Targeting of multiple tumor-associated antigens by individual T cell receptors during successful cancer immunotherapy.

The T cells of the immune system can target tumors and clear solid cancers following tumor-infiltrating lymphocyte (TIL) therapy. We used combinatorial peptide libraries and a proteomic database to reveal the antigen specificities of persistent cancer-specific T cell receptors (TCRs) following successful TIL therapy for stage IV malignant melanoma. Remarkably, individual TCRs could target multiple different tumor types via the HLA A∗02:01-restricted epitopes EAAGIGILTV, LLLGIGILVL, and NLSALGIFST from Melan A, BST2, and IMP2, respectively. Atomic structures of a TCR bound to all three antigens revealed the importance of the shared x-x-x-A/G-I/L-G-I-x-x-x recognition motif. Multi-epitope targeting allows individual T cells to attack cancer in several ways simultaneously. Such "multipronged" T cells exhibited superior recognition of cancer cells compared with conventional T cell recognition of individual epitopes, making them attractive candidates for the development of future immunotherapies.

Author Correction: Genome-wide CRISPR-Cas9 screening reveals ubiquitous T cell cancer targeting via the monomorphic MHC class I-related protein MR1.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Genome-wide CRISPR-Cas9 screening reveals ubiquitous T cell cancer targeting via the monomorphic MHC class I-related protein MR1.

Human leukocyte antigen (HLA)-independent, T cell-mediated targeting of cancer cells would allow immune destruction of malignancies in all individuals. Here, we use genome-wide CRISPR-Cas9 screening to establish that a T cell receptor (TCR) recognized and killed most human cancer types via the monomorphic MHC class I-related protein, MR1, while remaining inert to noncancerous cells. Unlike mucosal-associated invariant T cells, recognition of target cells by the TCR was independent of bacterial loading. Furthermore, concentration-dependent addition of vitamin B-related metabolite ligands of MR1 reduced TCR recognition of cancer cells, suggesting that recognition occurred via sensing of the cancer metabolome. An MR1-restricted T cell clone mediated in vivo regression of leukemia and conferred enhanced survival of NSG mice. TCR transfer to T cells of patients enabled killing of autologous and nonautologous melanoma. These findings offer opportunities for HLA-independent, pan-cancer, pan-population immunotherapies.

Author Correction: Tertiary lymphoid structures improve immunotherapy and survival in melanoma.

An Amendment to this paper has been published and can be accessed via a link at the top of the paper.

Tertiary lymphoid structures improve immunotherapy and survival in melanoma.

Checkpoint blockade therapies that reactivate tumour-associated T cells can induce durable tumour control and result in the long-term survival of patients with advanced cancers1. Current predictive biomarkers for therapy response include high levels of intratumour immunological activity, a high tumour mutational burden and specific characteristics of the gut microbiota2,3. Although the role of T cells in antitumour responses has thoroughly been studied, other immune cells remain insufficiently explored. Here we use clinical samples of metastatic melanomas to investigate the role of B cells in antitumour responses, and find that the co-occurrence of tumour-associated CD8+ T cells and CD20+ B cells is associated with improved survival, independently of other clinical variables. Immunofluorescence staining of CXCR5 and CXCL13 in combination with CD20 reveals the formation of tertiary lymphoid structures in these CD8+CD20+ tumours. We derived a gene signature associated with tertiary lymphoid structures, which predicted clinical outcomes in cohorts of patients treated with immune checkpoint blockade. Furthermore, B-cell-rich tumours were accompanied by increased levels of TCF7+ naive and/or memory T cells. This was corroborated by digital spatial-profiling data, in which T cells in tumours without tertiary lymphoid structures had a dysfunctional molecular phenotype. Our results indicate that tertiary lymphoid structures have a key role in the immune microenvironment in melanoma, by conferring distinct T cell phenotypes. Therapeutic strategies to induce the formation of tertiary lymphoid structures should be explored to improve responses to cancer immunotherapy.

