There is a strong correlation between myeloid-derived suppressor cells (MDSC) and resistance to immune checkpoint blockade (ICB), but the detailed mechanisms underlying this correlation are largely unknown. Using single-cell RNA sequencing analysis in a bilateral tumor model, we found that immunosuppressive myeloid cells with characteristics of fatty acid oxidative metabolism dominate the immune-cell landscape in ICB-resistant subjects. In addition, we uncovered a previously underappreciated role of a serine/threonine kinase, PIM1, in regulating lipid oxidative metabolism via PPARγ-mediated activities. Enforced PPARγ expression sufficiently rescued metabolic and functional defects of Pim1−/− MDSCs. Consistent with this, pharmacologic inhibition of PIM kinase by AZD1208 treatment significantly disrupted the myeloid cell–mediated immunosuppressive microenvironment and unleashed CD8+ T-cell–mediated antitumor immunity, which enhanced PD-L1 blockade in preclinical cancer models. PIM kinase inhibition also sensitized nonresponders to PD-L1 blockade by selectively targeting suppressive myeloid cells. Overall, we have identified PIM1 as a metabolic modulator in MDSCs that is associated with ICB resistance and can be therapeutically targeted to overcome ICB resistance.

Intratumoral Tregs are key mediators of cancer immunotherapy resistance, including anti–programmed cell death (ligand) 1 [anti–PD-(L)1] immune checkpoint blockade (ICB). The mechanisms driving Treg infiltration into the tumor microenvironment (TME) and the consequence on CD8 ⁺ T cell exhaustion remain elusive. Here, we report that heat shock protein gp96 (also known as GRP94) was indispensable for Treg tumor infiltration, primarily through the roles of gp96 in chaperoning integrins. Among various gp96-dependent integrins, we found that only LFA-1 (αL integrin), and not αV, CD103 (αE), or β7 integrin, was required for Treg tumor homing. Loss of Treg infiltration into the TME by genetic deletion of gp96/LFA-1 potently induced rejection of tumors in multiple ICB-resistant murine cancer models in a CD8 ⁺ T cell–dependent manner, without loss of self-tolerance. Moreover, gp96 deletion impeded Treg activation primarily by suppressing IL-2/STAT5 signaling, which also contributed to tumor regression. By competing for intratumoral IL-2, Tregs prevented the activation of CD8 ⁺ tumor-infiltrating lymphocytes, drove thymocyte selection-associated high mobility group box protein (TOX) induction, and induced bona fide CD8 ⁺ T cell exhaustion. By contrast, Treg ablation led to striking CD8 ⁺ T cell activation without TOX induction, demonstrating clear uncoupling of the 2 processes. Our study reveals that the gp96/LFA-1 axis plays a fundamental role in Treg biology and suggests that Treg-specific gp96/LFA-1 targeting represents a valuable strategy for cancer immunotherapy without inflicting autoinflammatory conditions.

In cancer, CD8⁺ T cells enter a dysfunctional state which prevents them from effectively targeting and killing tumor cells. Tumor-infiltrating CD8⁺ T cells consist of a heterogeneous population of memory-like progenitor, effector, and terminally exhausted cells that exhibit differing functional and self-renewal capacities. Our recently published work has shown that interleukin (IL)-21-producing CD4⁺ T cells help to generate effector CD8⁺ T cells within the tumor, which results in enhanced tumor control. However, the molecular mechanisms by which CD4⁺ helper T cells regulate the differentiation of effector CD8⁺ T cells are not well understood. In this study, we found that Basic Leucine Zipper ATF-Like Transcription Factor (BATF), a transcription factor downstream of IL-21 signaling, is critical to maintain CD8⁺ T cell effector function within the tumor. Using mixed bone marrow chimeras, we demonstrated that CD8⁺ T cell-specific deletion of BATF resulted in impaired tumor control. In contrast, overexpressing BATF in CD8⁺ T cells enhanced effector function and resulted in improved tumor control, bypassing the need for CD4⁺ helper T cells. Transcriptomic analyses revealed that BATF-overexpressing CD8⁺ T cells had increased expression of costimulatory receptors, effector molecules, and transcriptional regulators, which may contribute to their enhanced activation and effector function. Taken together, our study unravels a previously unappreciated CD4⁺ T cell-derived IL-21–BATF axis that could provide therapeutic insights to enhance effector CD8⁺ T cell function to fight cancer.

