The following articles have been recommended for further reading in the field of cancer immunotherapy by Dr. Cornelis J. M. Melief, JITC Co-Section Editor for the Basic Tumor Immunology Section.
“Sensitive identification of neoantigens and cognate TCRs in human solid tumors” by Marion Arnaud et al
Identification of patient-specific neoantigens and cognate T cell receptors historically has been hindered by a low frequency of tumor-specific tumor infiltrating lymphocytes (TILs) and discordance between peripheral blood lymphocytes and TILs. NeoScreen is an in vitro TIL expansion and screening tool described by Marion Arnaud et al that optimizes sensitivity of antigen identification and enriches for rare tumor-antigen-specific CD8+ T cells. Unlike conventional culture methods, TILs sourced from whole tumor fragments or dissociated tumor cells are exposed to autologous CD40-activated B cells presenting the antigen of choice and engineered to optimize potency by co-electroporation of RNA encoding the immune stimulatory ligands 4-1BB and OX40 and IL-2. Compared to conventional TIL cultures, NeoScreen identified a significantly higher frequency of tumor antigen-specific TILs, some of which were observed exclusively with NeoScreen. An average of three tumor epitopes per patient were identified with NeoScreen compared to one tumor epitope with conventional techniques. NeoScreen enriched for TILs reactive to neoepitopes or tumor-associated antigens and the frequency for targeting epitopes was ~67-fold higher compared to conventional TIL cultures. TCRbeta sequencing confirmed a marked expansion of TILs specific for tumor antigens identified in ex vivo tumor samples with NeoScreen. In vitro, T cell lines expressing TCRs identified by NeoScreen were reactive against autologous tumor for 31 different neoantigens. Furthermore, adoptive transfer of peripheral blood T cells transduced with tumor antigen-specific TCRs cloned from NeoScreen TILs led to regression of established patient-derived xenograft tumors in human IL-2-expressing immunodeficient murine models.
Why this matters: NeoScreen uses a novel approach to increase the sensitivity of tumor-specific antigen identification and cognate TCR isolation, a technique that may improve the efficacy of cancer vaccines and adoptive cell therapy.
“Determinants of anti-PD-1 response and resistance in clear cell renal cell carcinoma” by Lewis Au et al
A common feature of clear cell renal cell carcinoma (ccRCC) tumors is intratumor heterogeneity, which has historically resulted in inconsistent analyses of the tumor immune microenvironment (TME). The phase II ADAPTeR study, reported by Lewis Au and colleagues, assessed the TME in multiple regions of primary and metastatic ccRCC tumors throughout the course of immune checkpoint inhibitor (ICI) treatment including at the time of surgery or disease progression. Biopsies from 15 treatment-naïve patients encompassing 115 multi-regional tumor samples were analyzed using numerous techniques, including whole exome sequencing, RNA sequencing, T cell receptor (TCR) sequencing, immunohistochemistry, multiplex immunofluorescence, flow cytometry, and single-cell RNA and TCR sequencing. Overall, no pre-treatment genomic nor mutational characteristics were able to differentiate between responding and non-responding tumors. However, pre-treatment ccRCC-specific human endogenous retrovirus (HERV) expression was significantly higher in non-responsive tumors. TCR analyses revealed that intratumoral T cells had higher TCR clonality at baseline in tumors that responded, and the expanded clones were maintained with ICI treatment. However, post-treatment novel T cell clonal expansion did not correlate with response. Single-cell analyses revealed an association between expanded, ICI-bound, granzyme B-expressing CD8+ T cells and response.
Why this matters: The multi-omic analyses of ccRCC samples in this study reveal important features of the ccRCC TME associated with clinical benefit from ICIs. Multi-regional TME analyses are needed to understand the immunogenicity of ccRCC and elucidate combination approaches that may increase the number of patients who respond to immunotherapy.
“Mapping the evolution of T cell states during response and resistance to adoptive cellular therapy” by Pavan Bachireddy et al
Integrating single cell transcriptomic, chromatin accessibility, and T cell receptor sequencing data from bone marrow samples from 12 patients with relapsed leukemia pre- and post-donor lymphocyte infusion (DLI), Pavan Bachireddy and colleagues created longitudinal profiles associated with response versus resistance. Based on gene expression signatures, 43 distinct clusters corresponding to functional states in bone marrow-infiltrating T cells pre- and post-DLI were mapped. Long-term response to DLI was associated with enrichment for T cell activation and pre-infusion T cell exhaustion, while patients with non-responsive disease displayed T cells with multiple distinct dysfunctional states. Higher phenotypic diversity and enrichment for markers of a “late differentiated” state pre-DLI were seen in T cells from patients that achieved long-term disease control post-DLI, as well as a distinct temporal dynamics of expansion of early differentiated and contraction of late differentiated T cell subsets—immunophenotypes that were supported by expression of exhaustion and costimulatory markers. Chromatin accessibility was globally similar in T cells with responding disease both pre- and post-DLI. A probabilistic model of co-expression patterns, scRNA-seq, and chromatin accessibility identified master regulators for the phenotypic states associated with response and resistance to DLI, including some transcription factors known to mediate T cell exhaustion and dysfunction, such as EOMES and GATA3, respectively. Single-cell TCR paired with scRNA-seq revealed a common ancestry for roughly one third of the terminally exhausted and precursor exhausted clones in 2 patients that responded to DLI. Notably, after DLI, a minority of the precursor exhausted T cells shared TCR clonotypes with the DLI infusion products or had TCRs that were virus-recognizing.
