Methods for selecting active agents for cancer treatment

Abstract

The present invention provides methods for individualizing chemotherapy, and particularly methods for individualizing neoadjuvant chemotherapy. The present invention provides methods for predicting a cancer patient's response to neoadjuvant chemotherapy, including assessing the probability of a positive response upon treatment with candidate agents prior to surgery. In various aspects, the invention involves culturing a monolayer of malignant cells from an explant of a patient's biopsy specimen, such as a transcutaneous biopsy-sized specimen, and testing the malignant cells for resistance or sensitivity to one or a plurality of candidate agents for neoadjuvant therapy. In other aspects, the invention provides methods for accurately scoring and interpreting such assays, and discloses in vitro chemoresponse results that are predictive of a patient's pathological complete response (pCR) upon receiving the corresponding treatment regimen.

1 . A method for selecting a neoadjuvant chemotherapy for a cancer patient, comprising:
 preparing an explant for cell culture from a transcutaneous biopsy specimen from the patient;   growing a cell monolayer from said explant, the monolayer comprising malignant cells;   exposing cells from said monolayer to a panel of candidate chemotherapy agents or combinations of agents, for treatment of said cancer, and at a range of concentrations;   measuring a response of the cells to each of said candidate chemotherapy agents or combinations of agents within said range;   preparing a dose-response curve for each of said candidate chemotherapy agents or combinations of said agents; and   identifying the most effective chemotherapy agent for said patient from said panel of candidate chemotherapy agents or combinations of agents.   
     
     
         2 . The method of  claim 1 , wherein said patient is a breast cancer patient. 
     
     
         3 . The method of  claim 2 , wherein the breast cancer patient has stage II or stage III breast cancer. 
     
     
         4 . The method of any of  claims 1  to  3 , wherein said transcutaneous biopsy specimen is one or more core needle biopsies. 
     
     
         5 . The method of any of  claims 1  to  4 , wherein the transcutaneous biopsy specimen contains from about 25 mg to about 100 mg of tumor tissue. 
     
     
         6 . The method of any one of  claims 1  to  4 , wherein the transcutaneous biopsy specimen contains from about 10 mg to about 50 mg of tumor tissue. 
     
     
         7 . The method of any one of  claims 1  to  6 , wherein the explants are prepared by mincing tissue from the biopsy specimen. 
     
     
         8 . The method of  claim 7 , wherein said explant has a size of about 0.5 to about 1.5 mm 3 . 
     
     
         9 . The method of  claim 8 , wherein the tissue is not exposed to enzymatic treatment. 
     
     
         10 . The method of any one of  claims 1  to  9 , wherein said monolayer contains at least 65% epithelial cells. 
     
     
         11 . The method of any one of  claims 1  to  9 , wherein said monolayer contains at least 75% epithelial cells. 
     
     
         12 . The method of any one of  claims 1  to  11 , wherein the cells of said monolayer are suspended prior to confluency, and seeded for testing. 
     
     
         13 . The method of  claim 12 , wherein the cells of said monolayer are seeded in wells of one or more microtiter plate(s). 
     
     
         14 . The method of any one of  claims 1  to  13 , wherein said panel of candidate chemotherapy agents or combinations of agents, comprises two or more of cyclophosphamide, docetaxel, doxorubicin, 5-fluorouracil, germcitabine, irinotecan, methotrexate, paclitaxel, capecitabine, cisplatin, epirubicin, etoposide, vinorelbine, and combinations thereof. 
     
     
         15 . The method of  claim 14 , wherein said panel of candidate chemotherapy agents comprises at least five of cyclophosphamide, docetaxel, doxorubicin, 5-fluorouracil, germcitabine, irinotecan, methotrexate, paclitaxel, capecitabine, cisplatin, epirubicin, etoposide, vinorelbine, and combinations thereof. 
     
     
         16 . The method of  claim 14 , wherein said panel of candidate chemotherapy agents comprises 5-flurouracil, epirubicin, cyclophosphamide, docetaxel, and optionally combinations thereof. 
     
     
         17 . The method of any one of  claims 1  to  16 , wherein the panel comprises a docetaxel/5-fluorouracil combination. 
     
     
         18 . The method of  claim 17 , wherein said sensitivity to the docetaxel/5-flurouracil combination is predictive of a breast cancer patient's complete response to said neoadjuvant therapy. 
     
     
         19 . The method of any one of  claims 1  to  18 , wherein measuring the response of the cells comprises measuring cell growth or cell death in the presence of each agent or combination. 
     
     
         20 . The method of any one of  claims 1  to  19 , further comprising, calculating an adjusted area under said dose-response curve (aAUC) for each agent or combination in the panel. 
     
     
         21 . The method of  claim 20 , wherein sensitivity or resistance to each agent or combination is determined by comparing each aAUC score to a cut-off value. 
     
     
         22 . The method of any one of  claims 1  to  21 , wherein the method is fully or partially automated. 
     
     
         23 . The method of any one of  claims 1  to  22 , wherein the patient is a Her2 positive breast cancer patient. 
     
     
         24 . The method of any one of  claims 1  to  22 , wherein the patient is a Her2 negative breast cancer patient. 
     
     
         25 . The method of any one of  claims 1  to  24 , wherein the patient's cells exhibit a variable response to agents and combinations in the panel. 
     
     
         26 . The method of any one of  claims 1  to  25 , further comprising, generating a report comparing chemosensitivity of the patient's cells across the panel. 
     
     
         27 . A method for selecting a chemotherapy treatment for a cancer patient, comprising:
 preparing an explant for cell culture from a biopsy specimen from the patient;   growing a cell monolayer from said explant, the monolayer comprising malignant cells;   exposing cells from said monolayer to a panel of candidate chemotherapy agents or combinations of agents, for treatment of said cancer, and at a range of concentrations;   measuring a response of the cells to each of said candidate chemotherapy agents or combinations of agents within said range;   preparing a dose-response curve for each of said candidate chemotherapy agents or combinations of said agents;   determining an aAUC for each dose-response curve; and   identifying the most effective chemotherapy agent for said patient from said panel of candidate chemotherapy agents or combinations of agents.   
     
     
         28 . The method of  claim 27 , wherein the aAUC accounts for changes in cytotoxicity between dose points along a dose response curve, and assigns weights relative to the degree of changes in cytotoxicity between dose points. 
     
     
         29 . The method of  claim 28 , wherein the changes in cytotoxicity between dose points along the dose-response curve are represented by a local slope, and the local slopes and weighted along the dose-response curve to emphasize cytotoxicity. 
     
     
         30 . The method of any one of  claims 27 - 29 , wherein aAUC is determined by:
 calculating a Cytotoxity Index (CI) for each dose, where CI=Mean drug/Mean control;   calculating a local slope (S d ) at each dose point, where S d =(CI d −CI d-1 )/Unit of Dose, or S d =(CI d-1 −CI d )/Unit of Dose;   calculating slope weight at each dose point, where W d =1−S d ; and   calculating aAUC, where aAUC=ΣW d  CI d , and where, d=1, 2, . . . , 10.   
     
     
         31 . The method of any one of  claims 27  to  30 , wherein sensitivity or resistance to each agent or combination is determined by comparing each aAUC score to a cut-off value. 
     
     
         32 . The method of any one of  claims 27  to  30 , wherein an effective agent or combination is identified by comparing the aAUC scores for the plurality of agents or combinations.