and disorders characterized by NETosis.

What is claimed is: 
     
         1 . A DNase-Fc fusion polypeptide comprising the amino acid sequence shown in
 (i) residues 21-538 or 21-537 of SEQ ID NO:18;   (ii) residues 21-548 or 21-547 of SEQ ID NO:20;   (iii) residues 21-538 or 21-537 of SEQ ID NO:86;   (iv) residues 21-538 or 21-537 of SEQ ID NO:88; or   (v) residues 21-538 or 21-537 of SEQ ID NO:90.   
     
     
         2 . A dimeric protein comprising a first fusion polypeptide and a second fusion polypeptide, wherein each of the first and second fusion polypeptides is a fusion polypeptide as defined in  claim 1 . 
     
     
         3 . A polynucleotide encoding the fusion polypeptide of  claim 1 . 
     
     
         4 . An expression cassette comprising a DNA segment encoding the fusion polypeptide of  claim 1 , wherein the DNA segment is operably linked to a promoter. 
     
     
         5 . A cultured cell into which has been introduced the expression cassette of  claim 4 , wherein the cell expresses the DNA segment. 
     
     
         6 . The cultured cell of  claim 5 , wherein the cultured cell is a stable cell line comprising the expression cassette within its genomic DNA, and wherein the stable cell line constitutively expresses the DNA segment. 
     
     
         7 . The cultured cell of  claim 6 , wherein the stable cell line is a Chinese hamster ovary (CHO) cell line. 
     
     
         8 . A method of making a fusion polypeptide, the method comprising:
 culturing a cell into which has been introduced the expression cassette of  claim 4 , wherein the cell expresses the DNA segment and the encoded fusion polypeptide is produced; and   recovering the fusion polypeptide.   
     
     
         9 . The method of  claim 8 , wherein the encoded fusion polypeptide is produced in the cell and recovered as a dimeric protein. 
     
     
         10 . A method for treating a disease or disorder characterized by NETosis, the method comprising:
 administering to a subject having the disease or disorder characterized by NETosis an effective amount of a dimeric protein of  claim 2 .   
     
     
         11 . The method of  claim 10 , wherein the disease or disorder is selected from the group consisting of an inflammatory lung disease, an inflammatory skin disease, an autoimmune disease, an autoinflammatory disease, a neurological disease or disorder, a metabolic disease, a cardiovascular disease, a fibrotic disease, thrombosis, sepsis, and ischemia reperfusion. 
     
     
         12 . The method of  claim 11 , wherein the inflammatory lung disease is selected from the group consisting of chronic obstructive pulmonary disease (COPD), asthma, cystic fibrosis (CF), bronchiectasis, hypoxia, acute respiratory distress syndrome (ARDS), and interstitial lung disease. 
     
     
         13 . The method of  claim 11 , wherein the inflammatory skin disease is selected from the group consisting of psoriasis and atopic dermatitis. 
     
     
         14 . The method of  claim 11 , wherein the autoimmune disease is selected from the group consisting of systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), psoriasis, type 1 diabetes mellitus, antiphospholipid syndrome, vasculitis, and systemic sclerosis. 
     
     
         15 . The method of  claim 11 , wherein the autoinflammatory disease is an inflammatory bowel disease (MD). 
     
     
         16 . The method of  claim 11 , wherein the neurological disease or disorder is selected from the group consisting of a chronic neurodegenerative disease, a central nervous system (CNS) infection, and ischemic stroke. 
     
     
         17 . The method of  claim 11 , wherein the metabolic disease is selected from the group consisting of type 2 diabetes and obesity. 
     
     
         18 . The method of  claim 11 , wherein the fibrotic disease is selected from the group consisting of systemic sclerosis, systemic lupus erythematosus, an inflammatory lung disease, a chronic liver disease, and a chronic kidney disease. 
     
     
         19 . The method of  claim 10 , wherein the disease or disorder is a cancer. 
     
     
         20 . The method of  claim 19 , wherein the cancer is selected from the group consisting of breast cancer, colorectal cancer, hepatocellular carcinoma, melanoma, mesothelioma, non-small cell lung cancer, small cell lung cancer, renal cell carcinoma, basal cell carcinoma, cutaneous squamous cell carcinoma, esophageal squamous cell carcinoma, Hodgkin lymphoma, head and neck squamous cell carcinoma, urothelial carcinoma, cervical cancer, endometrial carcinoma, esophageal carcinoma, gastric carcinoma, Merkel cell carcinoma, and large B cell lymphoma. 
     
     
         21 . The method of  claim 19 , wherein the method is a combination therapy comprising an immune checkpoint inhibitor. 
     
     
         22 . The method of  claim 21 , wherein the immune checkpoint inhibitor is selected from the group consisting of anti-PD-1/PD-L1, anti-CTLA-4, and a combination thereof.