Bortezomib-based immunosuppression after reduced-intensity conditioning hematopoietic stem cell transplantation: randomized phase II results.

Aprior phase I/II trial of bortezomib/tacrolimus/methotrexate prophylaxis after human leukocyte antigen (HLA)-mismatched reduced intensity conditioning allogeneic hematopoietic stem cell transplantation documented low acute graft-versus-host disease incidence, with promising overall and progression-free survival. We performed an open-label three-arm 1:1:1 phase II randomized controlled trial comparing grade II-IV acute graft-versus-host disease between conventional tacrolimus/methotrexate (A) versus bortezomib/tacrolimus/methotrexate (B), and versus bortezomib/sirolimus/tacrolimus (C), in reduced intensity conditioning allogeneic transplantation recipients lacking HLA-matched related donors. The primary endpoint was grade II-IV acute graft-versus-host disease incidence rate by day +180. One hundred and thirty-eight patients (A 46, B 45, C 47) with a median age of 64 years (range: 24-75), varying malignant diagnoses and disease risk (low 14, intermediate 96, high/very high 28) received 7-8/8 HLA-mismatched (40) or matched unrelated donor (98) grafts. Median follow up in survivors was 30 months (range: 14-46). Despite early immune reconstitution differences, day +180 grade II-IV acute graft-versus-host disease rates were similar (A 32.6%, B 31.1%, C 21%; P=0.53 for A vs B, P=0.16 for A vs C). The 2-year non-relapse mortality incidence was similar (A 14%, B 16%, C 6.4%; P=0.62), as were relapse (A 32%, B 32%, C 38%; P=0.74), chronic graft-versus-host disease (A 59%, B 60% C 55%; P=0.66), progression-free survival (A 54%, B 52%, C 55%; P=0.95), and overall survival (A 61%, B 62%, C 62%; P=0.98). Overall, the bortezomib-based regimens evaluated did not improve outcomes compared with tacrolimus/methotrexate therapy. clinicaltrials.gov Identifier: 01754389.

Comprehensive immunophenotyping of solid tumor-infiltrating immune cells reveals the expression characteristics of LAG-3 and its ligands.

Background: Immune cell expression profiling from patient samples is critical for the successful development of immuno-oncology agents and is useful to understand mechanism-of-action, to identify exploratory biomarkers predictive of response, and to guide treatment selection and combination therapy strategies. LAG-3 is an inhibitory immune checkpoint that can suppress antitumor T-cell responses and targeting LAG-3, in combination with PD-1, is a rational approach to enhance antitumor immunity that has recently demonstrated clinical success. Here, we sought to identify human immune cell subsets that express LAG-3 and its ligands, to characterize the marker expression profile of these subsets, and to investigate the potential relationship between LAG-3 expressing subsets and clinical outcomes to immuno-oncology therapies. Methods: Comprehensive high-parameter immunophenotyping was performed using mass and flow cytometry of tumor-infiltrating lymphocytes (TILs) and peripheral blood mononuclear cells (PBMCs) from two independent cohorts of samples from patients with various solid tumor types. Profiling of circulating immune cells by single cell RNA-seq was conducted on samples from a clinical trial cohort of melanoma patients treated with immunotherapy. Results: LAG-3 was most highly expressed by subsets of tumor-infiltrating CD8 T central memory (TCM) and effector memory (TEM) cells and was frequently co-expressed with PD-1. We determined that these PD-1+ LAG-3+ CD8 memory T cells exhibited a unique marker profile, with greater expression of activation (CD69, HLA-DR), inhibitory (TIM-3, TIGIT, CTLA-4) and stimulatory (4-1BB, ICOS) markers compared to cells that expressed only PD-1 or LAG-3, or that were negative for both checkpoints. In contrast to tumors, LAG-3 expression was more limited in circulating immune cells from healthy donors and solid tumor patients. Additionally, we found abundant expression of the LAG-3 ligands MHC-II and galectin-3 in diverse immune cell types, whereas FGL1 and LSECtin were minimally expressed by immune cells in the tumor microenvironment (TME). Lastly, we found an inverse relationship between baseline and on-treatment levels of circulating LAG3 transcript-expressing CD8 memory T cells and response to combination PD-1 and CTLA-4 blockade in a clinical trial cohort of melanoma patients profiled by scRNAseq. Conclusions: These results provide insights into the nature of LAG-3- and ligand-expressing immune cells within the TME, and suggest a biological basis for informing mechanistic hypotheses, treatment selection strategies, and combination immunotherapy approaches to support continued development of dual PD-1 and LAG-3 blockade.