Abstract 1068: Tumor infiltrating lymphocytes (TIL) recognize a unique and diverse number of non-synonymous somatic mutations in patients with metastatic breast cancer
Authors: Zacharakis Nikolaos , Kim Peter , Robbins Paul , Gartner Jared , Prickett Todd , Hill Victoria , Paria Biman , Feldman Steven , Goff Stephanie , Rosenberg Steven 
Published: September 2020

Introduction: Adoptive transfer of tumor infiltrating lymphocytes (TIL) can mediate long-term durable regression in patients with metastatic melanoma, a type of cancer which is characterized by a high number of mutated genes and pronounced lymphocytic infiltrate. In contrast, metastatic breast cancer expresses far fewer somatic mutations and may contain lower levels of TIL. This pilot study investigated the ability to identify immunogenic non-synonymous somatic mutations which can serve as targets for adoptively transferred autologous T cells into patients with metastatic breast cancer, refractory to other treatments.

Methods: WES and RNAseq were performed on metastatic lesions resected in the Surgery branch, NCI, NIH from patients with metastatic breast cancer. Non-synonymous somatic mutations were identified and, in parallel, TIL from the same lesions were grown ex vivo in the presence of IL-2. The somatic mutations were tested for recognition by the autologous TIL using previously described functional assays, involving tandem mini-genes (TMG), long peptides or predicted minimal peptide approaches. For some patients, the mutation-reactive T cells were sorted by FACS and the respective TCR was identified by single cell sequencing.

Results: Metastatic tumor lesions from 37 patients with breast cancer were studied and all of them were found to contain and express non-synonymous somatic mutations (range: 4-1788, median: 102). TIL were successfully grown ex vivo from the tumors of all patients. Following functional assays (IFN-γ secretion and 4-1BB/OX-40 upregulation), it was found that autologous TIL recognized at least one (range: 1-11, median: 3) mutated product in 25 of 37 patients (68%) tested. Seventy-five percent of those were recognized by CD4+ and 25% by CD8+ T cells. All identified immunogenic tumor mutations (n=88) were unique and not shared by other patients. Sixty-four unique TCR pairs were isolated from the reactive cells and verified to recognize 30 immunogenic tumor mutations (range 1-7 TCRs/neoantigen).

Conclusions: Patients with metastatic breast cancer contain a substantial number of non-synonymous somatic mutations and mutation-reactive TIL were found in the majority of those patients. The immunogenicity of tumor-specific mutations in metastatic breast cancer can be the platform for a highly personalized adoptive T cell transfer of selected TIL or mutation-reactive TCR transduced T cells targeting those tumor mutated genes.

Citation Format: Nikolaos Zacharakis, Peter Kim, Paul Robbins, Jared Gartner, Todd Prickett, Victoria Hill, Biman Paria, Steven Feldman, Stephanie Goff, Steven Rosenberg. Tumor infiltrating lymphocytes (TIL) recognize a unique and diverse number of non-synonymous somatic mutations in patients with metastatic breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1068.