Upregulation Of LAG-3 Research Articles

Single-Cell Transcriptomic Analysis of Epstein-Barr Virus-Associated Hemophagocytic Lymphohistiocytosis.

Epstein-Barr virus (EBV) infection can lead to infectious mononucleosis (EBV-IM) and, more rarely, EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH), which is characterized by a life-threatening hyperinflammatory cytokine storm with immune dysregulation. Interferon-gamma (IFNγ) has been identified as a critical mediator for primary HLH; however, the detailed role of IFNγ and other cytokines in EBV-HLH is not fully understood. In this study, we used single-cell RNA sequencing to characterize the immune landscape of EBV-HLH and compared it with EBV-IM. Three pediatric patients with EBV-HLH with different backgrounds,one with X-linked lymphoproliferative syndrome type 1 (XLP1), two with chronic active EBV disease (CAEBV), and two patients with EBV-IM were enrolled. The TUBA1B + STMN1 + CD8 + T cell cluster, a responsive proliferating cluster with rich mRNA detection, was explicitly observed in EBV-IM, and the upregulation of SH2D1A-the gene responsible for XLP1-was localized in this cluster. This proliferative cluster was scarcely observed in EBV-HLH cases. In EBV-HLH cases with CAEBV, upregulation of LAG3 was observed in EBV-infected cells, which may be associated with an impaired response by CD8 + T cells. Additionally, genes involved in type I interferon (IFN) signaling were commonly upregulated in each cell fraction of EBV-HLH, and activation of type II IFN signaling was observed in CD4 + T cells, natural killer cells, and monocytes but not in CD8 + T cells in EBV-HLH. In conclusion, impaired responsive proliferation of CD8 + T cells and upregulation of type I IFN signaling were commonly observed in EBV-HLH cases, regardless of the patients' background, indicating the key features of EBV-HLH.

Abstract CT112: An off-the-shelf multivalent vaccine containing cancer’s dark matter, DPV-001, combined with PD-1 +/- GITR in head & neck cancer: safety, efficacy, and immunodynamics from the phase 1 GITRVax trial

