Protocol to develop a chemotherapy drug screening process by constructing a cancer prognostic model using public databases.

Drug resistance is currently the biggest challenge in cancer chemotherapy. Here, we present a protocol to develop a chemotherapy drug screening process by constructing a cancer prognostic model (PM) using public databases. We describe steps for downloading code and data, preparing the expression matrix and metadata for analysis, screening modeling genes, and constructing a PM. We then detail procedures for constructing predictive websites for cancer patients' survival based on their age, tumor stage, gene expression levels, and risk scores. For complete details on the use and execution of this protocol, please refer to Bai et al.1.

Protocol to detect immune levels, abnormal metabolism, and signaling pathways in tumor tissue based on scRNA-seq obtained from patient databases.

Malignant tumor cells are typically more active in terms of metabolism and signal transduction compared to immune and normal cells. Here, we present a protocol to evaluate immune levels, abnormal metabolism, and signaling pathways in tumor tissue based on single-cell sequencing based on patient data obtained from the GEO database. We describe steps for tumor immune microenvironment (TIME)-based evaluation, tumor purity assessment, and identification of abnormal signal transduction and metabolic pathways. We then detail procedures for screening hub genes. For complete details on the use and execution of this protocol, please refer to Bai et al.1.

Screening for metastasis-related genes in mouse melanoma cells through sequential tail vein injection.

Tumor metastasis, responsible for approximately 90% of cancer-associated mortality, remains poorly understood. Here in this study, we employed a melanoma lung metastasis model to screen for metastasis-related genes. By sequential tail vein injection of mouse melanoma B16F10 cells and the subsequently derived cells from lung metastasis into BALB/c mice, we successfully obtained highly metastatic B16F15 cells after five rounds of in vivo screening. RNA-sequencing analysis of B16F15 and B16F10 cells revealed a number of differentially expressed genes, some of these genes have previously been associated with tumor metastasis while others are novel discoveries. The identification of these metastasis-related genes not only improves our understanding of the metastasis mechanisms, but also provides potential diagnostic biomarkers and therapeutic targets for metastatic melanoma.

The combined characteristics of cholesterol metabolism and the immune microenvironment may serve as valuable biomarkers for both the prognosis and treatment of hepatocellular carcinoma.

Background: Hepatocellular carcinoma (HCC) being a complex disease, commonly exhibits multifaceted presentations, rendering its treatment challenging and necessitating specific approaches. The tumor immune microenvironment is crucial in cancer treatment, and cholesterol metabolism is a key component that helps cells grow and produce vital metabolites. However, the reprogramming of cholesterol metabolism in the tumor microenvironment (TME) can promote HCC development, and cancer classifiers relating to cholesterol metabolism are currently limited. Despite significant progress, further research is needed to improve early detection, liver function, and treatment options to improve patient outcomes. Methods: To evaluate the expression abundance of tumor immune microenvironment (TIME) and cholesterol metabolism in 8 types of liver cancer cells, we comprehensively evaluated the immune cell composition, extracellular matrix alterations, and activity of relevant signaling pathways in the TIME through nine liver cancer patients, stromal scoring, immune scoring, tumor purity scoring, immune infiltration analysis, and pathway enrichment. Subsequently, we utilized machine learning techniques to construct prognostic models for both cholesterol metabolism and the tumor immune microenvironment, further exploring the tumor mutation burden, immune infiltration levels, and drug sensitivity in different subtypes of HCC patients. Results: Our study constructed three cancer screening models to identify HCC patients with high cholesterol metabolism and low TIME, who have a poorer prognosis. On the contrary, patients with low cholesterol metabolism and high TIME often have better prognosis. Furthermore, we identified chemical compounds, such as BPD-00008900, ML323, Doramapimod, and AZD2014, which display better chemotherapy results for high-risk patients in specific sub-groups.

The roles and mechanism of STIM1 in tumorigenesis and metastasis.

STIM1 (stromal interaction molecule 1) is one of the key components of the store operated Ca2+ entry channel (SOCE), which is located on the endoplasmic reticulum membrane and highly expressed in most kinds of tumors. STIM1 promotes tumorigenesis and metastasis by modulating the formation of invadopodia, promoting angiogenesis, mediating inflammatory response, altering the cytoskeleton and cell dynamics. However, the roles and mechanism of STIM1 in different tumors have not been fully elucidated. In this review, we summarize the latest progress and mechanisms of STIM1 in tumorigenesis and metastasis, thereby providing insights and references for the study on STIM1 in the field of cancer biology in the future.

The role of hepatocyte nuclear factor 4α (HNF4α) in tumorigenesis.

