Programmed death-ligand 1 (PD-L1) and its receptor programmed cell death protein 1 (PD-1) modulate antitumor immunity and are major targets of checkpoint blockade immunotherapy. However, clinical trials of anti-PD-L1 and anti-PD-1 antibodies in breast cancer demonstrate only modest efficacy. Furthermore, specific PD-L1 contributions in various tissue and cell compartments to antitumor immunity remain incompletely elucidated. Here we show that PD-L1 expression is markedly elevated in mature adipocytes versus preadipocytes. Adipocyte PD-L1 prevents anti-PD-L1 antibody from activating important antitumor functions of CD8+ T cells in vitro. Adipocyte PD-L1 ablation obliterates, whereas forced preadipocyte PD-L1 expression confers, these inhibitory effects. Pharmacologic inhibition of adipogenesis selectively reduces PD-L1 expression in mouse adipose tissue and enhances the antitumor efficacy of anti-PD-L1 or anti-PD-1 antibodies in syngeneic mammary tumor models. Our findings provide a previously unappreciated approach to bolster anticancer immunotherapy efficacy and suggest a mechanism for the role of adipose tissue in breast cancer progression.

Energy metabolism follows a diurnal pattern responding to the cycles of light and food exposures. Although food availability is a potent synchronizer of peripheral circadian clock in mammals, the underlying mechanism remains elusive. Here, we found that the temporal signals of fasting and refeeding hormones regulate the transcription of Bmal1, a key transcription activator of molecular clock, in the liver. During fasting, glucagon, a major fasting hormone, activates CREB/CRTC2 transcriptional complex that is recruited to Bmal1 promoter to induce its expression. Furthermore, we showed that CRTC2 is required for basal transcriptional regulation of Bmal1 by experiments using either adenovirus-mediated CRTC2 RNAi knockdown or primary Crtc2 null hepatocytes. On the other hand, insulin suppresses fasting-induced Bmal1 expression by inhibiting CRTC2 activity after refeeding. Taken together, our results indicate CRTC2 as a key component of the circadian oscillator that integrates the mammalian clock and energy metabolism.

The rapid proliferation of cancer cells and dysregulated vasculature within the tumor leads to limited nutrient accessibility. Cancer cells often rewire their metabolic pathways for adaption to nutrient stress, and the underlying mechanism remains largely unknown. Glutamate dehydrogenase 1 (GDH1) is a key enzyme in glutaminolysis that converts glutamate to α-ketoglutarate (α-KG). Here, we show that, under low glucose, GDH1 is phosphorylated at serine (S) 384 and interacts with RelA and IKKβ. GDH1-produced α-KG directly binds to and activates IKKβ and nuclear factor κB (NF-κB) signaling, which promotes glucose uptake and tumor cell survival by upregulating GLUT1, thereby accelerating gliomagenesis. In addition, GDH1 S384 phosphorylation correlates with the malignancy and prognosis of human glioblastoma. Our finding reveals a unique role of α-KG to directly regulate signal pathway, uncovers a distinct mechanism of metabolite-mediated NF-κB activation, and also establishes the critical role of α-KG-activated NF-κB in brain tumor development.

6-Phosphogluconate dehydrogenase (6PGD) is a key enzyme that converts 6-phosphogluconate into ribulose-5-phosphate with NADP+ as cofactor in the pentose phosphate pathway (PPP). 6PGD is commonly upregulated and plays important roles in many human cancers, while the mechanism underlying such roles of 6PGD remains elusive. Here we show that upon EGFR activation, 6PGD is phosphorylated at tyrosine (Y) 481 by Src family kinase Fyn. This phosphorylation enhances 6PGD activity by increasing its binding affinity to NADP+ and therefore activates the PPP for NADPH and ribose-5-phosphate, which consequently detoxifies intracellular reactive oxygen species (ROS) and accelerates DNA synthesis. Abrogating 6PGD Y481 phosphorylation (pY481) dramatically attenuates EGF-promoted glioma cell proliferation, tumor growth and resistance to ionizing radiation. In addition, 6PGD pY481 is associated with Fyn expression, the malignancy and prognosis of human glioblastoma. These findings establish a critical role of Fyn-dependent 6PGD phosphorylation in EGF-promoted tumor growth and radiation resistance.

Although food availability is a potent synchronizer of the peripheral circadian clock in mammals, the underlying mechanisms are unclear. Here, we show that hepatic Bmal1, a core transcription activator of the molecular clock, is post-transcriptionally regulated by signals from insulin, an important hormone that is temporally controlled by feeding. Insulin promotes postprandial Akt-mediated Ser42-phosphorylation of Bmal1 to induce its dissociation from DNA, interaction with 14-3-3 protein and subsequently nuclear exclusion, which results in the suppression of Bmal1 transcriptional activity. Inverted feeding cycles not only shift the phase of daily insulin oscillation, but also elevate the amplitude due to food overconsumption. This enhanced and reversed insulin signalling initiates the reset of clock gene rhythms by altering Bmal1 nuclear accumulation in mouse liver. These results reveal the molecular mechanism of insulin signalling in regulating peripheral circadian rhythms.

