Search

A key unknown of the functional space in tumor immunity is whether CD4 T cells depend on intratumoral MHCII cancer antigen recognition. MHCII-expressing, antigen-presenting cancer-associated fibroblasts (apCAFs) have been found in breast and pancreatic tumors and are considered to be immunosuppressive. This analysis shows that antigen-presenting fibroblasts are frequent in human lung non-small cell carcinomas, where they seem to actively promote rather than suppress MHCII immunity. Lung apCAFs directly activated the TCRs of effector CD4 T cells and at the same time produced C1q, which acted on T cell C1qbp to rescue them from apoptosis. Fibroblast-specific MHCII or C1q deletion impaired CD4 T cell immunity and accelerated tumor growth, while inducing C1qbp in adoptively transferred CD4 T cells expanded their numbers and reduced tumors. Collectively, we have characterized in the lungs a subset of antigen-presenting fibroblasts with tumor-suppressive properties and propose that cancer immunotherapies might be strongly dependent on in situ MHCII antigen presentation.

In situ antigen presentation is required to sustain active proliferating CD4+ T cells in tumours and to help form memory CD8+ T cells, but the antigen presenting cells (APCs) and pathways involved remain elusive. Cancer associated fibroblasts (CAFs) are prominent stromal constituent of solid tumors. Current immunological dogma considers that CAFs facilitate tumour immune escape. A new subset of MHCII antigen presenting cancer-associated fibroblasts (apCAFs) has been described, but their function remains unknown. Here we report a previously unrecognized function of apCAFs in sustaining CD4+ T cells in primary human and murine lung tumours. In response to IFNγ and oxidative stress in tumours CAFs up-regulated MHCII. Fibroblast-specific targeted ablation of MHCII induced a hypometabolic hypoproliferative state in CD4+ T cells, which impacted MHCII and MHCI immunity, accelerating tumor growth. apCAFs directly presented MHCII-peptide (MCHIIp) complexes to activate the TCRs of CD4+ T cell. Highthrough-put profiling and blocking assays unveiled a novel CAF to T cell communication pathway via complement 1q binding on membrane C1qbp. Thus, apCAFs sustain anti-tumor CD4+ T cells via MHCIIp-TCR and C1q-C1qbp binding. Our studies pave the way to the design of novel immunotherapeutic strategies that will harness apCAFs to help sustain T cells inside solid tumours. Statement of significance This work challenges the immunological dogma that dominant physiological significance in tumour specific immunity comes only through professional APCs, leading to the design of novel MHCII fibroblast-directed strategies to sustain endogenous or adoptively transferred CD4+ T cells within solid tumors.

Summary Chimeric antigen receptor anti-CD19 (CAR19)-T cell immunotherapy-induced clinical remissions in CD19+ B cell lymphomas are often short lived. We tested whether CAR19-engineering of the CD1d-restricted invariant natural killer T (iNKT) cells would result in enhanced anti-lymphoma activity. CAR19-iNKT cells co-operatively activated by CD1d- and CAR19-CD19-dependent interactions are more effective than CAR19-T cells against CD1d-expressing lymphomas in vitro and in vivo. The swifter in vivo anti-lymphoma activity of CAR19-iNKT cells and their enhanced ability to eradicate brain lymphomas underpinned an improved tumor-free and overall survival. CD1D transcriptional de-repression by all-trans retinoic acid results in further enhanced cytotoxicity of CAR19-iNKT cells against CD19+ chronic lymphocytic leukemia cells. Thus, iNKT cells are a highly efficient platform for CAR-based immunotherapy of lymphomas and possibly other CD1d-expressing cancers., Graphical Abstract, Highlights • Bespoke protocol for CAR19-iNKT cell transduction and clinical scale expansion • Higher CAR19-iNKT than CAR19-T cell expandability and killing of CD19+CD1d+ targets • CAR19-iNKT cell reactivity potentiation by αGalCer and ATRA • Prolonged survival and brain lymphoma eradication of CAR19-iNKT cell-treated mice, Rotolo et al. show that anti-CD19 chimeric antigen receptor (CAR19)-engineered CD1d-restricted invariant NKT cells (iNKT) are more effective than CAR19-T cells against CD1d-expressing lymphomas, including those in the brain. De-repression of CD1d expression further enhances the anti-tumor efficacy of CAR19-iNKT.

