Project description:Next Generation Sequencing (NGS) using capture or amplicons strategies allows the detection of a large number of mutations increasing the rate of positive diagnosis for the patients. However, most of the detected mutations are Single Nucleotide Variants (SNVs) or small indels. Structural Variants (SVs) are often underdiagnosed in inherited genetic diseases, probably because few user-friendly tools are available for biologists or geneticists to identify them easily. We present here the diagnosis of two brothers presenting a demyelinating motor-sensitive neuropathy: a presumed homozygous c.5744_5745delAT in exon 10 of SACS gene was initially detected, while actually these patients were heterozygous for this mutation and harbored a large deletion of SACS exon 10 in the other allele. This hidden mutation has been detected thanks to the user-friendly CovCopCan software. We recommend to systematically use such a software to screen NGS data in order to detect SVs, such as Copy Number Variations, to improve diagnosis of the patients.
Project description:Oncolytic viruses, including oncolytic herpes simplex virus (oHSV), have produced provocative therapeutic responses in patients with glioblastoma, the most aggressive brain tumor. Paradoxically, innate immune responses mediated by natural killer (NK) cells and macrophages/microglia appear to limit oHSV efficacy. Therefore, we investigated whether pretreatment with an immunosuppressive cytokine, TGFβ, might reverse these effects and thereby potentiate oHSV efficacy. TGFβ treatment of NK cells rendered them less cytolytic against oHSV-infected glioblastoma cells and stem-like cells in vitro. Furthermore, TGFβ treatment of NK cells, macrophages, or microglia increased viral titers of oHSV in cocultures with glioblastoma cells. In a syngeneic mouse model of glioblastoma, administering TGFβ prior to oHSV injection inhibited intracranial infiltration and activation of NK cells and macrophages. Notably, a single administration of TGFβ prior to oHSV therapy was sufficient to phenocopy NK-cell depletion and suppress tumor growth and prolong survival in both xenograft and syngeneic models of glioblastoma. Collectively, our findings show how administering a single dose of TGFβ prior to oncolytic virus treatment of glioblastoma can transiently inhibit innate immune cells that limit efficacy, thereby improving therapeutic responses and survival outcomes.
Project description:Despite expression of immunogenic polypeptides, tumors escape immune surveillance by engaging T cell checkpoint regulators and expanding Tregs, among other mechanisms. What orchestrates these controls is unknown. We report that free C3d, a fragment of the third component of complement, inside tumor cells - or associated with irradiated tumor cells and unattached to antigen - recruits, accelerates, and amplifies antitumor T cell responses, allowing immunity to reverse or even to prevent tumor growth. C3d enhances antitumor immunity independently of B cells, NK cells, or antibodies, but it does so by increasing tumor infiltrating CD8+ lymphocytes, by depleting Tregs, and by suppressing expression of programmed cell death protein 1 (PD-1) by T cells. These properties of C3d appear specific for the tumor and dependent on complement receptor 2, and they incur no obvious systemic toxicity. The heretofore unrecognized properties of free C3d suggest that protein might determine the effectiveness of immune surveillance and that increasing availability of the protein might prove advantageous in the treatment or prevention of cancer and premalignant conditions.
Project description:Microbes and their byproducts have been reported to regulate host health and immune functions. Here we demonstrated that microbial exopolysaccharide produced by Lactobacillus delbrueckii subsp. bulgaricus OLL1073R-1 (EPS-R1) induced CCR6+ CD8+ T cells of mice and humans. In mice, ingestion of EPS-R1 augmented antitumor effects of anti-CTLA-4 or anti-PD-1 monoclonal antibody against CCL20-expressing tumors, in which infiltrating CCR6+ CD8+ T cells were increased and produced IFNγ accompanied by a substantial immune response gene expression signature maintaining T-cell functions. Of note, the antitumor adjuvant effect of EPS-R1 was also observed in germ-free mice. Furthermore, the induction of CCR6 expression was mediated through the phosphorylated structure in EPS-R1 and a lysophosphatidic acid receptor on CD8+ T cells. Overall, we find that dietary EPS-R1 consumption induces CCR6+ CD8+ T cells in Peyer's patches, favoring a tumor microenvironment that augments the therapeutic effect of immune-checkpoint blockade depending on CCL20 production by tumors.SignificanceGut microbiota- and probiotic-derived metabolites are attractive agents to augment the efficacy of immunotherapies. Here we demonstrated that dietary consumption of Lactobacillus-derived exopolysaccharide induced CCR6+ CD8+ T cells in Peyer's patches and improved the tumor microenvironment to augment the therapeutic effects of immune-checkpoint blockade against CCL20-producing tumors. See related commentary by Di Luccia and Colonna, p. 1189. This article is highlighted in the In This Issue feature, p. 1171.
