The Hippo pathway noncanonically drives autophagy and cell survival in response to energy stress.

The Hippo pathway is known for its crucial involvement in development, regeneration, organ size control, and cancer. While energy stress is known to activate the Hippo pathway and inhibit its effector YAP, the precise role of the Hippo pathway in energy stress response remains unclear. Here, we report a YAP-independent function of the Hippo pathway in facilitating autophagy and cell survival in response to energy stress, a process mediated by its upstream components MAP4K2 and STRIPAK. Mechanistically, energy stress disrupts the MAP4K2-STRIPAK association, leading to the activation of MAP4K2. Subsequently, MAP4K2 phosphorylates ATG8-family member LC3, thereby facilitating autophagic flux. MAP4K2 is highly expressed in head and neck cancer, and its mediated autophagy is required for head and neck tumor growth in mice. Altogether, our study unveils a noncanonical role of the Hippo pathway in energy stress response, shedding light on this key growth-related pathway in tissue homeostasis and cancer.

IL-1ß-associated NNT acetylation orchestrates iron-sulfur cluster maintenance and cancer immunotherapy resistance.

Interleukin-1ß (IL-1ß) is a key protein in inflammation and contributes to tumor progression. However, the role of IL-1ß in cancer is ambiguous or even contradictory. Here, we found that upon IL-1ß stimulation, nicotinamide nucleotide transhydrogenase (NNT) in cancer cells is acetylated at lysine (K) 1042 (NNT K1042ac) and thereby induces the mitochondrial translocation of p300/CBP-associated factor (PCAF). This acetylation enhances NNT activity by increasing the binding affinity of NNT for NADP+ and therefore boosts NADPH production, which subsequently sustains sufficient iron-sulfur cluster maintenance and protects tumor cells from ferroptosis. Abrogating NNT K1042ac dramatically attenuates IL-1ß-promoted tumor immune evasion and synergizes with PD-1 blockade. In addition, NNT K1042ac is associated with IL-1ß expression and the prognosis of human gastric cancer. Our findings demonstrate a mechanism of IL-1ß-promoted tumor immune evasion, implicating the therapeutic potential of disrupting the link between IL-1ß and tumor cells by inhibiting NNT acetylation.

LncRNA CamK-A Regulates Ca2+-Signaling-Mediated Tumor Microenvironment Remodeling.

Cancer cells entail metabolic adaptation and microenvironmental remodeling to survive and progress. Both calcium (Ca2+) flux and Ca2+-dependent signaling play a crucial role in this process, although the underlying mechanism has yet to be elucidated. Through RNA screening, we identified one long noncoding RNA (lncRNA) named CamK-A (lncRNA for calcium-dependent kinase activation) in tumorigenesis. CamK-A is highly expressed in multiple human cancers and involved in cancer microenvironment remodeling via activation of Ca2+-triggered signaling. Mechanistically, CamK-A activates Ca2+/calmodulin-dependent kinase PNCK, which in turn phosphorylates IκBα and triggers calcium-dependent nuclear factor κB (NF-κB) activation. This regulation results in the tumor microenvironment remodeling, including macrophage recruitment, angiogenesis, and tumor progression. Notably, our human-patient-derived xenograft (PDX) model studies demonstrate that targeting CamK-A robustly impaired cancer development. Clinically, CamK-A expression coordinates with the activation of CaMK-NF-κB axis, and its high expression indicates poor patient survival rate, suggesting its role as a potential biomarker and therapeutic target.

dSCOPE: a software to detect sequences critical for liquid-liquid phase separation.

Membrane-based cells are the fundamental structural and functional units of organisms, while evidences demonstrate that liquid-liquid phase separation (LLPS) is associated with the formation of membraneless organelles, such as P-bodies, nucleoli and stress granules. Many studies have been undertaken to explore the functions of protein phase separation (PS), but these studies lacked an effective tool to identify the sequence segments that critical for LLPS. In this study, we presented a novel software called dSCOPE (http://dscope.omicsbio.info) to predict the PS-driving regions. To develop the predictor, we curated experimentally identified sequence segments that can drive LLPS from published literature. Then sliding sequence window based physiological, biochemical, structural and coding features were integrated by random forest algorithm to perform prediction. Through rigorous evaluation, dSCOPE was demonstrated to achieve satisfactory performance. Furthermore, large-scale analysis of human proteome based on dSCOPE showed that the predicted PS-driving regions enriched various protein post-translational modifications and cancer mutations, and the proteins which contain predicted PS-driving regions enriched critical cellular signaling pathways. Taken together, dSCOPE precisely predicted the protein sequence segments critical for LLPS, with various helpful information visualized in the webserver to facilitate LLPS-related research.