The Unc-51 like autophagy activating kinase 1 (ULK1) complex plays a central role in the initiation stage of autophagy. However, the function of ULK1 in the late stage of autophagy is unknown. Here, we report that ULK1, a central kinase of the ULK1 complex involved in autophagy initiation, promotes autophagosome-lysosome fusion. PKCα phosphorylates ULK1 and prevents autolysosome formation. PKCα phosphorylation of ULK1 does not change its kinase activity; however, it decreases autophagosome-lysosome fusion by reducing the affinity of ULK1 for syntaxin 17 (STX17). Unphosphorylated ULK1 recruited STX17 and increased STX17's affinity towards synaptosomal-associated protein 29 (SNAP29). Additionally, phosphorylation of ULK1 enhances its interaction with heat shock cognate 70 kDa protein (HSC70) and increases its degradation through chaperone-mediated autophagy (CMA). Our study unearths a key mechanism underlying autolysosome formation, a process in which the kinase activity of PKCα plays an instrumental role, and reveals the significance of the mutual regulation of macroautophagy and CMA in maintaining the balance of autophagy.

Macrophages are a dominant leukocyte population in the tumor microenvironment and actively promote cancer progression. However, the molecular mechanism underlying the role of macrophages remains poorly understood. Here we show that polarized M2 macrophages enhance 3-phosphoinositide-dependent protein kinase 1 (PDPK1)-mediated phosphoglycerate kinase 1 (PGK1) threonine (T) 243 phosphorylation in tumor cells by secreting interleukin-6 (IL-6). This phosphorylation facilitates a PGK1-catalyzed reaction toward glycolysis by altering substrate affinity. Inhibition of PGK1 T243 phosphorylation or PDPK1 in tumor cells or neutralization of macrophage-derived IL-6 abrogates macrophage-promoted glycolysis, proliferation, and tumorigenesis. In addition, PGK1 T243 phosphorylation correlates with PDPK1 activation, IL-6 expression, and macrophage infiltration in human glioblastoma multiforme (GBM). Moreover, PGK1 T243 phosphorylation also correlates with malignance and prognosis of human GBM. Our findings demonstrate a novel mechanism of macrophage-promoted tumor growth by regulating tumor cell metabolism, implicating the therapeutic potential to disrupt the connection between macrophages and tumor cells by inhibiting PGK1 phosphorylation.

Anti-programmed death-ligand 1 (αPD-L1) immunotherapy is approved to treat bladder cancer (BC) but is effective in <30% of patients. Interleukin (IL)-2/αIL-2 complexes (IL-2c) that preferentially target IL-2 receptor β (CD122) augment CD8+ antitumor T cells known to improve αPD-L1 efficacy. We hypothesized that the tumor microenvironment, including local immune cells in primary versus metastatic BC, differentially affects immunotherapy responses and that IL-2c effects could differ from, and thus complement αPD-L1.

Tumor cell-intrinsic programmed death-ligand 1 (PD-L1) signals mediate immunopathologic effects in breast, colon, and ovarian cancers and in melanomas, but bladder cancer (BC) effects are unreported. We show here that BC cell-intrinsic PD-L1 signals in mouse MB49 and human RT4, UM-UC3, and UM-UC-14 BC cells regulate important pathologic pathways and processes, including effects not reported in other cancers. α-PD-L1 antibodies reduced BC cell proliferation in vitro, demonstrating direct signaling effects. BC cell-intrinsic PD-L1 promoted mammalian target of rapamycin complex 1 (mTORC1) signals in vitro and augmented in vivo immune-independent cell growth and metastatic cancer spread, similar to effects we reported in melanoma and ovarian cancer. BC cell-intrinsic PD-L1 signals also promoted basal and stress-induced autophagy, whereas these signals inhibited autophagy in melanoma and ovarian cancer cells. BC cell-intrinsic PD-L1 also mediated chemotherapy resistance to the commonly used BC chemotherapy agents cis-platinum and gemcitabine and to the mTORC1 inhibitor, rapamycin. Thus, BC cell-intrinsic PD-L1 signals regulate important virulence and treatment resistance pathways that suggest novel, actionable treatment targets meriting additional studies. As a proof-of-concept, we showed that the autophagy inhibitor chloroquine improved cis-platinum treatment efficacy in vivo, with greater efficacy in PD-L1 null versus PD-L1-replete BC.