Abstract Background: Immune responses to shared cancer antigens have correlated with durable clinical responses. DPV-001 is a novel microvesicle vaccine that contains >300 proteins for genes commonly overexpressed by cancer and is enriched for short-lived proteins (SLiPs), and defective ribosomal products (DRiPs), including non-canonical peptides, representing potential shared non-mutated alternative cancer neoantigens (dark matter). Preclinical studies identified increased therapeutic efficacy when this vaccine strategy was combined with anti-PD-1 and anti-GITR, leading to this clinical trial for patients with advanced or metastatic HNSCC. Methods: Following safety run-in, eligible pts were randomly assigned 1:1 to receive DPV-001 +/- GITR agonist mAb (INCAGN-1949; q2wks). All received sequenced PD-1 mAb (retifanlimab; q4wks) starting D15. Safety was evaluated as the primary endpoint and efficacy was evaluated as a secondary endpoint. Tumor biopsies were taken pre-treatment, Wk 2 and 8, and assessed by CITE-seq, scRNA-seq, BCR-seq, and TCR-seq. Multiplex immunofluoresence was performed on biopsies. Blood samples were taken pre-treatment and at multiple timepoints and analyzed by flow cytometry and seromics. Results: 18 pts received backbone therapy of DPV-001 + sequenced PD-1; of these, 9 pts also received GITR starting D1 (Arm 2). G3 irAE's included: hypothyroidism & mucositis (Arm 1, same pt); pneumonitis and adrenal insufficiency (Arm 2, separate pts). No G4 toxicity was observed. Combining both arms, the response rate (RR) for PD-1 naïve pts was 55% (5/9), and 33% (3/9) for PD-1 experienced pts. Two CR's have been observed in Arm 2. Durability of response (DOR) and progression-free survival (PFS) will be reported at the meeting. DPV-001 induced increased Ki67+ CD4 EM (p<0.042) by D15, prior to first dose of PD-1. Addition of GITR at D1 further increased Ki67+ CD4 (p=0.008) & CD8 EM cells (P=0.0006) by D15. Activated or proliferating CD4 and CD8 EM were significantly increased in both arms by D30 (p<0.025). Across above specified analyses, pts receiving GITR had the greatest increase. Evaluable pts had significantly (all p<0.032) increased numbers of TIL expressing IFN-γ and/or GZMB, LAG-3, and TIM3 in on-treatment biopsies. TCR, BCR, and seromic analyses, as well as studies to identify targets of anti-cancer immunity, are ongoing. Conclusions: This 18 pt trial of DPV-001 and sequenced PD-1 +/- GITR shows a promising RR and evidence of increased activation and expansion of effector T cells in PBL and tumor. Upregulation of LAG-3 and TIM3 by T cells that infiltrate the tumor and have increased in number, provide a rationale for including inhibitors for both in this treatment strategy. Current efforts include evaluating whether immune responses target shared non-canonical alternative neoantigens, or Dark Matter, contained in DPV-001, and whether synchronized antibody and cellular response is evidenced. Acknowledgements: Incyte, Providence Portland Medical Foundation, Steve and Cindy Harder, Trial Registry: NCT04470024, Ethics Approval: PSJH IRB# 2020000480 Citation Format: Rom S. Leidner, Christopher Paustian, Shawn M. Jensen, Tarsem Moudgil, Yoshinobu Koguchi, Noah Simons, Venkatesh Rajamanickam, Ryan Meng, Marcus A. Couey, Quyen Vu, Matthew H. Taylor, Anne Long, Abigail J. Staeck, Lakhvir Kaur, Tijana Jovanovic, Brady Bernard, Tanisha Christie, Noriko Iwamoto, Yuriko Minegishi, Koji Ueda, William Redmond, Lessli M. Rushforth, Takashi Shimada, Patrick Rethwisch, Ashish Patel, Richard B. Bell, Carlo B. Bifulco, Traci L. Hilton, Hong-Ming Hu, Brian Piening, Walter J. Urba, Bernard A. Fox. An off-the-shelf multivalent vaccine containing cancer’s dark matter, DPV-001, combined with PD-1 +/- GITR in head & neck cancer: safety, efficacy, and immunodynamics from the phase 1 GITRVax trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT112.

Circulating extracellular vesicles derived from tumor endothelial cells hijack the local and systemic anti-tumor immune response: Role of mTOR/G-CSF pathway

Circulating tumour-derived extracellular vesicles are supposed to contribute to the spreading of distant metastasis. In this study, we investigated the impact of circulating extracellular vesicles derived from tumour-endothelial cells (TEVs) in the expansion of the metastatic bulk. We focus on the role of immune cells in controlling this process using the 4T1 triple negative breast cancer (TNBC) syngeneic model.4T1 cells were intravenously injected and exposed to circulating TEVs from day 7. The lung, spleen, and bone marrow (BM) were recovered and analysed. We demonstrated that circulating TEVs boost lung metastasis and angiogenesis. FACS and immunohistochemically analyses revealed a significant enrichment of Ly6G+/F4/80+/CD11b+ cells and Ly6G+/F4/80-/CD11b+ in the lung and in the spleen, while Ly6G+/F4/80-/CD11b+ in the BM, indicating the occurrence of a systemic and local immune suppression. TEV immune suppressive properties were further supported by the increased expression of PD-L1, PD-1, and iNOS in the tumour mass. In addition, in vitro experiments demonstrated an increase of CD11+ cells, PD-L1+ myeloid and cancer cells, upregulation of LAG3, CTLA4 and PD-1 in T-cells, release of ROS and NOS, and impaired T-cell-mediated cytotoxic effect in co-culture of TEVs-preconditioned PBMCs and cancer cells. Granulocyte-colony stimulating factor (G-CSF) level was increased in vivo, and was involved in reshaping the immune response. Mechanistically, we also found that mTOR enriched TEVs support G-CSF release and trigger the phosphorylation of the S6 (Ser235/236) mTOR downstream target. Overall, we provided evidence that circulating TEVs enriched in mTOR supported G-CSF release thereby granting tumour immune suppression and metastasis outgrowth.

