Cancer cells that succeed in forming metastasis need to be reprogrammed to evade immune surveillance and survive in a new microenvironment. This is facilitated by metastatic niches that are either postformed through reciprocal signaling between tumor cells and local stromal cells or preformed as premetastatic niches before tumor cell arrival. IL6/STAT3 signaling is aberrantly activated in lung tumorigenesis and metastasis, however, the roles and mechanisms of action of IL6 remain controversial. Here, we showed that blockade of intrinsic STAT3 signaling in lung tumor cells suppressed lung metastasis in immune-competent syngeneic mice, but not in immune-deficient nude mice. Consistently, repression of STAT3 signaling in tumor cells made them susceptible to T-cell-mediated cytotoxicity. Thus, STAT3mediated immunosuppression is crucial for metastasis. Noticeably, lung metastasis was greatly increased in Gprc5a-knockout (ko; 5a À/À ) mice compared with wild-type mice, which correlated with upre-gulated IL6 in the tumor microenvironment. Depletion of IL6 via combined deletion of Il6 and Gprc5a genes almost completely eliminated lung metastasis in Gprc5a-ko/Il6-ko (5a À/À ;Il6 À/À ) mice. Mechanistically, dysregulated IL6 reprogrammed the STAT3 pathway in metastatic tumor cells, and induced recruitment of myeloid-derived suppressor cells and polarized macrophages to evade host immunity. Consistently, IHC staining showed that activated STAT3 correlated with repressed infiltration of CD8 þ T cells in non-small cell lung cancer. Therefore, IL6/STAT3 signaling is crucial for orchestrating premetastatic niche formation and immunosuppression in lung.Significance: IL6 plays important roles not only in cell autonomous propensity for metastasis, but also in establishing the metastatic niche.

Chronic inflammation has been linked to promotion of tumorigenesis and metastasis in lung. However, due to lack of a relevant animal model for characterization, the underlying mechanism remains elusive. Lung tumor suppressor gene Gprc5a-knockout (ko) mice are susceptible to lung inflammation, tumorigenesis and metastasis, which resembles the pathological features in human patients. Here, we showed that PTGES/PGE 2 signaling was highly associated with lung tumorigenesis and metastasis in Gprc5a-ko mice. Interestingly, Ptges-knockout in mouse lung tumor cells, although reduced their stemness and EMT-like features, still formed tumors and lung metastasis in immune-deficient nude mice, but not in immune-competent mice. This suggests that the major role of PTGES/PGE 2 signaling in tumorigenicity and lung metastasis is through immunosuppression. Mechanistically, PTGES/PGE 2 signaling intrinsically endows tumor cells resistant to T-cell cytotoxicity, and induces cytokines extrinsically for MDSC recruitment, which is crucial for suppression of T-cell immunity. Importantly, targeting PGE 2 signaling in Gprc5a-ko mice by PTGES inhibitor suppressed MDSC recruitment, restored T cells, and significantly repressed lung metastasis. Thus, PTGES/PGE 2 signaling links immunosuppression and metastasis in an inflammatory lung microenvironment of Gprc5a-ko mouse model.

The major role of aldehyde dehydrogenase 2 family (ALDH2) is to detoxify acetaldehyde (ACE) to non-toxic acetic acid. Many evidences suggest that ALDH2 dysfunction contributes to a variety of human diseases including cancer. However, the biological function and molecular mechanism of ALDH2 in tumor progression remain elusive. In this study, we found that ALDH2 repression was associated with poor prognosis in lung adenocarcinoma. Overexpression of ALDH2 inhibited malignant features of lung adenocarcinoma cells, such as proliferation, stemness and migration, whereas ALDH2 knockdown increased these features. Mechanistically, ALDH2 repression led to accumulation of ACE; whereas ACE enhanced the migration features of lung adenocarcinoma cells, which was associated with increased DNA damage. Importantly, accumulated ACE and increased DNA damage were identified in Aldh2-knockout (KO) mouse lung tissues in vivo. Consistent with this concept, treatment of lung adenocarcinoma cells with ALDH2 agonist Alda-1 suppressed the proliferation, stemness and migration features of lung adenocarcinoma cells. Thus, activating ALDH2, such as via its agonist, may provide a novel strategy for treatment of lung cancer.

