Deciphering the role of alternative splicing in neoplastic diseases for immune-oncological therapies.

Alternative splicing (AS) is an important molecular biological mechanism regulated by complex mechanisms involving a plethora of cis and trans-acting elements. Furthermore, AS is tissue specific and altered in various pathologies, including infectious, inflammatory, and neoplastic diseases. Recently developed immuno-oncological therapies include monoclonal antibodies (mAbs) and chimeric antigen receptor (CAR) T cells targeting, among others, immune checkpoint (ICP) molecules. Despite therapeutic successes have been demonstrated, only a limited number of patients showed long-term benefit from these therapies with tumor entity-related differential response rates were observed. Interestingly, splice variants of common immunotherapeutic targets generated by AS are able to completely escape and/or reduce the efficacy of mAb- and/or CAR-based tumor immunotherapies. Therefore, the analyses of splicing patterns of targeted molecules in tumor specimens prior to therapy might help correct stratification, thereby increasing therapy success by antibody panel selection and antibody dosages. In addition, the expression of certain splicing factors has been linked with the patients' outcome, thereby highlighting their putative prognostic potential. Outstanding questions are addressed to translate the findings into clinical application. This review article provides an overview of the role of AS in (tumor) diseases, its molecular mechanisms, clinical relevance, and therapy response.

Role of HLA-G in tumors and upon COVID-19 infection.

HLA-G expression of tumors and upon viral infections is involved in their immune escape leading to the evasion from both T and NK cell recognition. The underlying mechanisms of HLA-G expression in both pathophysiologic conditions are broad and range from genetic abnormalities to epigenetic, transcriptional and posttranscriptional regulation. This review summarizes the current knowledge of the frequency, regulation and clinical relevance of HLA-G expression upon neoplastic and viral transformation, its interaction with components of the microenvironment as well as its potential as diagnostic marker and/or therapeutic target. In addition, it discusses urgent topics, which have to be addressed in HLA-G research.

Prognostic impact of the bone marrow tumor microenvironment, HLA-I and HLA-Ib expression in MDS and CMML progression to sAML.

Genetic aberrations and immune escape are fundamental in MDS and CMML initiation and progression to sAML. Therefore, quantitative and spatial immune cell organization, expression of immune checkpoints (ICP), classical human leukocyte antigen class I (HLA-I) and the non-classical HLA-Ib antigens were analyzed in 274 neoplastic and 50 non-neoplastic bone marrow (BM) biopsies using conventional and multiplex immunohistochemistry and correlated to publicly available dataset. Higher numbers of tissue infiltrating lymphocytes (TILs) were found in MDS/CMML (8.8%) compared to sAML (7.5%) and non-neoplastic BM (5.3%). Higher T cell abundance, including the CD8+ T cell subset, inversely correlated with the number of pathogenic mutations and was associated with blast BM counts, ICP expression, spatial T cell distribution and improved patients' survival in MDS and CMML. In MDS/CMML, higher PD-1/PD-L1/PD-L2 and HLA-I, but lower HLA-G expression correlated with a significantly better patients' outcome. Moreover, a closer spatial proximity of T cell subpopulations and their proximity to myeloid blasts showed a stronger prognostic impact when compared to TIL numbers. In sAML - the continuum of MDS and CMML - the number of TILs had no impact on prognosis, but higher CD28 and HLA-I expression correlated with a better outcome of sAML patients. This study underlines the independent prognostic value of the tumor microenvironment in MDS/CMML progression to sAML, which shows the most pronounced immune escape. Moreover, new prognostic markers, like HLA-G expression and spatial T cell distribution, were described for the first time, which might also serve as therapeutic targets.

Correlation of Increased Soluble Tumor Necrosis Factor Receptor 1 with Mortality and Dependence on Treatment in Non-Small-Cell Lung Cancer Patients: A Longitudinal Cohort Study.