17th International Conference on Malignant Lymphoma, Palazzo dei Congressi, Lugano, Switzerland, 13 - 17 June, 2023.

Background: Classical Hodgkin lymphoma (cHL), the most common lymphoma in young adults, is a disease of poorly understood heterogeneity. The need for enriching tumor cells before assaying has limited the granularity of its molecular sub-classification. Although the cure rates of patients with cHL have increased over time, new biomarkers are still needed for more precise treatment decision-making in this complex patient population. Patients with cHL frequently have significant amounts of circulating tumor DNA (ctDNA), suggesting an alternative approach for studying this enigmatic tumor. Methods: IOSI-EMA003 (NCT03280394) is a prospective, observational, multi-center study of adult patients with newly diagnosed cHL aiming at: (i) identifying subgroups of patients with phenotype- and outcome-associated molecular signatures; (ii) testing and validating baseline ctDNA load as a prognostic biomarker; (iii) testing if ctDNA can be used as biomarker for the early identification of chemoresistance. Blood samples were collected at staging and disease response assessment. PET scans were centrally and blindly reviewed. ctDNA was analyzed by using deep targeted and low pass whole genome sequencing to measure ctDNA load, capture mutations and profile ctDNA fragmentation patterns. Single-cell RNA sequencing has been used to deconvolute the composition of the tumor microenvironment. Results: A total of 215 patients were enrolled. Based on ctDNA fragmentation patterns reflecting chromatin accessibility in the regulatory region of germinal center (GC) B-cell genes, we segregated cHL into two previously unknown subgroups that we named SNCD (for sub-nucleosomal cfDNA) and NCD (for nucleosomal cfDNA). We hypothesized that SNCD cHL stems from a cell that is closer to the GC B-cell differentiation stage than NCD cHL. SNCD cHL and NCD cHL differed in many aspects, including activation induced cytidine deaminase (AID)-hypermutation profile, neoantigen load, immune editing mechanism and response to chemotherapy and checkpoint inhibitors. Clinically, SNCD cHL displayed less sensitivity to both chemotherapy and anti-PD1 antibodies. Immune editing of SNCD cHL points loss of MHC-I, recruitment of Treg and upregulation of LAG3 as prominent immune checkpoint. High load of pre-treatment ctDNA nominated high-risk patients more accurately than clinico-radiological features. In addition, persistence of residual 18FDG avid lesions along with measurable ctDNA after two chemotherapy courses was a better proof of chemoresistance than the sole persistence of residual 18FDG avid lesions. Also, persistence of measurable ctDNA after the end of therapy invariably predicted progression. Conclusions: Collectively, our results provide a roadmap for cHL subtyping and for the clinical use of ctDNA as a biomarker to aid risk stratification and guide treatment decisions in a more personalized approach. Encore Abstract - previously submitted to ASCO 2023 The research was funded by: Research Advisory Board of the Ente Ospedaliero Cantonale, ABREOC 2019-22514, Bellinzona, Switzerland; European Research Council (ERC) Consolidator Grant CLLCLONE, ID: 772051; Swiss Cancer Research Foundation, ID 3746, 4395, 4660, 5257, Bern, Switzerland; Swiss National Science Foundation, ID 320030_169670/1, 310030_192439, 320036_179318, Berne, Switzerland; The Leukemia & Lymphoma Society, Translational Research Program, ID 6594-20, New York; AFRI, Ente Ospedaliero Cantonale, Bellinzona, Switzerland; ASH Global Research Award, Washington DC, USA; Worldwide Cancer Research (ID: 10583); ISREC Foundation; Helmut Horten Foundation, Switzerland; Fondazione Fidinam, Lugano, Switzerland; Nelia & Amadeo Barletta Foundation, Lausanne, Switzerland; Fond’Action, Lausanne, Switzerland; Fondazione Jacob, Lugano, Switzerland; Fondazione Dr. Ettore Balli, Lugano, Switzerland; Fondazione Ticinese per la Ricerca sul Cancro, Bellinzona, Switzerland; Jacques & Gloria Gossweiler Foundation, Berne, Switzerland. Keywords: Diagnostic and Prognostic Biomarkers, Hodgkin lymphoma, Liquid biopsy Conflicts of interests pertinent to the abstract. M. C. Pirosa Educational grants: BeiGene A. Condoluci Honoraria: AbbVie, AstraZeneca and Janssen Research funding: Gilead and Pfizer A. Moccia Honoraria: Janssen and Takeda A. Stathis Honoraria: Eli Lilly, Bayer, Roche, Novartis, Janssen Oncology, AstraZeneca Research funding: ADC Therapeutics, Pfizer, Roche, Novartis, Bayer, Eli Lilly, MEI Pharma, Cellestia, Debiopharm Group, Merck/MSD, Abbvie, Amgen, AstraZeneca, Incyte, Loxo, and Philogen C. Carlo-Stella Consultant or advisory role: Celgene/Bristol-Myers Squibb, ADC Therapeutics, Karyopharm 767 Therapeutics, Roche, and Sanofi Honoraria: Bristol-Myers Squibb, Merck Sharp & Dohme, Janssen Oncology, AstraZeneca, Incyte, Novartis, Takeda, ADC Therapeutics Research funding: ADC Therapeutics, Sanofi, Roche G. Gaidano Honoraria: Abbvie, Astra Zeneca, BeiGene, Incyte, Janssen E. Zucca Honoraria: BeiGene, BMS, Celltion Healthcare, Curis, Eli/Lilly, Incyte, Ipsen, Janssen, Kyte, Merck, Miltenyi, Roche Research funding: AstraZeneca, BMS, Incyte, Janssen, Merck, Roche Educational grants: Abbvie, BeiGene, Janssen and Roche D. Rossi Honoraria: AbbVie, AstraZeneca, BeiGene, BMS, Janssen Research funding: AbbVie, AstraZeneca, Janssen Educational grants: AstraZeneca and Janssen.

