Abstract PR11: Enhanced in vivo persistence and therapeutic activity of combined transferred polyclonal melanoma-reactive CTL generated in presence of IL-21 and anti-CTLA4.
Authors: Chapuis Aude G. , Thompson John A. , Margolin Kim A. , Byrd David , Bhatia Shailender , Lee Sylvia M. , Lai Ivy P. , Roberts Ilana M. , Sloan Heather L. , Greenberg Philip D. , Yee Cassian 
Published: August 2014

Abstract
Monotherapy with a blocking human anti-CTLA4 mAb achieves disease control in 20-28% of patients (pts), but failure to benefit in the remaining patients may reflect inadequate induction and/or expansion of melanoma reactive T cells following CTLA4 blockade. We have been exploring a combined biologic strategy involving the adoptive transfer of melanoma-reactive CD8+ cytotoxic T lymphocytes (CTL) with concurrent administration of anti-CTLA4 as a means to enhance and broaden the anti-tumor effect of the transferred CTL. Autologous melanoma-specific CTL were generated by stimulating CTL lines in the presence of the γc-chain cytokine Interleukin-21 (IL-21), which programs responding antigen-specific cells to retain characteristics of central memory T cells (Tcm). Responding cells were sorted for binding of a peptide-MHC (pMHC) multimer to obtain a purified tumor-reactive polyclonal product. We report on 7 pts with bulky, therapy-resistant melanoma who received 1010/m2 polyclonal IL-21-derived CTL lines targeting MART1. The infusions were preceded by intravenous cyclophosphamide 300 mg/m2 and followed by low-dose subcutaneous IL-2 (250,000 IU/m2 BID) x 14 days to promote in vivo expansion and survival. Anti-CTLA4 (ipilimumab) was administered at 3mg/kg every three weeks for four doses, starting on the day after the CTL infusion. The treatment was well tolerated, with no unexpected or serious (grade ≥3) related toxicities. 5/7 evaluable pts exhibited stable disease (SD) or partial responses (PR) by RECIST 1.1 criteria at 6 weeks, and 4/6 exhibited SD at 12 weeks. One pt who had failed previous ipilimumab monotherapy and prior CTL therapy targeting MART1 without CTLA4 blockade had a noteworthy response to the current protocol. He had SD at 12 weeks, then experienced a delayed objective response with 66% reduction in tumor mass at 28+ weeks. Persistence of the infused CTL at frequencies &gt;2x baseline levels was documented at 6 weeks (6/6 evaluable pts) and 12 weeks (4/4 evaluable pts), whereas such levels of persistence beyond 6 weeks was previously typically observed in only 15-20% of pts in previous studies. The cells infused into each pt expressed moderate/detectable levels of CD28 prior to infusions, but analysis of persisting CTL detected by specific multimer binding in vivo in responding pts revealed progressively higher expression of CD28, consistent with enhanced CD28:CD80/86 interactions in the presence of CTLA4 blockade providing survival advantages. Additionally, infused multimer+ T cells continued to proliferate (multimer+ cells expressed Ki-67), secreted multiple cytokines (IFNγ, TNFα and IL-2) in response to cognate antigen, and acquired phenotypic characteristics of Tcm (including expression of CD27, CD127, CD62L and CCR7) within 12 weeks of observation after infusion. Most significantly, all pts who had clinical benefit (prolonged SD or PR) developed de novo anti-tumor responses (epitope spreading) to non-targeted antigens (NY-ESO-1, Tyrosinase, gp100 and MAGE A3) within 12 weeks, potentially reflecting release of tumor antigens following tumor lysis by the infused CTL in a pro-inflammatory, pro-immunogenic context induced by concurrent anti-CTLA4 blockade. Our preliminary results suggest that treatment with polyclonal, melanoma-targeting CTL generated in the presence of IL-21 and combined with ipilimumab is safe. Blockade of CTLA4 may enhance the activity/persistence of the infused melanoma-specific T cells in vivo. Additionally, CTL-induced tumor cell lysis in the presence of CTLA4 blockade may promote induction of anti-tumor responses to non-targeted antigens, enhancing the biologic activity of the combined regimen.
This abstract is also presented as Poster A14.
Citation Format: Aude G. Chapuis, John A. Thompson, Kim A. Margolin, David Byrd, Shailender Bhatia, Sylvia M. Lee, Ivy P. Lai, Ilana M. Roberts, Heather L. Sloan, Philip D. Greenberg, Cassian Yee. Enhanced in vivo persistence and therapeutic activity of combined transferred polyclonal melanoma-reactive CTL generated in presence of IL-21 and anti-CTLA4. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr PR11.