BACKGROUND: Tumor necrosis factor (TNF) is a multipotent cytokine involved in inflammation and anti-tumor activity. TNF-α exerts its function upon binding to TNF-receptor 1 (TNF-R1) and TNF-receptor 2 (TNF-R2). This study investigates the relationship of soluble (s) TNF-R1 levels in non-small-cell lung cancer (NSCLC) patients with treatment and overall survival. METHODS: In total, 134 NSCLC patients treated at the Medical Faculty of Martin Luther University Halle-Wittenberg between 2017 and 2019 were included in this study. Serum levels of sTNF-R1 were measured via ELISA at baseline and during and after treatment. A linear mixed-effects model was used to assess sTNF-R1 changes over time. Linear regression was applied to investigate the association between clinical characteristics and changes in sTNF-R1. Cox regression models were used to estimate associations with overall mortality. RESULTS: The estimated average sTNFR-1 at baseline was 2091.71 pg/mL, with a change of 6.19 pg/mL per day. Cox models revealed that the individual change in sTNF-R1 was more strongly associated with mortality than its baseline value, especially after adjusting for covariates. CONCLUSIONS: This study provides evidence that the individual change in sTNF-R1 levels during and after treatment were associated with the risk of mortality, suggesting the use of the sTNF-R1 trajectory as a prognostic marker.

BRAF and MEK inhibitor combinations induce potent molecular and immunological effects in NRAS-mutant melanoma cells: Insights into mode of action and resistance mechanisms.

About 25% of melanoma harbor activating NRAS mutations, which are associated with aggressive disease therefore requiring a rapid antitumor intervention. However, no efficient targeted therapy options are currently available for patients with NRAS-mutant melanoma. MEK inhibitors (MEKi) appear to display a moderate antitumor activity and also immunological effects in NRAS-mutant melanoma, providing an ideal backbone for combination treatments. In our study, the MEKi binimetinib, cobimetinib and trametinib combined with the BRAF inhibitors (BRAFi) encorafenib, vemurafenib and dabrafenib were investigated for their ability to inhibit proliferation, induce apoptosis and alter the expression of immune modulatory molecules in sensitive NRAS-mutant melanoma cells using two- and three-dimensional cell culture models as well as RNA sequencing analyses. Furthermore, NRAS-mutant melanoma cells resistant to the three BRAFi/MEKi combinations were established to characterize the mechanisms contributing to their resistance. All BRAFi induced a stress response in the sensitive NRAS-mutant melanoma cells thereby significantly enhancing the antiproliferative and proapoptotic activity of the MEKi analyzed. Furthermore, BRAFi/MEKi combinations upregulated immune relevant molecules, such as ICOS-L, components of antigen-presenting machinery and the "don't eat me signal" molecule CD47 in the melanoma cells. The BRAFi/MEKi-resistant, NRAS-mutant melanoma cells counteracted the molecular and immunological effects of BRAFi/MEKi by upregulating downstream mitogen-activated protein kinase pathway molecules, inhibiting apoptosis and promoting immune escape mechanisms. Together, our study reveals potent molecular and immunological effects of BRAFi/MEKi in sensitive NRAS-mutant melanoma cells that may be exploited in new combinational treatment strategies for patients with NRAS-mutant melanoma.

Identification and characterization of the anti-viral interferon lambda 3 as direct target of the Epstein-Barr virus microRNA-BART7-3p.

The human Epstein-Barr virus (EBV), as a member of the human γ herpes viruses (HHV), is known to be linked with distinct tumor types. It is a double-stranded DNA virus and its genome encodes among others for 48 different microRNAs (miRs). Current research demonstrated a strong involvement of certain EBV-miRs in molecular immune evasion mechanisms of infected cells by, e.g., the disruption of human leukocyte antigen (HLA) class Ia and NKG2D functions. To determine novel targets of EBV-miRs involved in immune surveillance, ebv-miR-BART7-3p, an EBV-encoded miR with high expression levels during the different lytic and latent EBV life cycle phases, was overexpressed in human HEK293T cells. Using a cDNA microarray-based comparative analysis, 234 (229 downregulated and 5 upregulated) deregulated human transcripts were identified in ebv-miR-BART7-3p transfectants, which were mainly involved in cellular processes and molecular binding. A statistically significant downregulation of the anti-proliferative and tumor-suppressive hsa-miR-34A and the anti-viral interferon lambda (IFNL)3 mRNA was found. The ebv-miR-BART7-3p-mediated downregulation of IFNL3 expression was due to a direct interaction with the IFNL3 3'-untranslated region (UTR) as determined by luciferase reporter gene assays including the identification of the accurate ebv-miR-BART7-3p binding site. The effect of ebv-miR-BART7-3p on the IFNL3 expression was validated both in human cell lines in vitro and in human tissue specimen with known EBV status. These results expand the current knowledge of EBV-encoded miRs and their role in immune evasion, pathogenesis and malignant transformation.