Circulating tumor DNA of classical Hodgkin lymphoma.

7516 Background: Classical Hodgkin lymphoma (cHL) is the most common lymphoma in young adults. The morphologic peculiarity that makes cHL unique are the few tumor cells surrounded by a polytypic inflammatory microenvironment. It is also the cancer with the highest sensitivity to immune checkpoint inhibitors. But why is it so immunogenic and how it escapes the immune control at the same time have only been partially elucidated. Although cure rates of cHL have constantly increased over the years there is still a high need for identification of biomarkers for more precise treatment decisions. The need for enriching tumor cells before assaying has limited cHL biological studies. Patients with cHL have significant amounts of circulating tumor DNA (ctDNA), suggesting an alternative approach for studying this enigmatic tumor. Methods: IOSI-EMA003 (NCT03280394) is a prospective, observational, multi-center study of adult patients with previously untreated cHL aiming at: i) identifying subgroups of patients with phenotype- and outcome-associated molecular signatures; ii) testing and validating baseline ctDNA load as a prognostic biomarker; iii) testing if ctDNA can be used for the early identification of chemoresistance. Blood samples were collected at staging and disease response assessment. PET scans were centrally and blindly reviewed. ctDNA was analyzed by using deep targeted and shallow whole genome sequencing to measure ctDNA load, capture mutations and profile ctDNA fragmentation patterns. The composition of the tumor microenvironment of cHL was probed by single cell RNA-seq. Results: A total of 215 patients were enrolled. Based on ctDNA fragmentation patterns reflecting chromatin accessibility in the regulatory region of germinal center (GC) B-cell genes, we segregated cHL into two subgroups that we named SNCD (for sub-nucleosomal cfDNA) and NCD (for nucleosomal cfDNA). We hypothesized that SNCD cHL stems from a cell that is closer to the GC B-cell differentiation stage than NCD cHL. SNCD cHL and NCD cHL differed in many aspects, including activation induced cytidine deaminase (AID)-hypermutation profile, neoantigen load, immune editing mechanism and response to chemotherapy and checkpoint inhibitors. Clinically, SNCD cHL displayed less sensitivity to both chemotherapy and anti-PD1 antibodies. Immune editing of SNCD cHL points loss of MHC-I, recruitment of Treg and upregulation of LAG3 as prominent immune checkpoint. High load of pre-treatment ctDNA nominated high-risk patients more accurately than clinico-radiological features. In addition, persistence of residual 18FDG avid lesions and measurable ctDNA was a better proof of chemoresistance than the sole persistence of residual 18FDG avid lesions. Conclusions: Our results provide a roadmap for cHL subtyping and molecular classification. We also propose ctDNA as a quantifiable and radiation-free biomarker to be used in clinical trials testing a more personalized treatment approach. Clinical trial information: NCT03280394 .