Methionine deficiency facilitates antitumour immunity by altering m6A methylation of immune checkpoint transcripts.

OBJECTIVE: Methionine metabolism is involved in a myriad of cellular functions, including methylation reactions and redox maintenance. Nevertheless, it remains unclear whether methionine metabolism, RNA methylation and antitumour immunity are molecularly intertwined. DESIGN: The antitumour immunity effect of methionine-restricted diet (MRD) feeding was assessed in murine models. The mechanisms of methionine and YTH domain-containing family protein 1 (YTHDF1) in tumour immune escape were determined in vitro and in vivo. The synergistic effects of MRD or YTHDF1 depletion with PD-1 blockade were also investigated. RESULTS: We found that dietary methionine restriction reduced tumour growth and enhanced antitumour immunity by increasing the number and cytotoxicity of tumour-infiltrating CD8+ T cells in different mouse models. Mechanistically, the S-adenosylmethionine derived from methionine metabolism promoted the N6-methyladenosine (m6A) methylation and translation of immune checkpoints, including PD-L1 and V-domain Ig suppressor of T cell activation (VISTA), in tumour cells. Furthermore, MRD or m6A-specific binding protein YTHDF1 depletion inhibited tumour growth by restoring the infiltration of CD8+ T cells, and synergised with PD-1 blockade for better tumour control. Clinically, YTHDF1 expression correlated with poor prognosis and immunotherapy outcomes for cancer patients. CONCLUSIONS: Methionine and YTHDF1 play a critical role in anticancer immunity through regulating the functions of T cells. Targeting methionine metabolism or YTHDF1 could be a potential new strategy for cancer immunotherapy.

Monitoring tumour resistance to the BRAF inhibitor combination regimen in colorectal cancer patients via circulating tumour DNA.

AIMS: This study aimed to identify mechanisms of drug resistance to the combination of vemurafenib, irinotecan, and cetuximab (VIC) in BRAFV600E metastatic colorectal cancer (mCRC). METHODS: Forty-one patients with BRAFV600E mCRC from July 2018 and June 2020 were evaluated, with tissue and/or plasma samples collected. We profiled tissue and plasma samples using whole-exome sequencing and targeted sequencing of 425 cancer-relevant genes. Clinical cohort analysis from published studies was performed to consolidate our findings. RESULTS: BRAF mutant in baseline plasma and its dynamics are significantly associated with VIC-related response, and concurrent RNF43 mutation significantly sensitises tumour to VIC treatment. VIC resistance frequently involves genes in PI3K, MAPK pathway, and several novel resistance mechanisms such as TGFBR2 and SMAD4 mutations, and copy-number gains in PTK2, MYC, and GATA6 have been identified. We also firstly describe acquired altered genes in DNA damaging repair pathway, occurring in 33 % of patients after VIC treatment, and particularly, patients with this pre-treatment resistance subclones developed inferior responses, along with higher tumour mutation burden both at baseline and progression plasma. CONCLUSION: Analysis of ctDNA can provide novel insights into molecular resistance mechanisms to VIC in BRAFV600E mCRC patients, allowing accurate guidance for clinicians in personalised treatment strategies.

Novel Genetic and Epigenetic Biomarkers of Prognostic and Predictive Significance in Stage II/III Colorectal Cancer.

The therapeutic strategies of stage II/III colorectal cancer (CRC) patients after curative surgery remain controversial. In the clinical decision-making process, oncologists need to answer questions such as whether adjuvant chemotherapy is necessary or which therapeutic regimen should be given to each patient. At present, whether adjuvant chemotherapy should be applied is primarily based on histopathological features and clinical risk factors. However, only a fraction of patients can benefit from it. More rigorous stratifying biomarkers are urgently needed to help further distinguishing these populations of patients. Recent progress in next-generation sequencing and high-throughput technologies has greatly promoted biomarker discovery as well as our understanding of the underlying mechanisms in CRC. Novel genetic and epigenetic biomarkers that are associated with prognosis or therapeutic responses have emerged. In this review, we discuss the strategies of biomarker discovery and summarize the status and assess the utility of previously published biomarkers in CRC.