Comparative Immune Response after Vaccination with SOBERANA® 02 and SOBERANA® plus Heterologous Scheme and Natural Infection in Young Children.

(1) Background: In children, SARS-CoV-2 infection is mostly accompanied by mild COVID-19 symptoms. However, multisystem inflammatory syndrome (MIS-C) and long-term sequelae are often severe complications. Therefore, the protection of the pediatric population against SARS-CoV-2 with effective vaccines is particularly important. Here, we compare the humoral and cellular immune responses elicited in children (n = 15, aged 5-11 years) vaccinated with the RBD-based vaccines SOBERANA® 02 and SOBERANA® Plus combined in a heterologous scheme with those from children (n = 10, aged 4-11 years) who recovered from mild symptomatic COVID-19. (2) Methods: Blood samples were taken 14 days after the last dose for vaccinated children and 45-60 days after the infection diagnosis for COVID-19 recovered children. Anti-RBD IgG and ACE2-RBD inhibition were assessed by ELISA; IgA, cytokines, and cytotoxic-related proteins were determined by multiplex assays. Total B and T cell subpopulations and IFN-γ release were measured by multiparametric flow cytometry using a large panel of antibodies after in vitro stimulation with S1 peptides. (3) Results: Significant higher levels of specific anti-RBD IgG and IgA and ACE2-RBD inhibition capacity were found in vaccinated children in comparison to COVID-19 recovered children. Th1-like and Th2-like CD4+ T cells were also significantly higher in vaccinated subjects. IFN-γ secretion was higher in central memory CD4+ T cells of COVID-19 recovered children, but no differences between both groups were found in the CD4+ and CD8+ T cell effector, terminal effector, and naïve T cell subpopulations. In contrast to low levels of IL-4, high levels of IL-2, IL-6, IFN-γ, and IL-10 suggest a predominant Th1 cell polarization. Cytotoxic-related proteins granzyme A and B, perforin, and granulin were also found in the supernatant after S1 stimulation in both vaccinated and recovered children. (4) Conclusions: Vaccination with the heterologous scheme of SOBERANA® 02/SOBERANA® Plus induces a stronger antibody and cellular immune response compared to natural infections in young children.

Blood Immune Cells as Biomarkers in Long-Term Surviving Patients with Advanced Non-Small-Cell Lung Cancer Undergoing a Combined Immune/Chemotherapy.

An important challenge remains in identifying the baseline characteristics of cancer patients who will mostly benefit from immune checkpoint inhibitor (ICI) therapies. Furthermore, biomarkers could help in the choice of an optimal therapy duration after a primary therapy response. In this pilot study, the time courses of four different immune cell parameters were followed in 12 patients with advanced non-small-cell lung cancer (NSCLC) undergoing ICI therapy combined with chemotherapy and surviving at least 12 months. Blood was collected at the time point of the first and third antibody administration, as well as after 12 months of patients' survival. Using multi-color flow cytometry, two suppressive markers (neutrophil/lymphocyte ratio (NLR) and the frequency of circulating HLA-DRlow monocytes), as well as two markers of an ongoing immune response (6-Sulfo LacNAc (slan)+ non-classical monocytes and dendritic cell (DC) subtypes), were determined. In most of those who survived > 12 months, a low NLR and a low number of HLA-DRlow monocytes combined with clearly detectable numbers of slan+ non-classical monocytes and of DC subtypes were seen. Two of the patients had an increase in the suppressive markers paired with a decrease in slan+ non-classical monocytes and in DC subtypes, which, in at least one patient, was the correlate of an ongoing clinical progression. Our results implicate that the NLR, specific subtypes of monocytes, and the number of blood DCs might be useful predictive biomarkers for cancer patients during long-term treatment with ICI/chemotherapy.

Tumor immunology meets oncology (TIMO) XVII, April 20-22 2023 in Halle/Saale, Germany.

During the TIMO meeting 2023, national and international scientists as well as clinicians gave novel insights as well as perspectives into basic and translational tumor immunology. https://dgfi.org/arbeitskreise/ak-tumorimmunologie/meeting/.

Combination of multiple omics techniques for a personalized therapy or treatment selection.