A 3D Ex Vivo Tumor-Immune Coculture System Mimicking In Vivo Tumor Environmental Stress on CD8+ T Cells Exhaustion.

Dissection of exhaustion trajectories of immune cells under tumor selection pressure in the tumor microenvironment (TME) elucidates the underlying machinery in anti-tumor immunity, which still lacks easy-to-use models to decipher. Herein,gelatin methacryloyl (GelMA)-poly (ethylene oxide) (PEO) based 3D hydrogel microspheroids are constructed with non-immunogenicity and controllable macroporous structure to establish a tumor-immune cell coculture (3D-HyGTIC) system. In 3D-HyGTIC system, when immune cells embarked, stepwise up-regulation of main immune checkpoints (ICs) molecules is observed with compromised cytokine production in CD8+ T cells, the trajectory of which is in lineage correlation with in vivo grafted tumors. Reinvigoration of CD8+ T cells is more obvious with the addition of an anti-PD-1 regimen at the early time point, which is recapitulated during the coculture of patient-derived tumor fragments (PDTF) and autologous T cells. Moreover, the upregulation of LAG-3 on CD8+ T cells after anti-PD-1 treatment is uncovered. Sequential addition of anti-LAG-3 successfully rescues the otherwise failed reactivation of CD8+ T cells. Therefore, the 3D-HyGTIC system is not only inclined to mimic the early differentiation trajectories of tumor-infiltrating CD8+ T cells but also may facilitate an evaluation of the efficacy of IC blockades and guide the designing of combination immunotherapy.

Abstract 2874: The tetravalent structure of FS118, a bispecific antibody targeting LAG-3 and PD-L1, is required for its novel mechanism of LAG-3 shedding