Glycosphingolipids (GSLs) are essential constituents of cell membranes and lipid rafts and can modulate signal transduction events. The contribution of GSLs in osteoclast (OC) activation and osteolytic bone diseases in malignancies such as the plasma cell dyscrasia multiple myeloma (MM) is not known. Here, we tested the hypothesis that pathological activation of OCs in MM requires de novo GSL synthesis and is further enhanced by myeloma cell-derived GSLs. Glucosylceramide synthase (GCS) inhibitors, including the clinically approved agent N-butyl-deoxynojirimycin (NB-DNJ), prevented OC development and activation by disrupting RANKL-induced localization of TRAF6 and c-SRC into lipid rafts and preventing nuclear accumulation of transcriptional activator NFATc1. GM3 was the prevailing GSL produced by patient-derived myeloma cells and MM cell lines, and exogenous addition of GM3 synergistically enhanced the ability of the pro-osteoclastogenic factors RANKL and insulin-like growth factor 1 (IGF-1) to induce osteoclastogenesis in precursors. In WT mice, administration of GM3 increased OC numbers and activity, an effect that was reversed by treatment with NB-DNJ. In a murine MM model, treatment with NB-DNJ markedly improved osteolytic bone disease symptoms. Together, these data demonstrate that both tumor-derived and de novo synthesized GSLs influence osteoclastogenesis and suggest that NB-DNJ may reduce pathological OC activation and bone destruction associated with MM.

Multiple myeloma (MM) is an incurable tumor of the plasma cells, the terminally differentiated immunoglobulin secreting B lineage cells. The genetic make-up of MM has been extensively characterized but its impact on the biology of the disease is incomplete without more precise knowledge of the identity and functional role of cells with multiple myeloma propagating activity (MMPA). We review here recent data that link MMPA with myeloma clonotypic populations organized in a cellular hierarchy that mirrors normal B cell development and also with drug resistance and disease relapse. We further propose a conceptual framework which, with optimal use of recent technological advances in genomics and phenomics, could allow dissection of the cellular and molecular properties of cells with MMPA, drug resistance and in vivo relapse in an integrated and patient-specific manner. There is real hope that these approaches will significantly contribute to further improvements in disease control, overall survival, and possibly cure of patients with MM. Stem Cells 2015;33:3205–3211

Introduction: Development of mature B cells depends upon the expression of a functional B cell receptor (BCR) and of its immunoglobulin (Ig) heavy (H) and light (L) chains. The molecular hallmark of B cell development, somatic recombination of the genes that encode the IgH (V, D and J) and IgL (V and J) chains, takes place in B cells in primary B lymphopoiesis sites throughout development (i.e. fetal liver (FL), fetal bone marrow (FBM) and adult BM). However, the spatiotemporal relationship between the IgH repertoire in FL with that in FBM, and the impact of the fetal Ig repertoire on the long-term repertoire present in post-natal life, as well as the link between this and the development of disease, are largely unknown. Aim: To gain insights into the ontogeny of the human innate B cell repertoire by performing high resolution analysis of the IgH-Cmu repertoire of human FL, FBM and post-natal B cells. Methods: CD34negCD19pos B cells from human 2nd trimester FL and FBM, child peripheral blood (cPB) and adult peripheral blood (aPB) were isolated by flow sorting. For initial assessment of the IgM repertoire ontogeny in FL and FBM B cells, we performed CDR3 fragment length analysis of VH1-VH6 families in the B cells. To gain further insights into the ontogeny of IgH diversification, we employed 454 next generation sequencing (NGS)-based analysis of the IgVH-Cmu mRNA repertoire in FL, FBM and postnatal B cells. In total, 20 libraries generated from 17 individual, flow-sorted CD34negCD19pos B cell samples were sequenced: 5 FL (4 performed in independent duplicate libraries; GA 14-18 weeks), 3 FBM (GA 13-17 weeks), 3 cPB (age 4mo-4yrs) and 5 aPB (age 29-53 yrs) samples. Bioinformatic analysis was performed with the ARResT/Interrogate immunoprofiler, IMGT tools, and appropriate statistics. Results: Spectratyping and NGS data showed that the molecular mechanisms responsible for VDJ recombination-dependent repertoire diversification, such as convergent recombination, are active in early B cell development and as efficient as postnatal B cells in both FL and FBM. Comparably diversified B cell lymphopoiesis exists contemporaneously in FL and FBM. However, analysis of the top 100 most abundant clonotypes in each developmental stage showed that their mean abundance in FBM B cells was 10 times lower (0.12%) than in FL B cells (1.2%; p Conclusions: The lack of mature, antigen-responsive B cells in FBM and the pronounced clonotypic expansions in FL suggest that FL is the main source of IgM natural immunity during the 2nd trimester. Our results are consistent with persisting fetal IgM+ B cells being an important source of the natural IgM repertoire also in adult life. Further, the origins of specific stereotypic IgM+ BCR associated with CLL, can be traced back to fetal B cell lymphopoiesis, suggesting that persisting fetal B cells can be subject to malignant transformation late in life. Overall, these novel data provide unique insights into the ontogeny of physiological and malignant B lymphopoiesis that spans the human lifetime. Disclosures No relevant conflicts of interest to declare.