Performance of common genetic variants in risk prediction for colorectal cancer in Chinese: A two-stage and multicenter study.

The efficacy of susceptible variants derived from genome-wide association studies (GWAs) optimizing discriminatory accuracy of colorectal cancer (CRC) in Chinese remains unclear. In the present validation study, we assessed 75 recently identified variants from GWAs. A risk predictive model combining 19 variants using the least absolute shrinkage and selection operator (LASSO) statistics offered certain clinical advantages. This model demonstrated an area under the receiver operating characteristic (AUC) of 0.61 during training analysis and yielded robust AUCs from 0.59 to 0.61 during validation analysis in three independent centers. The individuals carrying the highest quartile of risk score revealed over 2-fold risks of CRC (ranging from 2.12 to 2.90) compared with those who presented the lowest quartile of risk score. This genetic model offered the possibility of partitioning risk within the average risk population, which might serve as a first step toward developing individualized CRC prevention strategies in China.

College of American Pathologists Tumor Regression Grading System for Long-Term Outcome in Patients with Locally Advanced Rectal Cancer.

BACKGROUND: The National Comprehensive Cancer Network's Rectal Cancer Guideline Panel recommends American Joint Committee of Cancer and College of American Pathologists (AJCC/CAP) tumor regression grading (TRG) system to evaluate pathologic response to neoadjuvant chemoradiotherapy for locally advanced rectal cancer (LARC). Yet, the clinical significance of the AJCC/CAP TRG system has not been fully defined. MATERIALS AND METHODS: This was a multicenter, retrospectively recruited, and prospectively maintained cohort study. Patients with LARC from one institution formed the discovery set, and cases from external independent institutions formed a validation set to verify the findings from discovery set. Overall survival (OS), disease-free survival (DFS), local recurrence-free survival (LRFS), and distant metastasis-free survival (DMFS) were assessed by Kaplan-Meier analysis, log-rank test, and Cox regression model. RESULTS: The discovery set (940 cases) found, and the validation set (2,156 cases) further confirmed, that inferior AJCC/CAP TRG categories were closely /ccorrelated with unfavorable survival (OS, DFS, LRFS, and DMFS) and higher risk of disease progression (death, accumulative relapse, local recurrence, and distant metastasis) (all p < .05). Significantly, pairwise comparison revealed that any two of four TRG categories had the distinguished survival and risk of disease progression. After propensity score matching, AJCC/CAP TRG0 category (pathological complete response) patients treated with or without adjuvant chemotherapy displayed similar survival of OS, DFS, LRFS, and DMFS (all p > .05). For AJCC/CAP TRG1-3 cases, adjuvant chemotherapy treatment significantly improved 3-year OS (90.2% vs. 84.6%, p < .001). Multivariate analysis demonstrated the AJCC/CAP TRG system was an independent prognostic surrogate. CONCLUSION: AJCC/CAP TRG system, an accurate prognostic surrogate, appears ideal for further strategizing adjuvant chemotherapy for LARC. IMPLICATIONS FOR PRACTICE: The National Comprehensive Cancer Network recommends the American Joint Committee of Cancer and College of American Pathologists (AJCC/CAP) tumor regression grading (TRG) four-category system to evaluate the pathologic response to neoadjuvant treatment for patients with locally advanced rectal cancer; however, the clinical significance of the AJCC/CAP TRG system has not yet been clearly addressed. This study found, for the first time, that any two of four AJCC/CAP TRG categories had the distinguished long-term survival outcome. Importantly, adjuvant chemotherapy may improve the 3-year overall survival for AJCC/CAP TRG1-3 category patients but not for AJCC/CAP TRG0 category patients. Thus, AJCC/CAP TRG system, an accurate surrogate of long-term survival outcome, is useful in guiding adjuvant chemotherapy management for rectal cancer.

Long Noncoding RNA p53-Stabilizing and Activating RNA Promotes p53 Signaling by Inhibiting Heterogeneous Nuclear Ribonucleoprotein K deSUMOylation and Suppresses Hepatocellular Carcinoma.