Project description:PurposePD-1/PD-L1 signaling promotes tumor growth while inhibiting effector cell-mediated antitumor immune responses. Here, we assessed the impact of single and dual blockade of PD-1/PD-L1, alone or in combination with lenalidomide, on accessory and immune cell function as well as multiple myeloma cell growth in the bone marrow (BM) milieu.Experimental designSurface expression of PD-1 on immune effector cells, and PD-L1 expression on CD138(+) multiple myeloma cells and myeloid-derived suppressor cells (MDSC) were determined in BM from newly diagnosed (ND) multiple myeloma and relapsed/refractory (RR) multiple myeloma versus healthy donor (HD). We defined the impact of single and dual blockade of PD-1/PD-L1, alone and with lenalidomide, on autologous anti-multiple myeloma immune response and tumor cell growth.ResultsBoth ND and RR patient multiple myeloma cells have increased PD-L1 mRNA and surface expression compared with HD. There is also a significant increase in PD-1 expression on effector cells in multiple myeloma. Importantly, PD-1/PD-L1 blockade abrogates BM stromal cell (BMSC)-induced multiple myeloma growth, and combined blockade of PD-1/PD-L1 with lenalidomide further inhibits BMSC-induced tumor growth. These effects are associated with induction of intracellular expression of IFNγ and granzyme B in effector cells. Importantly, PD-L1 expression in multiple myeloma is higher on MDSC than on antigen-presenting cells, and PD-1/PD-L1 blockade inhibits MDSC-mediated multiple myeloma growth. Finally, lenalidomide with PD-1/PD-L1 blockade inhibits MDSC-mediated immune suppression.ConclusionsOur data therefore demonstrate that checkpoint signaling plays an important role in providing the tumor-promoting, immune-suppressive microenvironment in multiple myeloma, and that PD-1/PD-L1 blockade induces anti-multiple myeloma immune response that can be enhanced by lenalidomide, providing the framework for clinical evaluation of combination therapy.
Project description:Immune checkpoint blockade (ICB) therapy, particularly PD1/PDL1 inhibition, has demonstrated success in bolstering durable responses in patients. However, the response rate remains below 30 %. In this study, we developed a polymeric bispecific antibody (BsAb) targeting PD1/PDL1 to enhance ICB therapy. Specifically, poly(L-glutamic acid) (PGLU) was conjugated with a double cyclic Fc binding peptide, Fc-III-4C, through condensation reactions between the -COOH group of PGLU and the -NH2 group of Fc-III-4C. This conjugate was then mixed with αPD1 and αPDL1 monoclonal antibodies (mAbs) in an aqueous solution. Mechanistically, the PD1/PDL1 BsAb (BsAbαPD1+αPDL1) acts as a bridge between tumor cells and CD8+ T cells, continuously activating CD8+ T cells to a greater extent. This leads to significantly suppressed tumor growth and prolonged survival in a mouse model of colon cancer compared to treatment with either a single mAb or a mixture of free mAbs. The tumor suppression rate achieved by the BsAbαPD1+αPDL1 was 90.1 %, with a corresponding survival rate of 83.3 % after 48 days. Thus, this study underscores the effectiveness of the BsAbαPD1+αPDL1 as a synchronizing T cell engager and dual ICBs, offering theoretical guidance for clinical ICB therapy.
Project description:BackgroundThe emergence of immunotherapy has revolutionized the paradigm of cancer treatment with immune checkpoint blockades (ICB) in solid cancers, including colorectal cancer (CRC). However, only a small subset of CRC patients harboring deficient mismatch repair (dMMR) or microsatellite instability-high (MSI-H) benefits from ICB therapy. A very limited response to ICB therapy has been achieved in MMR-proficient CRC, representing a significant challenge limiting the clinical application of immunotherapy. MMR is the critical DNA repair pathway that maintains genomic integrity by correcting DNA mismatches, which is mediated by the MutSα or MutSβ complex consisting of MSH2 with MSH6 and MSH3, respectively. Given that MMR status directs effective immune response, we sought to determine whether targeting MMR capacity boosts ICB efficacy.MethodsAzoxymethane/dextran sodium sulfate (AOM/DSS)-induced CRC and xenograft model were used to evaluate the function of PRMT6 and response to PRMT6 inhibitor EPZ020411 and combination therapy of PD1 and EPZ020411. Biochemical assays were performed to elucidate the underlying mechanism of PRMT6-mediated MSH2 methylation and immune evasion.ResultsWe have identified PRMT6 as a crucial regulator of MMR capacity via MSH2 dimethylation at R171 and R219. Such a modification abrogates its MMR capacity and prevents the recruitment of MSH3 and MSH6. PRMT6 loss or inhibition triggers cytosolic DNA accumulation and cGAS-STING signaling activation, leading to enhanced immune response in PRMT6-deficient colon tumors or xenografts. Pharmacological inhibition of PRMT6 using EPZ020411 promotes mutagenesis and destabilizes MutSα or MutSβ assembly, and prolonged EPZ020411 exposure maintains an MSI-like phenotype in microsatellite stability (MSS) cells. EPZ020411 treatment sensitizes ICB efficacy of MSS cells, but not MSI cells in vivo. Similar effects have been observed in MSS colon tumors induced by AOM/DSS.ConclusionsOur study provides a preclinical proof of concept to overcome resistance to immunotherapy by targeting PRMT6 in CRC with MSS.