Despite targeted therapies and immunotherapies have revolutionized the treatment of cancer patients, only a limited number of patients have long-term responses. Moreover, due to differences within cancer patients in the tumor mutational burden, composition of the tumor microenvironment as well as of the peripheral immune system and microbiome, and in the development of immune escape mechanisms, there is no "one fit all" therapy. Thus, the treatment of patients must be personalized based on the specific molecular, immunologic and/or metabolic landscape of their tumor. In order to identify for each patient the best possible therapy, different approaches should be employed and combined. These include (i) the use of predictive biomarkers identified on large cohorts of patients with the same tumor type and (ii) the evaluation of the individual tumor with "omics"-based analyses as well as its ex vivo characterization for susceptibility to different therapies.

Correlation of the tumor escape phenotype with loss of PRELP expression in melanoma.

BACKGROUND: Despite immunotherapies having revolutionized the treatment of advanced cutaneous melanoma, effective and durable responses were only reported in a few patients. A better understanding of the interaction of melanoma cells with the microenvironment, including extracellular matrix (ECM) components, might provide novel therapeutic options. Although the ECM has been linked to several hallmarks of cancer, little information is available regarding the expression and function of the ECM protein purine-arginine-rich and leucine-rich protein (PRELP) in cancer, including melanoma. METHODS: The structural integrity, expression and function of PRELP, its correlation with the expression of immune modulatory molecules, immune cell infiltration and clinical parameters were determined using standard methods and/or bioinformatics. RESULTS: Bioinformatics analysis revealed a heterogeneous, but statistically significant reduced PRELP expression in available datasets of skin cutaneous melanoma when compared to adjacent normal tissues, which was associated with reduced patients' survival, low expression levels of components of the MHC class I antigen processing machinery (APM) and interferon (IFN)-γ signal transduction pathway, but increased expression of the transforming growth factor (TGF)-ß isoform 1 (TFGB1) and TGF-ß receptor 1 (TGFBR1). In addition, a high frequency of intra-tumoral T cells directly correlated with the expression of MHC class I and PRELP as well as the T cell attractant CCL5 in melanoma lesions. Marginal to low PRELP expression levels were found in the 47/49 human melanoma cell lines analysis. Transfection of PRELP into melanoma cell lines restored MHC class I surface expression due to transcriptional upregulation of major MHC class I APM and IFN-γ pathway components. In addition, PRELP overexpression is accompanied by high CCL5 secretion levels in cell supernatant, an impaired TGF-ß signaling as well as a reduced cell proliferation, migration and invasion of melanoma cells. CONCLUSIONS: Our findings suggest that PRELP induces the expression of MHC class I and CCL5 in melanoma, which might be involved in an enhanced T cell recruitment and immunogenicity associated with an improved patients' outcome. Therefore, PRELP might serve as a marker for predicting disease progression and its recovery could revert the tumorigenic phenotype, which represents a novel therapeutic option for melanoma.

Correlation between Circulating miR-16, miR-29a, miR-144 and miR-150, and the Radiotherapy Response and Survival of Non-Small-Cell Lung Cancer Patients.

Despite the success of current therapy concepts, patients with advanced non-small-cell lung cancer (NSCLC) still have a very poor prognosis. Therefore, biological markers are urgently needed, which allow the assessment of prognosis, or prediction of the success of therapy or resistance in this disease. Circulating microRNAs (miRs) have potential as biomarkers for the prognosis and prediction of response to therapy in cancer patients. Based on recent evidence that circulating miR-16, miR-29a, miR-144 and miR-150 can be regulated by ionizing radiation, the concentration of these four miRs was assessed in the plasma of NSCLC patients at different time points of radiotherapy by digital droplet PCR (ddPCR). Furthermore, their impact on patients' prognosis was evaluated. The mean plasma levels of miR-16, miR-29a, miR-144 and miR-150 significantly differed intra- and inter-individually, and during therapy in NSCLC patients, but showed a strong positive correlation. The individual plasma levels of miR-16, miR-29a and miR-144 had prognostic value in NSCLC patients during or at the end of radiotherapy in Cox's regression models. NSCLC patients with low levels of these three miRs at the end of radiotherapy had the worst prognosis. However, miR-150 plasma levels and treatment-dependent changes were not predictive. In conclusion, circulating miR-16, miR-29a and miR-144, but not miR-150, have a prognostic value in NSCLC patients undergoing radiotherapy.

A novel ACE2 decoy for both neutralization of SARS-CoV-2 variants and killing of infected cells.