To identify hepatocellular carcinoma (HCC)-implicated long noncoding RNAs (lncRNAs), we performed an integrative omics analysis by integrating mRNA and lncRNA expression profiles in HCC tissues. We identified a collection of candidate HCC-implicated lncRNAs. Among them, we demonstrated that an lncRNA, which is named as p53-stabilizing and activating RNA (PSTAR), inhibits HCC cell proliferation and tumorigenicity through inducing p53-mediated cell cycle arrest. We further revealed that PSTAR can bind to heterogeneous nuclear ribonucleoprotein K (hnRNP K) and enhance its SUMOylation and thereby strengthen the interaction between hnRNP K and p53, which ultimately leads to the accumulation and transactivation of p53. PSTAR is down-regulated in HCC tissues, and the low PSTAR expression predicts poor prognosis in patients with HCC, especially those with wild-type p53. Conclusion: This study sheds light on the tumor suppressor role of lncRNA PSTAR, a modulator of the p53 pathway, in HCC.

Circular RNA: metabolism, functions and interactions with proteins.

Circular RNAs (CircRNAs) are single-stranded, covalently closed RNA molecules that are ubiquitous across species ranging from viruses to mammals. Important advances have been made in the biogenesis, regulation, localization, degradation and modification of circRNAs. CircRNAs exert biological functions by acting as transcriptional regulators, microRNA (miR) sponges and protein templates. Moreover, emerging evidence has revealed that a group of circRNAs can serve as protein decoys, scaffolds and recruiters. However, the existing research on circRNA-protein interactions is quite limited. Hence, in this review, we briefly summarize recent progress in the metabolism and functions of circRNAs and elaborately discuss the patterns of circRNA-protein interactions, including altering interactions between proteins, tethering or sequestering proteins, recruiting proteins to chromatin, forming circRNA-protein-mRNA ternary complexes and translocating or redistributing proteins. Many discoveries have revealed that circRNAs have unique expression signatures and play crucial roles in a variety of diseases, enabling them to potentially act as diagnostic biomarkers and therapeutic targets. This review systematically evaluates the roles and mechanisms of circRNAs, with the hope of advancing translational medicine involving circRNAs.

Association between adjuvant chemotherapy and survival in patients with rectal cancer and pathological complete response after neoadjuvant chemoradiotherapy and resection.

BACKGROUND: For patients with locally advanced rectal cancer (LARC), it is unclear whether neoadjuvant chemoradiotherapy-induced pathologic complete response (pCR) individuals would further benefit from adjuvant chemotherapy (ACT). METHODS: The pCR individuals who received different ACT cycles were paired by propensity score matching. Overall survival (OS), disease-free survival (DFS), local recurrence-free survival (LRFS), and distant metastasis-free survival (DMFS) were calculated by Kaplan-Meier and log-rank test. RESULTS: In total, 1041 pCR individuals were identified from 5567 LARC cases. Specifically, 303 pCR cases had no ACT treatment, and 738 pCR patients received fluoropyrimidine-based ACT (median, 4 cycles) treatment. After 1:3 propensity score matching, 297 cases without ACT treatment were matched to 712 cases who received ACT treatment. Kaplan-Meier analysis showed that pCR individuals treated with or without ACT had the similar 3-year outcome (OS, DFS, LRFS and DMFS) (all P > 0.05). Moreover, the pCR patients received different ACT cycle(s) (0 vs. 1-4 cycles, 0 vs. ≥5 cycles) had comparable 3-year OS, DFS, LRFS and DMFS (all P > 0.05). In stratified analysis, ACT treatment did not improve 3-year survival (OS, DFS, LRFS and DMFS) for the baseline high-risk (cT3-4/cN1-2) subgroup patients (all P > 0.05). CONCLUSION: ACT, which did not improve survival, is unnecessary to neoadjuvant treatment-induced pCR LARC patients. TRIAL REGISTRATION: 2019ZSLYEC-136 (24-6-2019).

METTL3 facilitates tumor progression via an m6A-IGF2BP2-dependent mechanism in colorectal carcinoma.