Fully Automated Workflow for Integrated Sample Digestion and Evotip Loading Enabling High-Throughput Clinical Proteomics.

Protein identification and quantification is an important tool for biomarker discovery. With the increased sensitivity and speed of modern mass spectrometers, sample preparation remains a bottleneck for studying large cohorts. To address this issue, we prepared and evaluated a simple and efficient workflow on the Opentrons OT-2 robot that combines sample digestion, cleanup, and loading on Evotips in a fully automated manner, allowing the processing of up to 192 samples in 6 h. Analysis of 192 automated HeLa cell sample preparations consistently identified ∼8000 protein groups and ∼130,000 peptide precursors with an 11.5 min active liquid chromatography gradient with the Evosep One and narrow-window data-independent acquisition (nDIA) with the Orbitrap Astral mass spectrometer providing a throughput of 100 samples per day. Our results demonstrate a highly sensitive workflow yielding both reproducibility and stability at low sample inputs. The workflow is optimized for minimal sample starting amount to reduce the costs for reagents needed for sample preparation, which is critical when analyzing large biological cohorts. Building on the digesting workflow, we incorporated an automated phosphopeptide enrichment step using magnetic titanium-immobilized metal ion affinity chromatography beads. This allows for a fully automated proteome and phosphoproteome sample preparation in a single step with high sensitivity. Using the integrated digestion and Evotip loading workflow, we evaluated the effects of cancer immune therapy on the plasma proteome in metastatic melanoma patients.

Frequency and characteristics of immune-related thyroid adverse events in patients with resected stage III/IV melanoma treated with adjuvant PD-1 inhibitors: a national cohort study.

PURPOSE: Immune-related thyroid adverse events (irTAEs) occur frequently following immune checkpoint inhibitor (ICI) therapy. The purpose of this study is to provide knowledge about the incidence, clinical timeline characteristics, associated factors of irTAEs, and potential impact on treatment efficacy in patients with melanoma receiving adjuvant ICI therapy. METHODS: A national multicenter retrospective cohort study of patients with resected stage III/IV melanoma treated with adjuvant PD-1 inhibitors between November 2018 and December 2020. Data were extracted from the Danish Metastatic Melanoma Database. The irTAEs were defined as two consecutive abnormal TSH values and subdivided into transient or persistent. RESULTS: Of 454 patients, 99 developed an irTAE (21.8%), of these were 46 transient (46.5%) and 53 persistent (53.5%). Median time to transient and persistent irTAE was 55 and 44 days, respectively (p = 0.57). A hyperthyroid phase followed by hypothyroidism was seen in 73.6% of persistent irTAEs, whereas 87% of transient irTAEs developed an isolated hypo- or hyperthyroid phase. Multiple variable analysis demonstrated an association between irTAE and female sex (HR 2.45; 95% CI 1.63-3.70; p < 0.001), but no association with recurrence-free survival (HR 0.86; 95% CI 0.50-1.48; p = 0.587) or overall survival (HR 1.05; 95% CI 0.52-2.12, p = 0.891). CONCLUSIONS: IrTAE is a common side effect to PD-1 inhibitors primarily occurring within the first 3 months, with a high risk of persistency. Female sex is a strong predictive factor. IrTAE was not associated with improved clinical outcome.

Differential ultracentrifugation enables deep plasma proteomics through enrichment of extracellular vesicles.

Human plasma is a rich source of biomedical information and biomarkers. However, the enormous dynamic range of plasma proteins limits its accessibility to mass spectrometric (MS) analysis. Here, we show that enrichment of extracellular vesicles (EVs) by ultracentrifugation increases plasma proteome depth by an order of magnitude. With this approach, more than two thousand proteins are routinely and reproducibly quantified by label-free quantification and data independent acquisition (DIA) in single-shot liquid chromatography tandem mass spectrometry runs of less than one hour. We present an optimized plasma proteomics workflow that enables high-throughput with very short chromatographic gradients analyzing hundred samples per day with deep proteome coverage, especially when including a study-specific spectral library generated by repeated injection and gas-phase fractionation of pooled samples. Finally, we test the workflow on clinical biobank samples from malignant melanoma patients in immunotherapy to demonstrate the improved proteome coverage supporting the potential for future biomarker discovery.

