Ruifang Teng

Significance Immune checkpoint blockade (ICB) to reinvigorate cytotoxic T lymphocytes using antibodies against CTLA4 or PD1 generates durable therapeutic responses in patients across a variety of cancer types. However, some cancers such as castration-resistant prostate cancer (CRPC) show overwhelming resistance to ICB. Here, we develop a nitroproteomic approach and uncover a potential mechanism for the immunotherapy resistance where a key protein for T cell activation, lymphocyte-specific protein tyrosine kinase (LCK), is nitrated and inactivated by myeloid-derived suppressor cell–generated reactive nitrogen species (RNS) in the resistant tumor. An agent that neutralizes RNS significantly sensitizes CRPC to ICB therapy in a mouse model. Therefore, our studies illuminate a clinical path hypothesis for combining ICB with RNS-reducing agents in the treatment of ICB-resistant cancers.

HSP60 is a major mitochondrial chaperone for maintaining mitochondrial proteostasis. Our previous studies showed that HSP60 was significantly downregulated in clear cell renal cell carcinoma (ccRCC), the most common type of kidney cancer characterized by the classic Warburg effect. Here, we analyzed datasets in The Cancer Genome Atlas and revealed that higher HSP60 expression correlated with better overall survival in ccRCC patients. We also stably knocked down or overexpressed HSP60 in ccRCC cells to investigate the effects of HSP60 expression on the transition between oxidative phosphorylation and glycolysis. We confirmed that HSP60 knockdown increased cell proliferation, whereas its overexpression decreased cell growth. Proteomics and metabolomics revealed that HSP60 knockdown promoted Warburg-like phenotypes with enhanced glycolysis and decreased mitochondrial activity. Consistent with this finding, isotope tracing showed that the metabolic flow from glycolysis to TCA was reduced. However, HSP60 silencing enhanced mitochondrial functions in glutamine-directed biosynthesis with increased flow in two parts of the TCA cycle: Gln→αKG→OAA→Asp and Gln→αKG→ISO→acetyl-CoA, resulting in elevated de novo nucleotide synthesis and lipid synthesis. Proteomic analysis indicated that HSP60 silencing activated NRF2-mediated oxidative stress responses, while glutamate generated from glutamine increased glutathione synthesis for quenching excessive reactive oxygen species (ROS) produced upon elevated cell growth. We further found that HSP60 silencing activated the MEK/ERK/c-Myc axis to promote glutamine addiction, and confirmed that ccRCC cells were susceptible to oxidative stress and glutaminase inhibition. Collectively, our data show that HSP60 knockdown drives metabolic reprogramming in ccRCC to promote tumor progression and enhances mitochondrial-dependent biosynthesis.

In the early stages of infection, Human Immunodeficiency Virus Type 1 (HIV-1) generally selects CCR5 as the primary coreceptor for entering the host cell. As infection progresses, the virus evolves and may exhibit a coreceptor-switch to CXCR4. Accurate determination coreceptor usage and identification key mutational patterns associated tropism switch are essential for selection of appropriate therapies and understanding mechanism of coreceptor change. We developed a classifier composed of two coreceptor-specific weight matrices (CMs) based on a full-scale dataset. For this classifier, we found an AUC of 0.97, an accuracy of 95.21% and an MCC of 0.885 (sensitivity 92.92%; specificity 95.54%) in a ten-fold cross-validation, outperforming all other methods on an independent dataset (13% higher MCC value than geno2pheno and 15% higher MCC value than PSSM). A web server (http://spg.med.tsinghua.edu.cn/CM.html) based on our classifier was provided. Patterns of genetic mutations that occur along with coreceptor transitions were further identified based on the score of each sequence. Six pairs of one-AA mutational patterns and three pairs of two-AA mutational patterns were identified to associate with increasing propensity for X4 tropism. These mutational patterns offered new insights into the mechanism of coreceptor switch and aided in monitoring coreceptor switch.

Mesenchymal stem cells (MSCs) show great promise in blood vessel restoration and vascularization enhancement in many therapeutic situations. Typically, the co-implantation of MSCs with vascular endothelial cells (ECs) is effective for the induction of functional vascularization in vivo, indicating its potential applications in regenerative medicine. The effects of MSCs-ECs-induced vascularization can be modeled in vitro, providing simplified models for understanding their underlying communication. In this article, a contact coculture model in vitro and an RNA-seq approach were employed to reveal the active crosstalk between MSCs and ECs within a short time period at both morphological and transcriptional levels. The RNA-seq results suggested that angiogenic genes were significantly induced upon coculture, and this prevascularization commitment might require the NF-κB signaling. NF-κB blocking and interleukin (IL) neutralization experiments demonstrated that MSCs potentially secreted IL factors including IL1β and IL6 to modulate NF-κB signaling and downstream chemokines during coculture. Conversely, RNA-seq results indicated that the MSCs were regulated by the coculture environment to a smooth muscle commitment within this short period, which largely induced myocardin, the myogenic co-transcriptional factor. These findings demonstrate the mutual molecular mechanism of MSCs-ECs-induced prevascularization commitment in a quick response.