Paulo C. Rodriguez

Inhibition of KRASG12C (G12Ci) has shown promising clinical activity in several cancers; however, emergence of resistance is a major limitation to long-term benefit. It remains unknown how G12Ci affects the tumor-immune microenvironment and its association with resistance. We investigated the immune-modulatory impact of the G12Ci sotorasib in KrasG12C/Trp53−/− lung and colon carcinoma models. Single-cell RNA sequencing of tumors in both models demonstrated sotorasib-induced loss of proliferative tumor cells and the enrichment of immune-active tumor cells with elevated NF-κB and IFN signaling and elevated MHC-I expression. These changes led to an increased proportion of effector T cells and MHC-IIhigh macrophages in the tumors and a reduced proportion of regulatory myeloid cells. Furthermore, sotorasib-resistant tumors exhibited a marked reduction of MHC-Ihigh immune-active tumor cells and immune cell type changes. Enrichment of immune-active tumor cells was associated directly with sotorasib-induced activation of NF-κB and STAT1, leading to heightened responsiveness to inflammatory cytokines and upregulation of MHC-I and chemokine expression. These findings indicate that modulation of immune-active tumor cell populations is a key mechanism associated with G12Ci therapeutic efficacy and the development of resistance. Watch first author Mohammed L. Ibrahim, B. Pharm, PhD, from California Northstate University College of Medicine describe this article: https://vimeo.com/showcase/12408339?video=1227805308.

Ubiquitination is an important posttranslational modification associated with essential cellular processes. Here we show that deletion of E3 ubiquitin ligase HECTD3 — germline or in hematopoietic compartment, including in Cd11c ⁺ cells — causes severe disease and increased production of proinflammatory cytokines in a murine model of ulcerative colitis (UC). HECTD3 mRNA levels were found reduced in colonic tissues of patients with UC, which highlights a potential role for HECTD3 in regulating inflammatory responses in UC. We identified MyD88, a central adaptor in the TLR/IL-1R signaling and inflammatory response, as a target for HECTD3 ubiquitination. We demonstrate that HECTD3 ubiquitinates MyD88 through K27-linked Poly-Ub chains in a nondegradative manner. HECTD3-KO GM-CSF–bone marrow–derived cells treated with the TLR4 ligand LPS produced more proinflammatory cytokines, showed elevated phosphorylation of NF-κB and IRAK4, as well as increased association of MyD88 with IRAK4, demonstrating that HECTD3 controls MyD88/IRAK4/NF-κB axis. Inhibition of MyD88 activity rescued colitis severity in the HECTD3-KO mice, including the elevated proinflammatory cytokine production. Thus, our results establish MyD88 as a target for HECTD3 nondegradative polyubiquitination and restriction of innate immune colonic inflammation.

Chimeric antigen receptor (CAR) T cell therapies have revolutionized B cell malignancy treatment, but many patients with large B cell lymphoma (LBCL) experience primary resistance or relapse. To uncover resistance mechanisms, here we examine pre-infusion tumor biopsies and observe that increased immunoregulatory macrophages correlate with poor clinical responses. In murine models, CAR T cell-produced interferon-gamma (IFN-γ) upregulates inducible nitric oxide synthase (iNOS, NOS2) in immunoregulatory macrophages, impairing CAR T cell function. Proteomic profiling reveals that iNOS-expressing macrophages promote apoptosis and cell cycle arrest while downregulating protein synthesis machinery in CAR T cells. Metabolically, CAR T cells exhibit reduced glycolytic intermediates and altered tricarboxylic acid (TCA) cycle activity. Pharmacological inhibition of iNOS enhances CAR T cell treatment efficacy in vivo. Elevated levels of iNOS⁺CD14⁺ monocytes in leukaphereses are associated with non-durable responses to CAR T cells. Targeting iNOS in immunoregulatory macrophages, potentially by modulating CAR T-produced IFN-γ, could improve LBCL outcomes.

Myeloid-derived suppressor cells (MDSCs) in the tumor microenvironment (TME) limit the efficacy of adoptive T cell therapies, highlighting the need to overcome tumor-associated immunosuppression. Sphingosine-1-phosphate (S1P), is an abundant signaling lipid in the TME. Here, we show that inhibition of sphingosine kinase-2 (SphK2), the enzyme generating S1P in MDSCs, reduces the suppressive activity of monocytic MDSCs (M-MDSCs) while promoting their differentiation toward a mature, immunogenic phenotype characterized by enhanced antigen presentation. Pharmacological SphK2 inhibition enhances the response to anti-PD-1 therapy in preclinical models of checkpoint-resistant breast, bladder, and melanoma cancers by mitigating MDSC-mediated suppression and limiting tumor progression. Mechanistically, S1P directly binds acetyl-CoA carboxylase-1 (ACC1) to inhibit its activity, thereby rewiring fatty-acid metabolism. Lowering intracellular S1P restores ACC activity, promotes phosphatidylcholine synthesis, and reduces MDSC immunosuppression. These findings identify the SphK2-ACC-phospholipid axis as a metabolic checkpoint controlling the immunogenicity of MDSCs and a potential therapeutic target for enhancing cancer immunotherapy.

GPR84 receptor modulation reduces apoptosis in CD8+ T cells.

GPR84 antagonism improves anti-tumor therapeutic vaccine efficacy in an in vivo multi-boost immunization regimen.

Top 50 differentially expressed genes between GLPG1205 and DMSO in CD8+ T cells.

Top 50 differentially expressed genes between DL175 and DMSO in CD8+ T cells.

BM-MDSCs express more GPR84 mRNA compared to T cells.

Validation of CRISPR-Cas9 and Global GPR84 knockout.