Adoptive cellular therapy (ACT) targeting neoantigens can achieve durable clinical responses in patients with cancer. Most neoantigens arise from patient-specific mutations, requiring highly individualized treatments. To broaden the applicability of ACT targeting neoantigens, we focused on TP53 mutations commonly shared across different cancer types. We performed whole-exome sequencing on 163 patients with metastatic solid cancers, identified 78 who had TP53 missense mutations, and through immunologic screening, identified 21 unique T-cell reactivities. Here, we report a library of 39 T-cell receptors (TCR) targeting TP53 mutations shared among 7.3% of patients with solid tumors. These TCRs recognized tumor cells in a TP53 mutation- and human leucocyte antigen (HLA)-specific manner in vitro and in vivo. Twelve patients with chemorefractory epithelial cancers were treated with ex vivo–expanded autologous tumor-infiltrating lymphocytes (TIL) that were naturally reactive against TP53 mutations. However, limited clinical responses (2 partial responses among 12 patients) were seen. These infusions contained low frequencies of mutant p53–reactive TILs that had exhausted phenotypes and showed poor persistence. We also treated one patient who had chemorefractory breast cancer with ACT comprising autologous peripheral blood lymphocytes transduced with an allogeneic HLA-A*02–restricted TCR specific for p53R175H. The infused cells exhibited an improved immunophenotype and prolonged persistence compared with TIL ACT and the patient experienced an objective tumor regression (-55%) that lasted 6 months. Collectively, these proof-of-concept data suggest that the library of TCRs targeting shared p53 neoantigens should be further evaluated for the treatment of patients with advanced human cancers. See related Spotlight by Klebanoff, p. 919

PURPOSE: Metastatic breast cancer (mBrCa) is most often an incurable disease with only modest responses to available immunotherapies. This study investigates the immunogenicity of somatic mutations in breast cancer and explores the therapeutic efficacy in a pilot trial of mutation-reactive tumor-infiltrating lymphocytes (TILs) in patients with metastatic disease. PATIENTS AND METHODS: Forty-two patients with mBrCa refractory to previous lines of treatment underwent surgical resection of a metastatic lesion(s), isolation of TIL cultures, identification of exomic nonsynonymous tumor mutations, and immunologic screening for neoantigen reactivity. Clinically eligible patients with appropriate reactivity were enrolled into one cohort of an ongoing phase II pilot trial of adoptive cell transfer of selected neoantigen-reactive TIL, with a short course of pembrolizumab (ClinicalTrials.gov identifier: NCT01174121 ). RESULTS: TILs were isolated and grown in culture from the resected lesions of all 42 patients with mBrCa, and a median number of 112 (range: 6-563) nonsynonymous mutations per patient were identified. Twenty-eight of 42 (67%) patients contained TIL that recognized at least one immunogenic somatic mutation (median: 3 neoantigens per patient, range: 1-11), and 13 patients demonstrated robust reactivity appropriate for adoptive transfer. Eight patients remained clinically eligible for treatment, and six patients were enrolled on a protocol of adoptive cell transfer of enriched neoantigen-specific TIL, in combination with pembrolizumab (≤ 4 doses). Objective tumor regression was noted in three patients, including one complete response (now ongoing over 5.5 years) and two partial responses (6 and 10 months). CONCLUSION: Most patients with breast cancer generated a natural immune response targeting the expressed products of their cancer mutations. Adoptive transfer of TIL is a highly personalized experimental option for patients with mBrCa shown to be capable of mediating objective responses in this pilot trial and deserves further study.

Malignant pleural mesothelioma (MPM) is an incurable, aggressive neoplasm with distinctive features, including preservation of wild-type p53, irrespective of histologic subtype. We posited that this consistent molecular characteristic represents an underexploited therapeutic target that can be approached by leveraging biologic effects of microRNA (miRNA). The Cancer Genome Atlas was surveyed to identify p53-responsive prognostic miRNA(s) in MPM. Using patient samples, in vitro MPM cell lines, and murine tumor xenograft models, we verified specific gene pathways targeted by these miRNAs, and we examined their therapeutic effects. miR-215-5p is a poor prognosis miRNA downregulated in MPM tissues, which has not been recognized previously. When miR-215-5p was ectopically re-expressed in MPM cells and delivered in vivo to tumor xenografts, it exerted significant cell killing by activating p53 function and inducing apoptosis. The mechanistic basis for this effect is due to combinatorial effects of a positive feedback loop of miR-215-MDM2-p53 signaling, additional mouse double minute 2 (MDM2)-p53 positive feedback loop(s) with other miRNAs such as miR-145-5p, and suppression of diverse gene targets associated with cell cycle dynamics not previously drug treatable in MPM clinical studies. Our results suggest a potential pathophysiologic role for and therapeutic significance of miR-215-5p in MPM.

Mesothelial cells are arranged as a monolayer on covering membranes that invest surfaces of body cavities like the pleura and peritoneum. Primary human mesothelial cell (HMC) cultures are needed for studying mesothelial cell homeostasis and developing disease models, such as wound healing or cancers. Remarkably, there is a paucity of useable HMC lines that are currently available that faithfully recapitulate normal in vivo phenotypic characteristics. Here, we present a strategy to recover HMC from human pleural tissue and to immortalize them for extended in vitro culturing. Human pleural membrane was harvested by minimally invasive surgical techniques. HMC were isolated using a two-step process combining explant cellular outgrowth from biopsy tissue and flow cytometry based on cell surface expression of cadherin-1 and CD71. Cell cultures were generated after lentiviral transfection with human telomerase. The new HMC cultures retain the same phenotypic traits and physiologic features as their in vivo counterparts, yet they can be adapted for short-term or long-term culture in large-scale in vitro experimentation. In particular, we generated a new HMC line harboring a germline mutation in breast cancer type-1-associated protein-1 ( BAP1), a causal tumor suppressor gene, that could be instrumental to malignant mesothelioma research. Patient-specific, normal HMC may serve as novel discovery tools allowing more powerful research models of both normal physiology and disease processes. Our surgically driven approach leads to a limitless resource of novel mesothelial cell cultures.