Earlier Recurrence Detection Using Routine FDG PET-CT Scans in Surveillance of Stage IIB to IIID Melanoma: A National Cohort Study of 1480 Patients.

BACKGROUND: The effect of routine imaging in melanoma surveillance is unknown. In 2016, Denmark was the first country in the world to implement routine imaging with positron emission tomography-computed tomography with fluorodeoxyglucose (FDG PET-CT) in a nationwide, population-based surveillance program. This study aimed to determine the impact of surveillance with routine FDG PET-CT on hazard, cumulative incidence, and absolute risk of overall, locoregional, and distant recurrence detection in patients with stage IIB to IIID cutaneous melanoma. METHODS: This retrospective, population-based, nationwide cohort study used prospectively collected data from five national health registries to compare hazard, cumulative incidence, and absolute risk of recurrence in patients with cutaneous melanoma diagnosed in 2008-2010 (cohort 1, followed with clinical examinations) and patients with cutaneous melanoma diagnosed in 2016-2017 (cohort 2, followed with clinical examinations and routine FDG PET-CT at 6, 12, 24, and 36 months). RESULTS: The study included 1480 patients with stage IIB to IIID cutaneous melanoma. Cumulative incidences of overall and distant recurrence were higher in cohort 2, with a peak difference at three years (32.3 % vs 27.5 % and 25.8 % vs. 18.5 %, respectively). The hazard of recurrence was higher in cohort 2 during the first two years, with hazard rates for overall and distant recurrence of 1.16 (95 % confidence interval [CI], 0.93-1.44) and 1.51 (95 % CI, 1.16-1.96), respectively. The patterns persisted in absolute risk estimates. CONCLUSIONS: Patients with stage IIB to IIID melanoma followed with routine FDG PET-CT had a 51 % increased hazard of distant recurrence detection within the first two years of surveillance. Future studies must determine whether this earlier recurrence detection translates into improved survival.

Combination therapy with immune check point inhibitors and acute kidney injury.

BACKGROUND: Immune checkpoint inhibitors have revolutionized the treatment of metastatic renal cell carcinoma and malignant melanoma but are also associated with a risk of severe side effects. Nephrotoxicity is an immune checkpoint inhibitor-related adverse effect, but acute kidney injury (AKI) can also be caused by other more common conditions. This study aimed to describe the incidence and causes of AKI in patients treated with combination therapy of immune checkpoint inhibitors. MATERIAL AND METHODS: This retrospective cohort study included 200 patients receiving ipilimumab and nivolumab for either metastatic renal cell carcinoma or malignant melanoma at the Department of Oncology at Copenhagen University Hospital, Herlev between 1 January 2019 and 31 December 2020. The incidence and cause of AKI within 6 months after treatment was determined. RESULTS: In the 96 patients treated for malignant melanoma 15 patients (16%) had an episode of AKI. Two of these patients had potential immune checkpoint inhibitor-related AKI both of which received treatment with a proton pump inhibitor (PPI). Of the 104 included patients with metastatic renal cell carcinoma 26 patients (25%) developed AKI. Five of these patients had potential immune checkpoint inhibitor-related AKI. Treatment with PPI before the development of AKI occurred in 4 out of these 5 patients. CONCLUSION: Patients receiving combination therapy with checkpoint inhibitors are at high risk of AKI, but different causes of AKI should always be considered. Use of PPI concurrently with ICIs is likely to increase the risk of AKI.

Quality of life and mental health in real-world patients with resected stage III/IV melanoma receiving adjuvant immunotherapy.

