Assays for detecting t cell immune subsets and methods of use thereof

Abstract

The present disclosure provides methods for measuring the number of CD4+ OX40+ Foxp3+ lymphocytes in a sample containing cancer cells and lymphocytes obtained from a subject by labeling lymphocytes that show CD4 expression in the sample, then labeling lymphocytes that show OX40 expression in the sample, then labeling lymphocytes that show Foxp3 expression in the sample, then measuring the number of CD4+ OX40+ Foxp3+ lymphocytes in the sample. Further provided are methods for determining the prognosis of a subject, predicting responsiveness of a subject having cancer to an OX40 agonist treatment, and methods for treating or delaying progression of cancer based on the number of CD4+ OX40+ Foxp3+ lymphocytes in a sample.

1 . A method for determining prognosis of a subject having cancer, comprising:
 (a) measuring the number of CD4+ OX40+ Foxp3+ lymphocytes in a sample comprising cancer cells and lymphocytes obtained from the subject; and   (b) determining the prognosis of the subject based on the number of CD4+ OX40+ Foxp3+ lymphocytes in the sample, as compared with a reference, wherein an increased number of CD4+ OX40+ Foxp3+ lymphocytes in the sample indicates that the subject may have an improved prognosis.   
     
     
         2 . The method of  claim 1 , wherein the improved prognosis comprises increased overall survival. 
     
     
         3 . The method of  claim 1 , wherein the improved prognosis comprises increased progression-free survival. 
     
     
         4 . A method for treating or delaying progression of cancer in a subject, comprising:
 (a) measuring the number of CD4+ OX40+ Foxp3+ lymphocytes in a sample comprising cancer cells and lymphocytes obtained from the subject;   (b) determining the number of CD4+ OX40+ Foxp3+ lymphocytes in the sample, as compared with a reference; and   (c) if the number of CD4+ OX40+ Foxp3+ lymphocytes in the sample is higher than the reference, administering to the subject an effective amount of an OX40 agonist.   
     
     
         5 . A method for treating or delaying progression of cancer in a subject, comprising administering to the subject an effective amount of an OX40 agonist, wherein a sample comprising cancer cells and lymphocytes obtained from the subject has an increased number of CD4+ OX40+ Foxp3+ lymphocytes, as compared with a reference. 
     
     
         6 . A method for predicting responsiveness of a subject having cancer to an OX40 agonist treatment, comprising:
 (a) measuring the number of CD4+ OX40+ Foxp3+ lymphocytes in a sample comprising cancer cells and lymphocytes obtained from the subject; and   (b) classifying the subject as a responsive or non-responsive subject based on the number of CD4+ OX40+ Foxp3+ lymphocytes in the sample, as compared with a reference, wherein an increased number of CD4+ OX40+ Foxp3+ lymphocytes in the sample indicates the subject may be responsive to the OX40 agonist treatment.   
     
     
         7 . The method of  claim 4 , wherein the number of CD4+ OX40+ Foxp3+ lymphocytes is a median, mean or average number of CD4+ OX40+ Foxp3+ lymphocytes in different regions of interest of the sample from the subject. 
     
     
         8 . The method of  claim 7 , wherein the number of CD4+ OX40+ Foxp3+ lymphocytes is normalized to total cells in the region of interest of the sample. 
     
     
         9 . The method of any  claim 4 , wherein the reference is based on the number of CD4+ OX40+ Foxp3+ lymphocytes in a sample comprising cancer cells and lymphocytes obtained from a cancer having the same type and/or stage as the cancer of the subject. 
     
     
         10 . The method of  claim 9 , wherein the reference is a median, mean, or average number of CD4+ OX40+ Foxp3+ lymphocytes in samples obtained from cancers having the same type and/or stage as the cancer of the subject. 
     
     
         11 . The method of  claim 4 , wherein the OX40 agonist is an agonist anti-human OX40 antibody. 
     
     
         12 - 16 . (canceled) 
     
     
         17 . The method of  claim 11 , wherein the antibody comprises (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:2, 8 or 9; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:3, 10, 11, 12, 13, or 14; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:4, 15 or 19; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:5; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:6; and (f) HVR-L3 comprising an amino acid sequence selected from SEQ ID NO:7, 22, 23, 24, 25, 26, 27 or 28. 
     
     
         18 - 20 . (canceled) 
     
     
         21 . The method of  claim 11 , wherein the antibody is MEDI6469 or MEDI0562. 
     
     
         22 . The method of  claim 4 , wherein the OX40 agonist comprises one or more extracellular domains of OX40L. 
     
     
         23 . The method of  claim 4 , wherein the OX40 agonist is MEDI6383. 
     