Abstract FS118 is a tetravalent bispecific antibody targeting LAG-3 and PD-L1 that can overcome immune suppressive signals with greater preclinical activity than a combination of monoclonal antibodies. In vivo, FS118 downregulates LAG-3 on tumor-infiltrating lymphocytes (TILs) and increases soluble LAG-3 (sLAG-3) in the serum of mice. In a Phase 1 trial, FS118 demonstrated a dose-dependent increase in sLAG-3 in the serum of patients with advanced malignancies. Here, we demonstrate that the tetravalent structure of FS118 is required to mediate this novel LAG-3 shedding mechanism. Human ex vivo assays were performed by co-culturing expanded CD4+ T cells with iDCs in the presence of Staphylococcal enterotoxin B and FS118, or controls. sLAG-3 was measured by ECLIA. The binding valency of FS118 was determined using chemically-crosslinked mass spectrometry mapping (XL/MS). Variants of FS118 and FS118 surrogate molecules were generated with differing valency for LAG-3 and PD-L1. MC38 tumor-bearing C57BL/6 mice were dosed once interperitoneally with FS118 surrogate or valency variants. TILs were analyzed by flow cytometry, and sLAG-3 was measured in the serum by ECLIA. In patients receiving FS118 treatment, a dose-dependent increase in sLAG-3 was detected in the blood. In an ex vivo human T cell assay, the FS118-mediated increase in sLAG-3 was greater than with the combination of the individual bispecific components. sLAG-3 increase by FS118 was mediated by ADAM10 and ADAM17. FS118 demonstrated tetravalent binding to its target antigens using XL/MS. All four binding sites simultaneously bound to target antigens, with preferential binding to PD-L1 in the presence of equal amounts of both LAG-3 and PD-L1. Variants of FS118 with reduced binding valency for target antigens mediated lower levels of LAG-3 release than tetravalent FS118. Compared to FS118, monovalent PD-L1 binding reduced sLAG-3 release and importantly, a variant of FS118 with monovalent LAG-3 binding generated the lowest level of sLAG-3. Data will be presented showing the role of binding valency on LAG-3 downregulation by TILs and T cell activation in ex vivo murine samples. FS118 is tetravalent and can bind to two LAG-3 and two PD-L1 molecules simultaneously. Bivalent LAG-3 and PD-L1 binding is required for enhanced shedding of cell surface LAG-3 in vitro. Removing LAG-3 from the surface of exhausted TILs is a novel mechanism attributed to the tetravalent structure of FS118, which may be important to overcome compensatory upregulation of LAG-3 induced by blockade of PD-L1 in patients. Citation Format: Claire S. Reader, Wenjia Liao, Beatrice J. Potter-Landau, Christel Séguy Veyssier, Martyn C. Rhoades, Claire J. Seal, Michelle Morrow, Neil Brewis. The tetravalent structure of FS118, a bispecific antibody targeting LAG-3 and PD-L1, is required for its novel mechanism of LAG-3 shedding [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2874.

Immunome Single Cell Profiling Reveals T Cell Exhaustion with Upregulation of Checkpoint Inhibitors LAG3 and Tigit on Marrow Infiltrating T Lymphocytes in Daratumumab and IMiDs Resistant Patients