INTRODUCTION: Treatment with immune checkpoint inhibitors (ICI) has expanded into the adjuvant setting enhancing the importance of knowledge on the immune-related toxicities and their impact on health-related quality of life (HRQoL). Large phase 3 trials of patients with resected Stage III/IV melanoma found no effect on HRQoL during adjuvant immunotherapy. This study investigates how HRQoL was affected during and after adjuvant immunotherapy in a real-world setting. METHODS: Patients with resected melanoma treated with adjuvant nivolumab from 2018 to 2021 in Denmark were identified using the Danish Metastatic Melanoma Database (DAMMED). The study was performed as a nationwide cross-sectional analysis as a questionnaire consisting of six different validated questionnaires on HRQoL, cognitive function, fatigue, depression, fear of recurrence, and decision regret was sent to all patients in March 2021. To evaluate HRQoL during and after adjuvant treatment, patients were divided into groups depending on their treatment status when answering the questionnaire; patients in active treatment for 0-6 months, patients in active treatment for >6 months, patients who ended treatment 0-6 months ago, and patients who ended treatment >6 months ago. RESULTS: A total of 271/412 (66%) patients completed the questionnaire. Patients who ended therapy 0-6 months ago had the lowest HRQoL and had more fatigue. Patients in active treatment for >6 months had lower HRQoL and more fatigue than patients who started treatment 0-6 months ago. Patients ending therapy >6 months ago had higher HRQoL and less fatigue compared to patients who ended therapy 0-6 months ago. Multivariable analysis showed an association between HRQoL and treatment status, comorbidity, civil status, and employment status. CONCLUSIONS: Adjuvant nivolumab may affect some aspects of QoL, but the influence seems temporary. Patient characteristics, such as civil status, employment status, and comorbidity were associated with HRQoL.

Fibrotic activity quantified in serum by measurements of type III collagen pro-peptides can be used for prognosis across different solid tumor types.

Due to activation of fibroblast into cancer-associated fibroblasts, there is often an increased deposition of extracellular matrix and fibrillar collagens, e.g. type III collagen, in the tumor microenvironment (TME) that leads to tumor fibrosis (desmoplasia). Tumor fibrosis is closely associated with treatment response and poor prognosis for patients with solid tumors. To assure that the best possible treatment option is provided for patients, there is medical need for identifying patients with high (or low) fibrotic activity in the TME. Measuring unique collagen fragments such as the pro-peptides released into the bloodstream during fibrillar collagen deposition in the TME can provide a non-invasive measure of the fibrotic activity. Based on data from 8 previously published cohorts, this review provides insight into the prognostic value of quantifying tumor fibrosis by measuring the pro-peptide of type III collagen in serum of a total of 1692 patients with different solid tumor types and discusses the importance of tumor fibrosis for understanding prognosis and for potentially guiding future drug development efforts that aim at overcoming the poor outcome associated with a fibrotic TME.

Clinical advances and ongoing trials on mRNA vaccines for cancer treatment.

Years of research exploring mRNA vaccines for cancer treatment in preclinical and clinical trials have set the stage for the rapid development of mRNA vaccines during the COVID-19 pandemic. Therapeutic cancer vaccines based on mRNA are well tolerated, and the inherent advantage in ease of production, which rivals the best available conventional vaccine manufacture methods, renders mRNA vaccines a promising option for cancer immunotherapy. Technological advances have optimised mRNA-based vaccine stability, structure, and delivery methods, and multiple clinical trials investigating mRNA vaccine therapy are now enrolling patients with various cancer diagnoses. Although therapeutic mRNA-based cancer vaccines have not yet been approved for standard treatment, encouraging results from early clinical trials with mRNA vaccines as monotherapy and in combination with checkpoint inhibitors have been obtained. This Review summarises the latest clinical advances in mRNA-based vaccines for cancer treatment and reflects on future perspectives and challenges for this new and promising treatment approach.

Clinical value of routine [18F]2-fluoro-2-deoxy-d-glucose positron emission tomography scans as a decision tool for early immunotherapy discontinuation in advanced melanoma.