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) led to millions of infections and deaths worldwide. As this virus evolves rapidly, there is a high need for treatment options that can win the race against new emerging variants of concern. Here, we describe a novel immunotherapeutic drug based on the SARS-CoV-2 entry receptor ACE2 and provide experimental evidence that it cannot only be used for (i) neutralization of SARS-CoV-2 in vitro and in SARS-CoV-2-infected animal models but also for (ii) clearance of virus-infected cells. For the latter purpose, we equipped the ACE2 decoy with an epitope tag. Thereby, we converted it to an adapter molecule, which we successfully applied in the modular platforms UniMAB and UniCAR for retargeting of either unmodified or universal chimeric antigen receptor-modified immune effector cells. Our results pave the way for a clinical application of this novel ACE2 decoy, which will clearly improve COVID-19 treatment.

Tumor immunology meets oncology (TIMO) XVI, July 07-09 2022 in Halle/Saale, Germany.

During the TIMO meeting 2022, national and international scientists as well as clinicians gave novel insights as well as perspectives into basic and translational tumor immunology.

Interferons and Resistance Mechanisms in Tumors and Pathogen-Driven Diseases-Focus on the Major Histocompatibility Complex (MHC) Antigen Processing Pathway.

Interferons (IFNs), divided into type I, type II, and type III IFNs represent proteins that are secreted from cells in response to various stimuli and provide important information for understanding the evolution, structure, and function of the immune system, as well as the signaling pathways of other cytokines and their receptors. They exert comparable, but also distinct physiologic and pathophysiologic activities accompanied by pleiotropic effects, such as the modulation of host responses against bacterial and viral infections, tumor surveillance, innate and adaptive immune responses. IFNs were the first cytokines used for the treatment of tumor patients including hairy leukemia, renal cell carcinoma, and melanoma. However, tumor cells often develop a transient or permanent resistance to IFNs, which has been linked to the escape of tumor cells and unresponsiveness to immunotherapies. In addition, loss-of-function mutations in IFN signaling components have been associated with susceptibility to infectious diseases, such as COVID-19 and mycobacterial infections. In this review, we summarize general features of the three IFN families and their function, the expression and activity of the different IFN signal transduction pathways, and their role in tumor immune evasion and pathogen clearance, with links to alterations in the major histocompatibility complex (MHC) class I and II antigen processing machinery (APM). In addition, we discuss insights regarding the clinical applications of IFNs alone or in combination with other therapeutic options including immunotherapies as well as strategies reversing the deficient IFN signaling. Therefore, this review provides an overview on the function and clinical relevance of the different IFN family members, with a specific focus on the MHC pathways in cancers and infections and their contribution to immune escape of tumors.

Regional Variation in the Tumor Microenvironment, Immune Escape and Prognostic Factors in Breast Cancer in Sub-Saharan Africa.

The low overall survival rates of patients with breast cancer in sub-Saharan Africa (SSA) are driven by regionally differing tumor biology, advanced tumor stages at diagnosis, and limited access to therapy. However, it is not known whether regional differences in the composition of the tumor microenvironment (TME) exist and affect patients' prognosis. In this international, multicentre cohort study, 1,237 formalin-fixed, paraffin-embedded breast cancer samples, including samples of the "African Breast Cancer-Disparities in Outcomes (ABC-DO) Study," were analyzed. The immune cell phenotypes, their spatial distribution in the TME, and immune escape mechanisms of breast cancer samples from SSA and Germany (n = 117) were investigated using histomorphology, conventional and multiplex IHC, and RNA expression analysis. The data revealed no regional differences in the number of tumor-infiltrating lymphocytes (TIL) in the 1,237 SSA breast cancer samples, while the distribution of TILs in different breast cancer IHC subtypes showed regional diversity, particularly when compared with German samples. Higher TIL densities were associated with better survival in the SSA cohort (n = 400), but regional differences concerning the predictive value of TILs existed. High numbers of CD163+ macrophages and CD3+CD8+ T cells accompanied by reduced cytotoxicity, altered IL10 and IFNγ levels and downregulation of MHC class I components were predominantly detected in breast cancer samples from Western SSA. Features of nonimmunogenic breast cancer phenotypes were associated with reduced patient survival (n = 131). We therefore conclude that regional diversity in the distribution of breast cancer subtypes, TME composition, and immune escape mechanisms should be considered for therapy decisions in SSA and the design of personalized therapies. See related Spotlight by Bergin et al., p. 705.

Monitoring Blood Immune Cells in Patients with Advanced Small Cell Lung Cancer Undergoing a Combined Immune Checkpoint Inhibitor/Chemotherapy.