Cancer cells need to generate large amounts of glutathione (GSH) to buffer oxidative stress during tumor development. A rate-limiting step for GSH biosynthesis is cystine uptake via a cystine/glutamate antiporter Xc-. Xc-is a sodium-independent antiporter passively driven by concentration gradients from extracellular cystine and intracellular glutamate across the cell membrane. Increased uptake of cystine via Xc-in cancer cells increases the level of extracellular glutamate, which would subsequently restrain cystine uptake via Xc-. Cancer cells must therefore evolve a mechanism to overcome this negative feedback regulation. In this study, we report that glutamate transporters, in particular SLC1A1, are tightly intertwined with cystine uptake and GSH biosynthesis in lung cancer cells. Dysregulated SLC1A1, a sodium-dependent glutamate carrier, actively recycled extracellular glutamate into cells, which enhanced the efficiency of cystine uptake via Xc-and GSH biosynthesis as measured by stable isotope-assisted metabolomics. Conversely, depletion of glutamate transporter SLC1A1 increased extracellular glutamate, which inhibited cystine uptake, blocked GSH synthesis, and induced oxidative stress-mediated cell death or growth inhibition. Moreover, glutamate transporters were frequently upregulated in tissue samples of nonsmall cell lung cancer patients. Taken together, active uptake of glutamate via SLC1A1 propels cystine uptake via Xc-for GSH biosynthesis in lung tumorigenesis. Significance: Cellular GSH in cancer cells is not only determined by upregulated Xc-but also by dysregulated glutamate transporters, which provides additional targets for therapeutic intervention. Research.

Iron dysregulation is associated with several diseases, including lung cancer, but the underlying mechanism is yet unknown. Iron directly binds CDK1, which is upregulated in several cancers, thereby promoting JAK1 phosphorylation and activation of STAT3 signaling to promote colorectal carcinogenesis. This study aimed to investigate the role of iron/CDK1/STAT3 signaling in lung carcinogenesis. We found that iron-dependent CDK1 activity upregulated IL-6 receptor subunit GP130 post-transcriptionally via phosphorylation of 4E-BP1, which is critical for activation of JAK/STAT3 signaling. CDK1 and STAT3 are essential for iron-mediated colony formation in lung cancer cell lines. CDK1 knockdown and iron chelator DFO decreased tumorigenicity and GP130/STAT3 signaling in vivo. Moreover, CDK1/GP130/STAT3 signaling were elevated in lung cancer tissues compared with adjacent normal lung tissues. Altogether, the present results suggest that CDK1 inhibition and iron deprivation are potential strategies to target GP130/STAT3 signaling to suppress lung cancer.

GPRC5A, a retinoic acid induced gene, is preferentially expressed in lung tissue. Gprc5a gene deletion leads to spontaneous lung tumor development. However, the mechanism of Gprc5a-mediated lung tumor suppression is not fully understood. Here we showed that MDM2, a p53-negative regulator, was dysregulated in Gprc5a-knockout (ko) mouse tracheal epithelial cells (KO-MTEC) compared to wild type ones. Targeting MDM2 in 1601-a Gprc5a-ko mouse derived lung tumor cell line-and A549-human lung cancer cells, by MDM2 inhibitor Nutlin-3a or small hairpin RNA (sh-RNA)-restored p53 signaling pathway, reduced cancer stem cell markers, and inhibited tumorigenicity. This suggests that dysregulated MDM2 pathway is essential for the oncogenic activities of these cells. MDM2 was found to be stabilized mainly by activated EGFR signaling as targeting EGFR by Erlotinib or sh-RNA repressed MDM2 in a transcription-independent manner. Importantly, overexpression of MDM2 and reduced GPRC5A expression at both protein and mRNA levels were frequently found in clinical human lung cancer tissues. Taken together, GPRC5A deficiency contributes to dysregulated MDM2 via activated EGFR signaling, which promotes lung tumor development.

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Lung squamous cell carcinomas (SCCs) are highly aggressive tumors, and there is currently no effective targeted therapy owing to the lack of specific mutation targets. Compared with lung adenocarcinoma (ADCs), lung SCCs reportedly utilized higher levels of glucose metabolism to meet the anabolic and catabolic needs required to sustain rapid tumor growth. Hexokinase 2 (HK2) is an enzyme that catalyzes the rate-limit and first committed step in glucose metabolism. Here, we investigated the expression and effect of HK2 in lung SCCs. We found a significantly higher HK2 expression in lung SCCs, but not lung ADC or normal tissues. HK2 depletion or inhibition decreased the glycolysis and tumor growth via activating AMPK signaling pathway, which downregulated mTORC1 activity. Furthermore, we found an increased oxygen respiration rate compensating for HK2 depletion. Thus, metformin treatment showed combinatorial therapeutic value, which resulted in greater induction of lung SCC apoptosis in vitro and in vivo. Our study suggests that HK2 depletion in combination with metformin might be a novel effective strategy for lung SCCs therapy.