Why this matters: Expansion of pre-existing precursor exhausted T cells and contraction of terminally exhausted T cells is linked to the efficacy of DLI. This investigation of dynamic shifts in the T cell repertoire serves as both a target for and a marker of the efficacy of anti-leukemic adoptive cellular therapy and sets the conceptual stage for studying determinants of response to other immunotherapy modalities.
Metastasis and immune evasion from extracellular cGAMP hydrolysis by Jun Li et al
Metastatic tumors subvert inflammatory signaling through cGAS and STING and the mechanisms by which malignant cells evade immune surveillance despite abundantly present cytosolic double-stranded DNA remain poorly understood. Jun Li and colleagues demonstrate a role for ENPP1 in establishing an immunosuppressive microenvironment through the degradation of the extracellular cGAS metabolite cGAMP. In multiple syngeneic murine models of breast and colorectal cancer, loss of ENPP1 was associated with longer survival and reduced incidences of metastasis. Conditioned media from cells lacking either cGAS or ENPP1 had roughly 40% higher levels of extracellular adenosine compared to the parental lines. Deficiency in cGAS or ENPP1 led to impaired migration in vitro and exogenous addition of cGAMP rescued migration only in the cGAS-deficient cells. Furthermore, in murine models of TNBC, combined knockout of STING and ENPP1 were additive in terms of reduced metastasis and extended survival. Tumors lacking ENPP1 had increased invasion of granulocytic cells, NK cells, and lymphocytes compared to the wild-type clones, which was associated with increased susceptibility to checkpoint blockade. Enforced overexpression of ENPP1 led to complete resistance to anti-CTLA-4 plus anti-PD-1 therapy. Data from The Cancer Genome Atlas revealed an inverse relationship between survival and expression of ENPP1 and the RNA and protein level across multiple tumor subtypes. High protein levels of ENPP1 was further associated with lower CD8+ T cell densities. In breast cancers, sarcomas, and melanomas, gene set enrichment analyses revealed upregulation of inflammatory pathways related to allograft rejection and interferon responses for tumors with high expression of cGAS, a signature that was further enhanced when ENPP1 was elevated.
Why this matters: These findings identify cGAMP as a major contributor to extracellular adenosine in the immunosuppressive tumor microenvironment and reveal ENPP1 as a potential target to increase immune-susceptibility of tumors with no obvious downsides.
“Overcoming PD-1 Blockade Resistance with CpG-A Toll-Like Receptor 9 Agonist Vidutolimod in Patients with Metastatic Melanoma” by Antoni Ribas et al
While anti-PD-1/PD-L1 immune checkpoint inhibitors (ICIs) have greatly improved outcomes for many patients with advanced melanoma, data are extremely limited to inform treatment for the subset of patients who progress on or after ICIs. Antoni Ribas and colleagues report results from a phase Ib trial evaluating safety and efficacy of combining intratumoral vidutolimod, a Toll-like receptor 9 (TLR9) agonist, and pembrolizumab in patients with advanced melanoma previously treated with an anti-PD-1 therapy. Most of the 44 patients enrolled in the study had received their last dose of prior anti-PD-1 therapy within 2 months of the study treatment and either had progressive or stable disease. After treatment, 25% of patients (95% CI 13% to 40%; n = 11) experienced either a partial response (n = 7) or complete response by (n = 4) RECIST v1.1. Responses were durable, with a median duration of response of 19.5 months (95% CI 5.8 to not estimable). Importantly, tumor regression was progressive and occurred in both injected and non-injected lesions, including visceral metastases. The combination treatment was well tolerated, with 20 patients experiencing grade 3−4 treatment-related adverse events (TRAEs), and two patients discontinuing treatment due to a TRAE. An important feature of vidutolimod is that it is contained in a virus-like particle (VLP) formed by a bacteriophage protein called Qbeta, which allows for co-activation of CD32 to further enhance dendritic cell type I IFN responses. Within 2 weeks following the first vidutolimod treatment, serum levels of anti-Qbeta antibodies and the type I IFN CXCL10 were elevated. Moreover, biopsies of injected and non-injected lesions in one patient who responded to treatment showed increased CD8+ T cell infiltration, PD-L1 expression, and IFN-related gene expression compared to baseline (pre-treatment), which was not present in biopsies from another patient with progressive disease.
Why this matters: The combination of vidutolimod and pembrolizumab is safe and effective in patients with melanoma that have progressed on anti-PD-1 therapy. Intratumoral approaches aimed at modulating the TME, such as vidutolimod, may help overcome resistance to anti-PD-1. Later-phase studies of these strategies are needed.
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