Routine [18F]2-fluoro-2-deoxy-d-glucose positron emission tomography (FDG-PET) may help predict clinical outcomes after response to immunotherapy. With a European Medicines Agency-recommended treatment length until disease progression or unacceptable toxicity, the optimal duration of immunotherapy remains to be defined. In a retrospective study, we retrieved from the Danish Metastatic Melanoma Database (DAMMED), all patients that were annotated as a partial or complete response based on the computed tomography (CT) of serial FDG-PET-CT scans. Patients treated with an anti-Programmed Death (PD)-1-containing regimen for <18 months, and ≥4 months without disease progression after halting anti-PD-1 were included. Cases were divided into an "elective" and a "toxicity" group based on the reason for treatment discontinuation. A total of 140 patients were included. At 29.3 months of median follow-up, a higher proportion of patients remained alive in the "elective" group (93% vs 75%, P = .0031) with an improved melanoma-specific (HR 0.07, 95% CI 0.02-0.32, P = .0041) survival (MSS). Patients without FDG-avid lesions at the time of treatment discontinuation had an improved MSS (HR 0.03, 95% CI 0.01-0.17, P = .0002), and the absence of FDG-avid lesions was the only independent predictive feature of improved MSS in multivariate analysis. In conclusion, patients with metastatic melanoma who obtain an early response and early discontinue immunotherapy have an excellent prognosis, especially in the absence of FDG-PET avid lesions when discontinuing treatment. These data support the option of early discontinuation, limiting possible overtreatment and thereby toxicity, health and economic expenses and improving logistics.

Transcriptomic signatures of tumors undergoing T cell attack.

BACKGROUND: Studying tumor cell-T cell interactions in the tumor microenvironment (TME) can elucidate tumor immune escape mechanisms and help predict responses to cancer immunotherapy. METHODS: We selected 14 pairs of highly tumor-reactive tumor-infiltrating lymphocytes (TILs) and autologous short-term cultured cell lines, covering four distinct tumor types, and co-cultured TILs and tumors at sub-lethal ratios in vitro to mimic the interactions occurring in the TME. We extracted gene signatures associated with a tumor-directed T cell attack based on transcriptomic data of tumor cells. RESULTS: An autologous T cell attack induced pronounced transcriptomic changes in the attacked tumor cells, partially independent of IFN-γ signaling. Transcriptomic changes were mostly independent of the tumor histological type and allowed identifying common gene expression changes, including a shared gene set of 55 transcripts influenced by T cell recognition (Tumors undergoing T cell attack, or TuTack, focused gene set). TuTack scores, calculated from tumor biopsies, predicted the clinical outcome after anti-PD-1/anti-PD-L1 therapy in multiple tumor histologies. Notably, the TuTack scores did not correlate to the tumor mutational burden, indicating that these two biomarkers measure distinct biological phenomena. CONCLUSIONS: The TuTack scores measure the effects on tumor cells of an anti-tumor immune response and represent a comprehensive method to identify immunologically responsive tumors. Our findings suggest that TuTack may allow patient selection in immunotherapy clinical trials and warrant its application in multimodal biomarker strategies.

Epigenetic therapy in combination with a multi-epitope cancer vaccine targeting shared tumor antigens for high-risk myelodysplastic syndrome - a phase I clinical trial.