BACKGROUND: Colorectal carcinoma (CRC) is one of the most common malignant tumors, and its main cause of death is tumor metastasis. RNA N6-methyladenosine (m6A) is an emerging regulatory mechanism for gene expression and methyltransferase-like 3 (METTL3) participates in tumor progression in several cancer types. However, its role in CRC remains unexplored. METHODS: Western blot, quantitative real-time PCR (RT-qPCR) and immunohistochemical (IHC) were used to detect METTL3 expression in cell lines and patient tissues. Methylated RNA immunoprecipitation sequencing (MeRIP-seq) and transcriptomic RNA sequencing (RNA-seq) were used to screen the target genes of METTL3. The biological functions of METTL3 were investigated in vitro and in vivo. RNA pull-down and RNA immunoprecipitation assays were conducted to explore the specific binding of target genes. RNA stability assay was used to detect the half-lives of the downstream genes of METTL3. RESULTS: Using TCGA database, higher METTL3 expression was found in CRC metastatic tissues and was associated with a poor prognosis. MeRIP-seq revealed that SRY (sex determining region Y)-box 2 (SOX2) was the downstream gene of METTL3. METTL3 knockdown in CRC cells drastically inhibited cell self-renewal, stem cell frequency and migration in vitro and suppressed CRC tumorigenesis and metastasis in both cell-based models and PDX models. Mechanistically, methylated SOX2 transcripts, specifically the coding sequence (CDS) regions, were subsequently recognized by the specific m6A "reader", insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2), to prevent SOX2 mRNA degradation. Further, SOX2 expression positively correlated with METTL3 and IGF2BP2 in CRC tissues. The combined IHC panel, including "writer", "reader", and "target", exhibited a better prognostic value for CRC patients than any of these components individually. CONCLUSIONS: Overall, our study revealed that METTL3, acting as an oncogene, maintained SOX2 expression through an m6A-IGF2BP2-dependent mechanism in CRC cells, and indicated a potential biomarker panel for prognostic prediction in CRC.

LncRNA LINRIS stabilizes IGF2BP2 and promotes the aerobic glycolysis in colorectal cancer.

BACKGROUND: Long noncoding RNAs (lncRNAs) play nonnegligible roles in the epigenetic regulation of cancer cells. This study aimed to identify a specific lncRNA that promotes the colorectal cancer (CRC) progression and could be a potential therapeutic target. METHODS: We screened highly expressed lncRNAs in human CRC samples compared with their matched adjacent normal tissues. The proteins that interact with LINRIS (Long Intergenic Noncoding RNA for IGF2BP2 Stability) were confirmed by RNA pull-down and RNA immunoprecipitation (RIP) assays. The proliferation and metabolic alteration of CRC cells with LINRIS inhibited were tested in vitro and in vivo. RESULTS: LINRIS was upregulated in CRC tissues from patients with poor overall survival (OS), and LINRIS inhibition led to the impaired CRC cell line growth. Moreover, knockdown of LINRIS resulted in a decreased level of insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2), a newly found N6-methyladenosine (m6A) 'reader'. LINRIS blocked K139 ubiquitination of IGF2BP2, maintaining its stability. This process prevented the degradation of IGF2BP2 through the autophagy-lysosome pathway (ALP). Therefore, knockdown of LINRIS attenuated the downstream effects of IGF2BP2, especially MYC-mediated glycolysis in CRC cells. In addition, the transcription of LINRIS could be inhibited by GATA3 in CRC cells. In vivo experiments showed that the inhibition of LINRIS suppressed the proliferation of tumors in orthotopic models and in patient-derived xenograft (PDX) models. CONCLUSION: LINRIS is an independent prognostic biomarker for CRC. The LINRIS-IGF2BP2-MYC axis promotes the progression of CRC and is a promising therapeutic target.

Hepatitis B virus infection is associated with younger median age at diagnosis and death in cancers.

