Adoptive Cellular Therapy with Autologous Tumor-Infiltrating Lymphocytes and T-cell Receptor–Engineered T Cells Targeting Common p53 Neoantigens in Human Solid Tumors

Abstract

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 .

Improving the sensitivity of in vivo CRISPR off-target detection with DISCOVER-Seq+

“…Next, we applied DISCOVER-Seq+ ex vivo to knock-in of a cancer neoantigen-specific transgenic T-cell receptor (tgTCR) construct into primary human T cells [31-32] . We electroporated Cas9 targeting TRA (T Cell Receptor Alpha Locus) along with a 4699 bp homology-directed repair template (HDRT) encoding a tgTCR specific for HLA-A*02 loaded with mutant p53 R175H peptide [31] , then performed DISCOVER-Seq+ 12 hours later (Fig. 3d) .…”

“…This study designed and validated DISCOVER-Seq+, the most sensitive method to-date for detecting CRISPR off-target activity in primary cells and in vivo. As CRISPR becomes an increasingly feasible approach for therapeutic genome editing [1,20,31,32,35-37] , evaluation of CRISPR off-target activity directly in clinically translatable applications is crucial. Even if a specific CRISPR gRNA does not have detectable off-target activity in one particular cell type, this may not translate to other cell types, tissues, or organisms.…”

“…For in vivo liver editing, such as to prevent cardiovascular disease [35] or to treat Transthyretin (ATTR) Amyloidosis [36] in a particular patient, initial off-target validation of gRNAs in non-human primates along with personalized validation in primary liver cells derived from that individual patient may be essential before therapy. Ex vivo generation of engineered T cells [31,32,37] further highlights a pressing need for personalized evaluation of off-target sites; since each patient receives a bespoke T cell line derived from their own cells, a precision medicine approach will be required to ensure each patient’s off-target profile is acceptable before therapy.…”

“…By leveraging MRE11 ChIP-seq with DNA-PKcs inhibition, DISCOVER-Seq+ provides the highest detection sensitivity to-date in systems ranging from ex vivo editing of primary human cells to in vivo editing of mice, setting the standard for genome-wide CRISPR off-target discovery (Table S1) . DISCOVER-Seq+ has the potential to validate the specificity profile of genome editing at numerous stages of the therapeutic development pipeline, from cell lines and primary cells to mice and potentially non-human primates [1,18,31,32,35-37] .…”

Neoadjuvant therapy alters the immune microenvironment in pancreatic cancer

“…Visceral metastases regressed in this patient, and the response was ongoing at 6 months ( 146 ). Since most neoantigens are derived from patient-specific mutations, neoantigen T-cell receptor gene therapy is highly personalized ( 147 ). In conclusion, we believe that cancer immunotherapy targeting neoantigens is a promising treatment option, but methods to reduce the cost of this highly personalized treatment warrant more research.…”