Devices and methods for isolating tumor infiltrating lymphocytes and uses thereof

Abstract

The present invention provides methods for isolating and cryopreserving tumor infiltrating lymphocytes (TILs) and producing therapeutic populations of TILs, including methods via use of a kit and a semi-automatic device for aseptic disaggregation, enrichment, and cryopreservation of a resected tumor prior to expansion of the TIL population. The present invention also provides methods for expansion, and/or stabilization of TILs, for instance UTILs, compositions involving the same and methods of treatment involving the same.

What is claimed is: 
     
         1 . A method of identifying potent genetically modified T-cells in a population of TILs, comprising:
 in an isolated and ex vivo expanded population of TILs derived from a subject's cancer tissue, identifying the presence of genetically modified T-cells expressing a combination of markers, the combination comprising each of:
 CD107a; 
 CD3; 
 IFN-γ; and, 
 a marker for identifying a genetic modification of the T-cells; 
   wherein, a genetically modified T-cell is determined to be potent by the presence of the combination of markers.   
     
     
         2 . The method of  claim 1 , wherein the genetic modification is transduction with a vector. 
     
     
         3 . The method of  claim 1 , wherein the isolated and ex vivo expanded population of TILs has undergone one or more processes selected from the group consisting of disaggregation, cryopreservation, thawing, incubation, washing, isoform enrichment, activation with an anti-CD3 antibody, co-culturing with an inactivated feeder cell, and co-culturing with an activator cell. 
     
     
         4 . The method of  claim 1 , wherein the isolated and ex vivo expanded population of TILs has undergone ex vivo expansion comprising outgrowth and rapid expansion. 
     
     
         5 . The method of  claim 1 , wherein the identifying comprises intracellular staining and flow cytometry. 
     
     
         6 . A method of determining potency of a population of TILs, comprising:
 in an isolated and ex vivo expanded population of TILs derived from a subject's cancer tissue, quantifying genetically modified T-cells expressing a combination of markers, the combination comprising each of:
 a T-cell expressing CD107a; 
 a T-cell expressing CD3; 
 a T-cell expressing IFN-γ; and, 
 a T-cell expressing a marker for identifying a genetic modification in the T-cell; 
   
       wherein, a genetically modified T-cell is determined to be potent by the presence of the combination of markers;
 quantifying the total number of genetically modified T-cells in the expanded population of TILs; and, 
 determining the percent potency of the expanded population of TILs by the ratio of genetically modified T-cells expressing the combination of markers to the total number of T-cells in the expanded population of TILs. 
 
     
     
         7 . The method of  claim 6 , wherein the genetic modification is transduction with a vector. 
     
     
         8 . The method of  claim 6 , wherein the isolated and ex vivo expanded population of TILs has undergone one or more processes selected from the group consisting of disaggregation, cryopreservation, thawing, incubation, washing, isoform enrichment, activation with an anti-CD3 antibody, co-culturing with an inactivated feeder cell, and co-culturing with an activator cell. 
     
     
         9 . The method of  claim 6 , wherein the isolated and ex vivo expanded population of TILs has undergone ex vivo expansion comprising outgrowth and rapid expansion. 
     
     
         10 . The method of  claim 6 , wherein the identifying comprises intracellular staining and flow cytometry. 
     
     
         11 . A method for treating cancer in a subject comprising identifying potent genetically modified T-cells in a population of TILs, comprising:
 in an isolated and ex vivo expanded population of TILs derived from a subject's cancer tissue, identifying the presence of genetically modified T-cells expressing a combination of markers, the combination comprising each of:
 CD107a; 
 CD3; 
 IFN-γ; and, 
 a marker for identifying a genetic modification of the T-cells, 
   wherein, a genetically modified T-cell is determined to be potent by the presence of the combination of markers,   and administering to the subject the identified potent genetically modified T-cells.   
     
     
         12 . The method of  claim 11 , wherein the genetic modification is transduction with a vector. 
     
     
         13 . The method of  claim 11 , wherein the isolated and ex vivo expanded population of TILs has undergone one or more processes selected from the group consisting of disaggregation, cryopreservation, thawing, incubation, washing, isoform enrichment, activation with an anti-CD3 antibody, co-culturing with an inactivated feeder cell, and co-culturing with an activator cell.