BACKGROUND: Standard care for patients with high-risk myelodysplastic syndrome (MDS) is hypomethylating agents such as azacitidine (AZA), which can induce expression of methylated tumor-associated antigens and therefore potentiate immunotherapeutic targeting. METHOD: In this phase 1 trial, we combined AZA with a therapeutic peptide vaccine targeting antigens encoded from NY-ESO-1, MAGE-A3, PRAME, and WT-1, which have previously been demonstrated to be upregulated by AZA treatment. RESULT: Five patients who had responded to AZA monotherapy were included in the study and treated with the vaccine. The combination therapy showed only few adverse events during the study period, whereof none classified as serious. However, no specific immune responses could be detected using intracellular cytokine staining or ELISpot assays. Minor changes in the phenotypic composition of immune cells and their expression of stimulatory and inhibitory markers were detected. All patients progressed to AML with a mean time to progression from inclusion (TTP) of 5.2 months (range 2.8 to 7.6). Mean survival was 18.1 months (range 10.9 to 30.6) from MDS diagnosis and 11.3 months (range 4.3 to 22.2) from inclusion. Sequencing of bone marrow showed clonal expansion of malignant cells, as well as appearance of novel mutations. CONCLUSION: The patients progressed to AML with an average time of only five months after initiating the combination therapy. This may be unrelated to the experimental treatment, but the trial was terminated early as there was no sign of clinical benefit or immunological response. Why the manuscript is especially interesting This study is the first to exploit the potential synergistic effects of combining a multi-peptide cancer vaccine with epigenetic therapy in MDS. Although our results are negative, they emphasize challenges to induce immune reactivity in patients with high-risk MDS.

Perspectives in Immunotherapy: meeting report from the Immunotherapy Bridge, December 1st-2nd, 2021.

Over the past decade, immunotherapy has become an increasingly fundamental modality in the treatment of cancer. The positive impact of immune checkpoint inhibition, especially anti-programmed death (PD)-1/PD-ligand (L)1 blockade, in patients with different cancers has focused attention on the potential for other immunotherapeutic approaches. These include inhibitors of additional immune checkpoints, adoptive cell transfer (ACT), and therapeutic vaccines. Patients with advanced cancers who previously had limited treatment options available may now benefit from immunotherapies that can offer durable responses and improved survival outcomes. However, despite this, a significant proportion of patients fail to respond to immunotherapy, especially those with less immunoresponsive cancer types, and there remains a need for new treatment strategies.The virtual Immunotherapy Bridge (December 1st-2nd, 2021), organized by the Fondazione Melanoma Onlus, Naples, Italy in collaboration with the Society for Immunotherapy of Cancer addressed several areas of current research in immunotherapy, including lessons learned from cell therapies, drivers of immune response, and trends in immunotherapy across different cancers, and these are summarised here.

Cardiotoxicity in metastatic melanoma patients treated with BRAF and MEK inhibitors in a real-world setting.

BACKGROUND: Combination therapy with BRAF and MEK inhibitors (BRAF/MEKi) has significantly improved the outcome for patients with BRAF-mutated melanoma. A reduction in left ventricular ejection fraction (LVEF) is a known side effect during treatment with BRAF/MEKi. This study aimed to analyze sequential multigated acquisition (MUGA) scans for the evaluation of LVEF and provide real-world data on cardiotoxicity induced by BRAF/MEKi in advanced melanoma. METHODS: All patients with advanced melanoma treated with dabrafenib and trametinib at Herlev and Gentofte Hospital, Denmark, between March 2015 and September 2019, were included retrospectively. MUGA scans performed at baseline and every three months during treatment were analyzed. Cardiotoxicity was defined as a decline of ≥10 percentage point (pp) to an LVEF <50% (major cardiotoxicity) or a decline in LVEF of ≥15 pp but remaining >50% (minor cardiotoxicity). RESULTS: A total of 139 patients were included. Forty-six patients (33%) met our criteria for cardiotoxicity; 31 patients (22%) experienced minor cardiotoxicity and 15 patients (11%) experienced major cardiotoxicity. Median time to decline in LVEF was 94 days, and all clinically significant declines in LVEF occurred before evaluation at six months. Reversibility of LVEF was seen in 80% of patients, three patients were not evaluable for reversibility. A low left ventricular peak emptying rate adjusted for heart rate (LVPERadj) at baseline was found a potential risk factor for the development of major cardiotoxicity (RR = 0.159, p = 0.001). CONCLUSION: A decline in LVEF is common for patients with advanced melanoma treated with BRAF/MEKi but rarely clinically significant. No significant decline in LVEF was observed after evaluation at six months, therefore routine monitoring of LVEF might be stopped after six to nine months of BRAF/MEKi therapy. A low LVPERadj might be a risk factor for the development of cardiotoxicity and is suggested for further investigation.