Defective mitophagy contributes to normal aging and various neurodegenerative and cardiovascular diseases. The newly developed methodologies to visualize and quantify mitophagy allow for additional progress in defining the pathophysiological significance of mitophagy in various model organisms. However, current knowledge regarding mitophagy relevant to human physiology is still limited. Model organisms such as mice might not be optimal models to recapitulate all the key aspects of human disease phenotypes. The development of the human-induced pluripotent stem cells (hiPSCs) may provide an exquisite approach to bridge the gap between animal mitophagy models and human physiology. To explore this premise, we take advantage of the pH-dependent fluorescent mitophagy reporter, mt-Keima, to assess mitophagy in hiPSCs and hiPSC-derived cardiomyocytes (hiPSC-CMs). We demonstrate that mt-Keima expression does not affect mitochondrial function or cardiomyocytes contractility. Comparison of hiPSCs and hiPSC-CMs during different stages of differentiation revealed significant variations in basal mitophagy. In addition, we have employed the mt-Keima hiPSC-CMs to analyze how mitophagy is altered under certain pathological conditions including treating the hiPSC-CMs with doxorubicin, a chemotherapeutic drug well known to cause life-threatening cardiotoxicity, and hypoxia that stimulates ischemia injury. We have further developed a chemical screening to identify compounds that modulate mitophagy in hiPSC-CMs. The ability to assess mitophagy in hiPSC-CMs suggests that the mt-Keima hiPSCs should be a valuable resource in determining the role mitophagy plays in human physiology and hiPSC-based disease models. The mt-Keima hiPSCs could prove a tremendous asset in the search for pharmacological interventions that promote mitophagy as a therapeutic target. Abbreviations: AAVS1: adeno-associated virus integration site 1; AKT/protein kinase B: AKT serine/threonine kinase; CAG promoter: cytomegalovirus early enhancer, chicken ACTB/β-actin promoter; CIS: cisplatin; CRISPR: clustered regularly interspaced short palindromic repeats; FACS: fluorescence-activated cell sorting; FCCP: carbonyl cyanide p-trifluoromethoxyphenylhydrazone; hiPSC: human induced pluripotent stem cell; hiPSC-CMs: human induced pluripotent stem cell-derived cardiomyocytes; ISO: isoproterenol; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MTOR: mechanistic target of rapamycin kinase; PI3K: phosphoinositide 3-kinase; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; RT: room temperature; SB: SBI-0206965; ULK1: unc-51 like autophagy activating kinase 1.

Tumor-infiltrating myeloid cells are a prominent pro-tumorigenic immune cell population that limit host anti-tumor immunity and present a significant obstacle for many cancer immunotherapies. Targeting the mechanisms regulating myeloid cell function within the tumor microenvironment may overcome immunotherapy resistance in some cancers. Recent discoveries in the emerging field of immunometabolism reveal that the metabolic profiles of intratumoral myeloid cells are rewired to adapt to the nutrition-limited tumor microenvironment, and this shapes their pro-tumor phenotypes. Interestingly, metabolic modulation can shift these myeloid cells toward the immune-stimulating anti-tumor phenotype. In this review, we will highlight the roles of specific metabolic pathways in the activation and function of myeloid cells, and discuss the therapeutic value of metabolically reprogramming myeloid cells to augment and improve outcomes with cancer immunotherapy.

Introduction: As one of the major components of the tumor microenvironment, tumor-associated macrophages (TAMs) possess profound inhibitory activity against T cells and facilitate tumor escape from immune checkpoint blockade therapy. Converting this pro-tumorigenic toward the anti-tumorigenic phenotype thus is an important strategy for enhancing adaptive immunity against cancer. However, a plethora of mechanisms have been described for pro-tumorigenic differentiation in cancer, metabolic switches to program the anti-tumorigenic property of TAMs are elusive. Materials and methods: From an unbiased analysis of single-cell transcriptome data from multiple tumor models, we discovered that anti-tumorigenic TAMs uniquely express elevated levels of a specific fatty acid receptor, G-protein-coupled receptor 84 (GPR84). Genetic ablation of GPR84 in mice leads to impaired pro-inflammatory polarization of macrophages, while enhancing their anti-inflammatory phenotype. By contrast, GPR84 activation by its agonist, 6-n-octylaminouracil (6-OAU), potentiates pro-inflammatory phenotype via the enhanced STAT1 pathway. Moreover, 6-OAU treatment significantly retards tumor growth and increases the anti-tumor efficacy of anti-PD-1 therapy. Conclusion: Overall, we report a previously unappreciated fatty acid receptor, GPR84, that serves as an important metabolic sensing switch for orchestrating anti-tumorigenic macrophage polarization. Pharmacological agonists of GPR84 hold promise to reshape and reverse the immunosuppressive TME, and thereby restore responsiveness of cancer to overcome resistance to immune checkpoint blockade.