title: Expansion and characterization of human melanoma tumor-infiltrating lymphocytes (TILs).
abstract: BACKGROUND
  Various immunotherapeutic strategies for cancer are aimed at augmenting the T cell response against tumor cells. Adoptive cell therapy (ACT), where T cells are manipulated ex vivo and subsequently re-infused in an autologous manner, has been performed using T cells from various sources. Some of the highest clinical response rates for metastatic melanoma have been reported in trials using tumor-infiltrating lymphocytes (TILs). These protocols still have room for improvement and furthermore are currently only performed at a limited number of institutions. The goal of this work was to develop TILs as a therapeutic product at our institution.
  
  
  RESULTS
  TILs from 40 melanoma tissue specimens were expanded and characterized. Under optimized culture conditions, 72% of specimens yielded rapidly proliferating TILs as defined as at least one culture reaching ≥3×10(7) TILs within 4 weeks. Flow cytometric analyses showed that cultures were predominantly CD3+ T cells, with highly variable CD4+:CD8+ T cell ratios. In total, 148 independent bulk TIL cultures were assayed for tumor reactivity. Thirty-four percent (50/148) exhibited tumor reactivity based on IFN-γ production and/or cytotoxic activity. Thirteen percent (19/148) showed specific cytotoxic activity but not IFN-γ production and only 1% (2/148) showed specific IFN-γ production but not cytotoxic activity. Further expansion of TILs using a 14-day "rapid expansion protocol" (REP) is required to induce a 500- to 2000-fold expansion of TILs in order to generate sufficient numbers of cells for current ACT protocols. Thirty-eight consecutive test REPs were performed with an average 1865-fold expansion (+/- 1034-fold) after 14 days.
  
  
  CONCLUSIONS
  TILs generally expanded efficiently and tumor reactivity could be detected in vitro. These preclinical data from melanoma TILs lay the groundwork for clinical trials of ACT.
authors.affiliation: Campbell Family Institute for Breast Cancer Research, Ontario Cancer Institute, University Health Network, Toronto, Canada.
authors.forename: Linh T
authors.initials: LT
authors.lastname: Nguyen
authors.forename: Pei Hua
authors.initials: PH
authors.lastname: Yen
authors.forename: Jessica
authors.initials: J
authors.lastname: Nie
authors.forename: Nicole
authors.initials: N
authors.lastname: Liadis
authors.forename: Danny
authors.initials: D
authors.lastname: Ghazarian
authors.forename: Ayman
authors.initials: A
authors.lastname: Al-Habeeb
authors.forename: Alexandra
authors.initials: A
authors.lastname: Easson
authors.forename: Wey
authors.initials: W
authors.lastname: Leong
authors.forename: Joan
authors.initials: J
authors.lastname: Lipa
authors.forename: David
authors.initials: D
authors.lastname: McCready
authors.forename: Michael
authors.initials: M
authors.lastname: Reedijk
authors.forename: David
authors.initials: D
authors.lastname: Hogg
authors.forename: Anthony M
authors.initials: AM
authors.lastname: Joshua
authors.forename: Ian
authors.initials: I
authors.lastname: Quirt
authors.forename: Hans
authors.initials: H
authors.lastname: Messner
authors.forename: Patricia
authors.initials: P
authors.lastname: Shaw
authors.forename: Michael
authors.initials: M
authors.lastname: Crump
authors.forename: Eran
authors.initials: E
authors.lastname: Sharon
authors.forename: Pamela S
authors.initials: PS
authors.lastname: Ohashi
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002478:Cells, Cultured; D018920:Coculture Techniques; D003602:Cytotoxicity, Immunologic / Q000276:immunology; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D016130:Immunophenotyping; D016219:Immunotherapy, Adoptive / Q000379:methods; D007371:Interferon-gamma / Q000276:immunology / Q000378:metabolism; D016246:Lymphocytes, Tumor-Infiltrating / Q000276:immunology / Q000473:pathology* / Q000637:transplantation; D008297:Male; D008545:Melanoma / Q000276:immunology / Q000473:pathology* / Q000628:therapy; D008875:Middle Aged; D014407:Tumor Cells, Cultured
chemical_list: D007371:Interferon-gamma