Several non-hepatocellular cancers were linked with hepatitis B virus (HBV) infection. This study was aimed to quantify the potential associations between HBV infection and multiple non-hepatocellular cancers. Continuous cases, including 5,715 non-cancer and 40,963 cancer cases diagnosed from 2008 to 2014 in Sun Yat-sen University Cancer Center were analyzed. HBV DNA and hepatitis B core antigen (HBcAg) were examed in gastric cancer tissues by polymerase chain reaction and immunohistochemical staining. After adjusting for age, sex, year of diagnosis, smoking, drinking and family history of cancer, significant associations were found between serum HBsAg and frequently reported HBV-related non-hepatocellular cancers, including non-Hodgkin's lymphoma, cholangiocarcinoma and pancreatic cancer [adjusted odds ratio (AOR) and 95% confidence interval (CI): 1.89 (1.65-2.16)], as well as total other non-hepatocellular cancers [AOR and 95% CI: 1.12 (1.03-1.22)]. The median ages at diagnosis, all-cause death and cancer-specific death of serum HBsAg positive cancer patients were all significantly younger than those with serum HBsAg negative. HBV DNA was detected in 12.4% (34/275) gastric cancer tissues and HBcAg was most commonly detected in lymphocytes. This was the first report that HBV infection had a modest but significant nonspecific association with total non-hepatocellular cancers. Median age at diagnosis and death was significantly younger in serum HBsAg positive cancer patients. The underlying mechanism needs further investigation.

Melatonin overcomes gemcitabine resistance in pancreatic ductal adenocarcinoma by abrogating nuclear factor-κB activation.

Constitutive activation and gemcitabine induction of nuclear factor-κB (NF-κB) contribute to the aggressive behavior and chemotherapeutic resistance of pancreatic ductal adenocarcinoma (PDAC). Thus, targeting the NF-κB pathway has proven an insurmountable challenge for PDAC therapy. In this study, we investigated whether the inhibition of NF-κB signaling pathway by melatonin might lead to tumor suppression and overcome gemcitabine resistance in pancreatic tumors. Our results showed that melatonin inhibited activities of NF-κB by suppressing IκBα phosphorylation and decreased the expression of NF-κB response genes in MiaPaCa-2, AsPc-1, Panc-28 cells and gemcitabine resistance MiaPaCa-2/GR cells. Moreover, melatonin not only inhibited cell proliferation and invasion in a receptor-independent manner, but also enhanced gemcitabine cytotoxicity at pharmacologic concentrations in these PDAC cells. In vivo, the mice treated with both agents experienced a larger reduction in tumor burden than the single drug-treated groups in an orthotopic xenograft mouse model. Taken together, these results indicate that melatonin inhibits proliferation and invasion of PDAC cells and overcomes gemcitabine resistance of pancreatic tumors through NF-κB inhibition. Our findings therefore provide novel preclinical knowledge about melatonin inhibition of NF-κB in PDAC and suggest that melatonin should be investigated clinically, alone or in combination with gemcitabine for PDAC treatment.

Tumor iron homeostasis and immune regulation.

Abnormal iron metabolism has long been regarded as a key metabolic hallmark of cancer. As a critical cofactor, iron contributes to tumor progression by participating in various processes such as mitochondrial electron transport, gene regulation, and DNA synthesis or repair. Although the role of iron in tumor cells has been widely studied, recent studies have uncovered the interplay of iron metabolism between tumor cells and immune cells, which may affect both innate and adaptive immune responses. In this review, we discuss the current understanding of the regulatory networks of iron metabolism between cancer cells and immune cells and how they contribute to antitumor immunity, and we analyze potential therapeutics targeting iron metabolism. Also, we highlight several key challenges and describe potential therapeutic approaches for future investigations.

Circular RNAs: Emerging regulators of glucose metabolism in cancer.

Aberrant glucose metabolism is one of the most striking characteristics of metabolic reprogramming in cancer. Thus, clarifying the regulatory mechanism of glucose metabolism is crucial to understanding tumor progression and developing novel therapeutic strategies for cancer patients. Recent developments in circular RNAs have explained the regulatory mechanism of glucose metabolism from a new dimension. In this review, we briefly summarize the recent advances in circRNA research on cancer glucose metabolism and emphasize the different regulatory mechanisms, including acting as miRNA sponges, interacting with proteins and being translated into proteins. Additionally, we discuss the future research directions of circular RNAs in the field of glucose metabolism.

The Macrophage-Associated LncRNA MALR Facilitates ILF3 Liquid-Liquid Phase Separation to Promote HIF1α Signaling in Esophageal Cancer.

SIGNIFICANCE: Secretion of TNFα by tumor-associated macrophages stimulates cancer cells to upregulate lncRNA MALR, which induces ILF3 liquid-liquid phase separation and activation of HIF1α signaling to promote cancer progression.