In this exploratory prospective observational study on 40 small cell lung cancer (SCLC) patients treated with a combination of chemotherapy and immune checkpoint inhibitors, blood immune cells were characterized by multi-color flow cytometry at the baseline and at the third therapy cycle. The numbers of neutrophils and of T-, B-, and NK cells, as well as the frequency of HLA-DRlow monocytes, 6-SulfoLacNAc (slan)+ non-classical monocytes and circulating dendritic cell (DC) subtypes were determined. The prognostic value of the parameters was evaluated by the patient's survival analysis with overall survival (OS) as the primary endpoint. In addition, blood cell parameters from SCLC patients were compared to those from non-SCLC (NSCLC). The global median OS of patients was 10.4 ± 1.1 months. Disease progression (15% of patients) correlated with a higher baseline neutrophil/lymphocyte ratio (NLR), more HLA-DRlow monocytes, and lower NK cell and DC numbers. The risk factors for poor OS were the presence of brain/liver metastases, a baseline NLR ≥ 6.1, HLA-DRlow monocytes ≥ 21% of monocytes, slan+ non-classical monocytes < 0.12%, and/or CD1c+ myeloid DC < 0.05% of leukocytes. Lymphocytic subpopulations did not correlate with OS. When comparing biomarkers in SCLC versus NSCLC, SCLC had a higher frequency of brain/liver metastases, a higher NLR, the lowest DC frequencies, and lower NK cell numbers. Brain/liver metastases had a substantial impact on the survival of SCLC patients. At the baseline, 45% of SCLC patients, but only 24% of NSCLC patients, had between three and five risk factors. A high basal NLR, a high frequency of HLA-DRlow monocytes, and low levels of slan+ non-classical monocytes were associated with poor survival in all lung cancer histotypes. Thus, the blood immune cell signature might contribute to a better prediction of SCLC patient outcomes and may uncover the pathophysiological peculiarities of this tumor entity.

MicroRNAs affecting the susceptibility of melanoma cells to CD8+ T cell-mediated cytolysis.

BACKGROUND: The regulatory functions of microRNAs (miRNAs) in anti-tumour immunity have been mainly described in immune effector cells. Since little is known about miRNA effects on the susceptibility of target cells during T cell-target cell interaction, this study focused on the identification of miRNAs expressed in tumour cells controlling their susceptibility to CD8+ T cell-mediated cytotoxicity. METHODS: Luciferase expressing B16F10 melanoma (B16F10 Luci+ ) cells transfected with individual miRNAs covering a comprehensive murine miRNA library were screened for their susceptibility to lysis by an established cytotoxic T lymphocyte (CTL) line (5a, clone Nß) specific for the melanoma-associated antigen tyrosinase-related protein 2. miRNAs with the most pronounced effects on T cell-mediated lysis were validated and stably expressed in B16F10 cells. In silico analyses identified common targets of miRNA sets determined by the screen, which were further confirmed by small interfering RNA (siRNA)-mediated silencing experiments modulating immune surveillance. The Ingenuity Pathway Analysis (IPA) software and RNA sequencing (RNA-seq) data from miRNA-overexpressing cell lines were applied to investigate the underlying mechanisms. The Cancer Genome Atlas (TCGA)-derived miRNA sequencing data were used to assess the correlation of miRNA expression with melanoma patients' survival. RESULTS: The miRNA screen resulted in the selection of seven miRNAs enhancing CTL-mediated melanoma cell killing in vitro. Upon stable overexpression of selected miRNAs, hsa-miR-320a-3p, mmu-miR-7037-5p and mmu-miR-666-3p were determined as most effective in enhancing susceptibility to CTL lysis. In silico analyses and subsequent siRNA-mediated silencing experiments identified Psmc3 and Ndufa1 as common miRNA targets possibly involved in the functional effects observed. The analyses of RNA-seq data with IPA showed pathways, networks, biological functions and key molecules potentially involved in the miRNA-mediated functional effects. Finally, based on TCGA data analysis, a positive correlation of the conserved miRNAs among the panel of the seven identified miRNAs with overall survival of melanoma patients was determined. CONCLUSIONS: For the first time, this study uncovered miRNA species that affect the susceptibility of melanoma cells to T cell-mediated killing. These miRNAs might represent attractive candidates for novel therapy approaches against melanoma and other tumour entities.