Hepatocyte Nuclear Factor 4 Alpha (HNF4α) is a master transcription factor mainly expressed in the liver, kidney, intestine and endocrine pancreas. It regulates multiple target genes involved in embryonic development and metabolism. HNF4α-related diseases include non-alcoholic fatty liver disease (NAFLD), obesity, hypertension, hyperlipidemia, metabolic syndrome and diabetes mellitus. Recently, HNF4α has been emerging as a key player in a variety of cancers. In this review, we summarized the role and mechanism of HNF4α in different types of cancers, especially in liver and colorectal cancer, aiming to provide additional guidance for intervention of these diseases.

Caenorhabditis elegans NHR-14/HNF4α regulates DNA damage-induced apoptosis through cooperating with cep-1/p53.

BACKGROUND: Nuclear hormone receptors are involved in transcriptional regulation and many important cellular processes including development and metabolism. However, its role in DNA damage-induced apoptosis remains elusive. METHODS: Synchronized young adult animals were irradiated with different doses of gamma-Ray, and then put back to culture at 20 °C. Germline cell apoptosis was scored at different time point. RESULTS: Deletion of nhr-14 led to decreased DNA damage-induced germline apoptosis, but not the physiological programmed cell death. We also demonstrate that nhr-14 functions downstream of the DNA damage checkpoint pathway. Moreover, we show that nhr-14 regulates egl-1 and ced-13 transcription upon DNA damage. Mechanistically, NHR-14 forms a complex with CEP-1/p53 and binds directly to the egl-1 promoter to promote egl-1 transcription.. CONCLUSIONS: Our results indicate that NHR-14/HNF4α cooperates with CEP-1/p53 to regulate DNA damage-induced apoptosis. Video abstract.

PYK2 mediates the BRAF inhibitor (vermurafenib)-induced invadopodia formation and metastasis in melanomas.

OBJECTIVE: The BRAF inhibitor, vemurafenib, has been widely used in the treatment of patients with melanoma-bearing BRAFV600E mutations. While the initial response to vemurafenib is usually excellent, the majority of patients eventually develop resistance and metastatic disease. However, the underlying molecular mechanism remains elusive. The objective of this study was therefore to identify additional molecular targets responsible for vemurafenib resistance. METHODS: Western blots and immunohistochemistry analyses were used to evaluate expressions of PYK2 and p-PYK2 in cultured cells and melanoma tissue microarrays. The relationships of p-PYK2 with clinicopathological parameters were statistically analyzed. Invadopodia cell invasion, and a Ca2+ assay were used to determine the effect of vemurafenib resistance-induced p-PYK2 on melanoma progression. A mouse model was used to assess the effects of PYK2 on melanoma metastasis. RESULTS: Elevated p-PYK2 levels were detected in vemurafenib-resistant melanoma cells, and PYK2 was shown to regulate invadopodia formation in melanoma cells. Vemurafenib triggered invadopodia formation by activation of PYK2. Inhibition of PYK2 with either shRNA or the small molecule inhibitor, PF562711, dramatically reduced vemurafenib-induced invadopodia formation. Furthermore, knockdown of PYK2 significantly reduced melanoma lung metastasis in vivo. Increased expressions of p-PYK2 in melanoma patients were positively correlated with advanced stage (P = 0.002), metastasis (P < 0.001), and Clark grade (P < 0.001), and were also associated with short overall survival [hazard ratio (HR) = 3.304, P = 0.007] and progression-free survival (HR = 2.930, P = 0.001). CONCLUSIONS: PYK2 mediated vemurafenib-induced melanoma cell migration and invasion. Inhibition of PYK2 resensitized melanoma cells to vemurafenib. Phospho-PYK2 was a prognostic biomarker in melanoma patients.

Mitochondrial Deoxyguanosine Kinase Regulates NAD+ Biogenesis Independent of Mitochondria Complex I Activity.

Overexpression of DGUOK promotes mitochondria oxidative phosphorylation and lung adenocarcinoma progression. However, the role and mechanism of DGUOK in regulation of mitochondria function and lung cancer progression still poorly understood. Here we demonstrated that DGUOK regulated NAD+ biogenesis. Depletion of the DGUOK significantly decreased NAD+ level. Furthermore, knockout of the DGUOK considerably reduced expression of the NMNAT2, a key molecule controlling NAD+ synthesis, at both mRNA and protein levels. Ectopic expression of the NMNAT2 abrogated the effect of knockdown of DGUOK on NAD+. Notably, this regulation is independent of DGUOK -mediated mitochondria complex I activity. We also showed that NMNAT2 was highly expressed in lung adenocarcinoma and negatively correlated with the patient overall survival. Our study suggested that DGUOK regulates NAD+ in a NMNAT2 dependent manner and DGUOK-NMNAT2-NAD+ axis could be a potential therapeutic target in lung adenocarcinoma.