Background: The addition of immunomodulatory (IMiDs) drugs to monoclonal antibodies targeting the transmembrane glycoprotein CD38 has demonstrated very encouraging and durable responses in myeloma patients. It is believed that the synergistic effects observed with anti-CD38 antibodies and IMiDs are derived from their co-modulation of the host adaptive and innate immunity and therefore it is plausible to speculate that acquired resistance to daratumumab and IMiDs may be largely immune-mediated. The aim of the present study was to 1) interrogate at the single cell level the bone marrow immune repertoire of daratumumab sensitive and resistant patients, 2) identify cellular mediators of resistance to anti-CD38 antibodies and 3) define potential means to reinstate sensitivity to daratumumab and IMiDs.Methods and Results: Serial BM aspirates (n=44) were collected from patients treated with single agent daratumumab (MMY3012 trial) or daratumumab + pomalidomide (MM014 trial) prior to initiation of therapy, C3D1 and at relapse. Bone marrow mononuclear fractions were isolated through ficoll density gradients coupled with magnetic sorting of CD138pos and CD138neg cells. Unbiased mRNA profiling of BM CD138neg cells was performed by single-cell RNA-seq (scRNA-seq) using the GemCode system (10x Genomics). Paired-end sequencing was performed on Illumina NEXTseq and NOVAseq platforms. Cell Ranger Single and Seurat were used for sample de-multiplexing, barcode processing, single-cell 3′ gene counting and data analysis. Sequencing data were analyzed by principal component analysis (PCA), clustering with multi-sample batch correction and then visualized by t-distributed stochastic neighbor embedding (t-SNE) projection. Comparison of the single cell transcriptomes of CD138neg cells from responding patients pre- and post- treatment revealed that Daratumumab and Pomalidomide dramatically modify the immune cells composition (immunome) of the bone marrow niches leading to: 1) expansion of effector T cells (KLRG1high, GZMAhigh, CCL5high), 2) significant depletion of CD38high / FCGR3Ahigh NK cells with retained population of cytotoxic NK cells (CD27high, KLRB1high, NCR3high, GZMApos, PRF1pos), 3) depletion of FCGR3Ahigh / CD14low monocytes, 4) expansion of M1 inflammatory macrophages and depletion of plasmacytoid dendritic cells. Similar changes were seen in patients treated with single agent daratumumab (without IMiDs) however with a lesser expansion of effector T cells and in particular reduced marrow infiltrating inflammatory macrophages. In contrast, the immunome of daratumumab and pomalidomide resistant patients was characterized by a reduced central memory T cells (TCM), and a largely exhausted effector T cells populations that are CD28neg and expressing checkpoint inhibitors (LAG3and TIGIT significantly more than PDCD1) as well as high expression of TIM3 (HAVCR2) on marrow macrophages. Upregulation of LAG3 and TIGIT expression on T cells was also confirmed at the antigenic level by flow cytometry. Consistent with the non-bystander and suppressive effect of LAG3 and TIGIT on the function of effector T cells, activation (CD107a expression) and proliferation of LAGpos and/or TIGITpos sorted bone marrow T cells from resistant patients were significantly reduced in response to autologous myeloma cells stimulation or CD3/CD28 crosslinking. Lastly, a higher proportion and number of clonal T cell (through single cell TCR sequencing) was also observed in responding (≥ PR) vs non-responding (< PR) patients. Interrogation of the myeloma cells transcriptome, showed little to no loss of CD38 transcript at the time of acquired resistance with rather upregulation of complement inhibitory molecule CD59 and NFκB signature genes.Conclusion: A systematic unsupervised interrogation of the bone marrow immunome of daratumumab and IMiDs treated MM patients demonstrated a significant activation of adaptive and innate immunity in responding patients and revealed an expansion of exhausted T cells with upregulation of the checkpoint inhibitors LAG3 and TIGIT in resistant patients. Our findings warrant the exploration of LAG3- and/or TIGIT-blocking strategies as potential means to reinstate sensitivity to daratumumab and IMiDs in myeloma patients. DisclosuresNeri:Janssen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria. McCulloch:Celgene: Honoraria; Takeda: Other: Travel expenses. Thakurta:Celgene Corporation: Employment, Equity Ownership. Bahlis:Celgene: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria; Janssen: Consultancy, Honoraria, Research Funding.

The upregulation of LAG-3 on T cells defines a subpopulation with functional exhaustion and correlates with disease progression in HIV-infected subjects.

T cells develop functional defects during HIV-1 infection, partially due to the upregulation of inhibitory receptors such as programmed death-1 (PD-1) and CTLA-4. However, the role of lymphocyte activation gene-3 (LAG-3; CD223), also known as an inhibitory receptor, in HIV infection remains to be determined. In this study, we revealed that LAG-3 on T cells delivers an inhibitory signal to downregulate T cell functionality, thereby playing an immunoregulatory role during persistent HIV-1 infection. We observed that HIV-1 infection results in a significant increase in LAG-3 expression in both the peripheral blood and the lymph nodes. The upregulation of LAG-3 is dramatically manifested on both CD4(+) and CD8(+) T cells and is correlated with disease progression. As expected, prolonged antiretroviral therapy reduces the expression of LAG-3 on both CD4(+) and CD8(+) T cells. The ex vivo blockade of LAG-3 significantly augments HIV-specific CD4(+) and CD8(+) T cell responses, whereas the overexpression of LAG-3 in T cells or the stimulation of LAG-3 on T cells leads to the reduction of T cell responses. Furthermore, most LAG-3 and PD-1 are expressed in different T cell subsets. Taken together, these data demonstrate that the LAG-3/MHC class II pathway plays an immunoregulatory role, thereby providing an important target for enhancing immune reconstitution in HIV-infected patients. Additionally, the LAG-3/MHC class II pathway may synergize with PD-1/PD ligand to enhance T cell-mediated immune responses.