Project description:BackgroundTransforming growth factor beta (TGFβ) is well-recognized as an immunosuppressive player in the tumor microenvironment but also has a significant impact on cancer cell phenotypes. Loss of TGFβ signaling impairs DNA repair competency, which is described by a transcriptomic score, βAlt. Cancers with high βAlt have more genomic damage and are more responsive to genotoxic therapy. The growing appreciation that cancer DNA repair deficits are important determinants of immune response prompted us to investigate βAlt's association with response to immune checkpoint blockade (ICB). We predicted that high βAlt tumors would be infiltrated with lymphocytes because of DNA damage burden and hence responsive to ICB.MethodsWe analyzed public transcriptomic data from clinical trials and preclinical models using transcriptomic signatures of TGFβ targets, DNA repair genes, tumor educated immune cells and interferon. A high βAlt, immune poor mammary tumor derived transplant model resistant to programmed death ligand 1 (PD-L1) antibodies was studied using multispectral flow cytometry to interrogate the immune system.ResultsMetastatic bladder patients in IMvigor 210 who responded to ICB had significantly increased βAlt scores and experienced significantly longer overall survival compared to those with low βAlt scores (hazard ratio 0.62, P=0.011). Unexpectedly, 75% of high βAlt cancers were immune poor as defined by low expression of tumor educated immune cell and interferon signatures. The association of high βAlt with immune poor cancer was also evident in TCGA and preclinical cancer models. We used a high βAlt, immune poor cancer to test therapeutic strategies to overcome its inherent anti-PD-L1 resistance. Combination treatment with radiation and TGFβ inhibition were necessary for lymphocytic infiltration and activated NK cells were required for ICB response. Bioinformatic analysis identified high βAlt, immune poor B16 and CT26 preclinical models and paired biopsies of cancer patients that also demonstrated NK cell activation upon response to ICB.ConclusionsOur studies confirm βAlt as a biomarker that predicts response to ICB in immune poor cancers., which has implications for the development of therapeutic strategies to increase the number of cancer patients who will benefit from immunotherapy.
Project description:The limited infiltration of CD8+ T cells in tumors hampers the effectiveness of T cell-based immunotherapy, yet the mechanisms that limit tumor infiltration by CD8+ T cells remain unclear. Through bulk RNA sequencing of human tumors, we identified a strong correlation between WNT7A expression and reduced CD8+ T-cell infiltration. Further investigation demonstrated that inhibiting WNT7A substantially enhanced MHC-I expression on tumor cells. Mechanistically, WNT7A inhibition inactivated the Wnt/β-catenin signaling pathway and thus resulted in reduced physical interaction between β-catenin and p65 in the cytoplasm, which increased the nuclear translocation of p65 and activated the NF-κB pathway, ultimately promoting the transcription of genes encoding MHC-I molecules. We found that our lead compound, 1365-0109, disrupted the protein-protein interaction between WNT7A and its receptor FZD5, resulting in the upregulation of MHC-I expression. In murine tumor models, both genetic and pharmaceutical suppression of WNT7A led to increased MHC-I levels on tumor cells, and consequently enhanced the infiltration and functionality of CD8+ T cells, which bolstered antitumor immunity and improved the effectiveness of immune checkpoint blockade therapy. These findings have elucidated the intrinsic mechanisms of WNT7A-induced immune suppression, suggesting that therapeutic interventions targeting WNT7A hold promise for enhancing the efficacy of immunotherapy.
Project description:BackgroundCoronavirus disease 2019 (COVID-19) is likely to have significant implications for the cardiovascular care of patients. In most countries, containment has already started (on 17 March 2020 in France), and self-quarantine and social distancing are reducing viral contamination and saving lives. However, these considerations may only be the tip of the iceberg; most resources are dedicated to the struggle against COVID-19, and this unprecedented situation may compromise the management of patients admitted with cardiovascular conditions.AimWe aimed to assess the effect of COVID-19 containment measures on cardiovascular admissions in France.MethodsWe asked nine major cardiology centres to give us an overview of admissions to their nine intensive cardiac care units for acute myocardial infarction or acute heart failure, before and after containment measures.ResultsBefore containment (02-16 March 2020), the nine participating intensive cardiac care units admitted 4.8±1.6 patients per day, versus 2.6±1.5 after containment (17-22 March 2020) (rank-sum test P=0.0006).ConclusionsWe confirm here, for the first time, a dramatic drop in the number of cardiovascular admissions after the establishment of containment. Many hypotheses might explain this phenomenon, but we feel it is time raise the alarm about the risk for patients presenting with acute cardiovascular disease, who may suffer from lack of attention, leading to severe consequences (an increase in the number of ambulatory myocardial infarctions, mechanical complications of myocardial infarction leading to an increase in the number of cardiac arrests, unexplained deaths, heart failure, etc.). Similar consequences can be feared for all acute situations, beyond the cardiovascular disease setting.