The risk of cardiac events in patients receiving immune checkpoint inhibitors: a nationwide Danish study.

AIMS: The study aimed to estimate the risk of cardiac events in immune checkpoint inhibitor (ICI)-treated patients with lung cancer or malignant melanoma. METHODS AND RESULTS: The study included consecutive patients with lung cancer or malignant melanoma in 2011-17 nationwide in Denmark. The main composite outcome was cardiac events (arrhythmia, peri- or myocarditis, heart failure) or cardiovascular death. Absolute risks were estimated and the association of ICI and cardiac events was analysed in multivariable Cox models. We included 25 573 patients with lung cancer. Of these, 743 were treated with programmed cell death-1 inhibitor (PD1i) and their 1-year absolute risk of cardiac events was 9.7% [95% confidence interval (CI) 6.8-12.5]. Of the 13 568 patients with malignant melanoma, 145 had PD1i and 212 had cytotoxic T-lymphocyte-associated protein-4 inhibitor (CTLA-4i) treatment. Their 1-year risks were 6.6% (1.8-11.3) and 7.5% (3.7-11.3). The hazard rates of cardiac events were higher in patients with vs. without ICI treatment. Within 6 months from 1st ICI administration, the hazard ratios were 2.14 (95% CI 1.50-3.05) in patients with lung cancer and 4.30 (1.38-13.42) and 4.93 (2.45-9.94) in patients with malignant melanoma with PD1i and CTLA-4i, respectively. After 6 months, HRs were 2.26 (1.27-4.02) for patients with lung cancer and 3.48 (1.91-6.35) for patients with malignant melanoma and CTLA-4i. CONCLUSIONS: Among patients with lung cancer and malignant melanoma, ICI treated had increased rates of cardiac events. The absolute risks were higher in these data compared with previous pharmacovigilance studies (e.g. 1.8% peri-/myocarditis 1-year risk).

The effects of targeted immune-regulatory strategies on tumor-specific T-cell responses in vitro.

BACKGROUND: Immune-related adverse events (IrAEs) are auto-immune reactions associated with immune checkpoint inhibitor-based therapy (ICI). Steroids are currently the first-line option for irAE management; however, recent studies have raised concerns regarding their potential impairment of tumor-specific immune responses. In this study, we investigated the in vitro effects of commonly used irAE treatment drugs on the anti-tumor activity of tumor-infiltrating lymphocytes (TILs). METHODS: Impairment of anti-tumor immune responses by four drugs (antibodies: vedolizumab and tocilizumab; small molecules: mycophenolate mofetil and tacrolimus) reported to be effective in treating irAEs was tested at clinically relevant doses in vitro and compared to a standard moderate dose of corticosteroids (small molecules) or infliximab (antibodies). TIL responses against autologous tumor cell lines, in the presence or absence of irAE drugs, were determined by flow cytometry (short-term tumor-specific T-cell activation) or xCELLigence (T-cell-mediated tumor killing). RESULTS: None of the tested antibodies influenced T-cell activation or T-cell-mediated tumor killing. Low-dose mycophenolate and tacrolimus did not influence T-cell activation, whereas higher doses of tacrolimus (> 1 ng/ml) impaired T-cell activation comparably to dexamethasone. All tested small molecules impaired T-cell-mediated tumor killing, with high-dose tacrolimus reducing killing at levels comparable to dexamethasone-mediated inhibition. In addition, mycophenolate and tacrolimus alone also demonstrated anti-proliferative effects on tumor cells. CONCLUSIONS: These data support clinical testing of targeted immune-regulatory strategies in the initial phase of irAE management, as a potential replacement for corticosteroids.