     
         24 . (canceled) 
     
     
         25 . A method for measuring the number of CD4+ OX40+ Foxp3+ lymphocytes in a sample comprising cancer cells and lymphocytes obtained from a subject, comprising the steps of:
 (a) labeling lymphocytes that show CD4 expression in the sample;   (b) labeling lymphocytes that show OX40 expression in the sample after step (a);   (c) labeling lymphocytes that show Foxp3 expression in the sample after step (b); and   (d) measuring the number of CD4+ OX40+ Foxp3+ lymphocytes in the sample after step (c).   
     
     
         26 - 30 . (canceled) 
     
     
         31 . The method of  claim 4 , wherein the cancer is selected from the group consisting of non-small cell lung cancer, renal cell carcinoma, bladder cancer, ovarian cancer, glioblastoma, neuroblastoma, melanoma, triple-negative breast carcinoma, gastric cancer, colorectal cancer, and hepatocellular carcinoma. 
     
     
         32 - 35 . (canceled) 
     
     
         36 . A method for determining prognosis of a subject having cancer, comprising:
 (a) measuring the number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in a sample comprising metastatic cancer cells and lymphocytes obtained from the subject; and   (b) determining the prognosis of the subject based on the number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in the sample, as compared with a reference, wherein an increased number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in the sample indicates that the subject may have an improved prognosis.   
     
     
         37 . The method of  claim 36 , wherein the improved prognosis comprises increased overall survival. 
     
     
         38 . The method of  claim 36 , wherein the improved prognosis comprises increased progression-free survival. 
     
     
         39 . A method for treating or delaying progression of cancer in a subject, comprising:
 (a) measuring the number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in a sample comprising metastatic cancer cells and lymphocytes obtained from the subject;   (b) determining the number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in the sample, as compared with a reference; and   (c) if the number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in the sample is higher than the reference, administering to the subject an effective amount of an OX40 agonist.   
     
     
         40 . A method for treating or delaying progression of cancer in a subject, comprising administering to the subject an effective amount of an OX40 agonist, wherein a sample comprising metastatic cancer cells and lymphocytes obtained from the subject has an increased number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes, as compared with a reference. 
     
     
         41 . A method for predicting responsiveness of a subject having cancer to an OX40 agonist treatment, comprising:
 (a) measuring the number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in a sample comprising metastatic cancer cells and lymphocytes obtained from the subject; and   (b) classifying the subject as a responsive or non-responsive subject based on the number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in the sample, as compared with a reference, wherein an increased number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in the sample indicates the subject may be responsive to the OX40 agonist treatment.   
     
     
         42 . The method of  claim 39 , wherein the number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes is a median, mean or average number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in different regions of interest in the sample from the subject. 
     
     
         43 . The method of  claim 42 , wherein the number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes is normalized to total cells in the region in the sample. 
     
     
         44 . The method of  claim 39 , wherein the reference is based on the number of OX40+, CD4+ OX40+ Foxp3+, or CD4+ OX40+ Foxp3− lymphocytes in a sample comprising metastatic cancer cells and lymphocytes obtained from a cancer having the same type and/or stage as the cancer of the subject. 
     
     
         45 . The method of  claim 44 , wherein the reference is a median, mean, or average number of CD4+ OX40+ Foxp3+ lymphocytes in samples obtained from cancers having the same type and/or stage as the cancer of the subject. 
     
     
         46 . The method of  claim 39 , wherein the OX40 agonist is an agonist anti-human OX40 antibody. 
     
     
         47 - 51 . (canceled) 
     
     
         52 . The method of  claim 46 , wherein the antibody comprises (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:2, 8 or 9; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:3, 10, 11, 12, 13, or 14; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:4, 15 or 19; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:5; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:6; and (f) HVR-L3 comprising an amino acid sequence selected from SEQ ID NO:7, 22, 23, 24, 25, 26, 27 or 28. 
     
     
         53 - 55 . (canceled) 
     
     
         56 . The method of  claim 46 , wherein the antibody is MEDI6469 or MEDI0562. 
     
     
         57 . The method of  claim 39 , wherein the OX40 agonist comprises one or more extracellular domains of OX40L. 
     
     
         58 . (canceled) 
     
     
         59 . (canceled) 
     
     
         60 . The method of  claim 39 , wherein the cancer is selected from the group consisting of non-small cell lung cancer, renal cell carcinoma, bladder cancer, ovarian cancer, glioblastoma, neuroblastoma, melanoma, triple-negative breast carcinoma, gastric cancer, colorectal cancer, and hepatocellular carcinoma. 
     
     
         61 . (canceled) 
     
     
         62 . (canceled)