Promotes Cancer Proliferation Research Articles

LncRNA CARMN m6A demethylation by ALKBH5 inhibits mutant p53-driven tumour progression through miR-5683/FGF2.

N-methyladenosine (m6A) represents a prevalent RNA modification observed in colorectal cancer. Despite its abundance, the biological implications of m6A methylation on the lncRNA CARMN remain elusive in colorectal cancer, especially for mutant p53 gain-of-function. Here, we elucidate that CARMN exhibits diminished expression levels in colorectal cancer patients with mutant p53, attributed to its rich m6A methylation, which promotes cancer proliferation, invasion and metastasis in vitro and in vivo. Further investigation illustrates that ALKBH5 acts as a direct demethylase of CARMN, targeting 477 methylation sites, thereby preserving CARMN expression. However, the interaction of mutant p53 with the ALKBH5 promoter impedes its transcription, enhancing m6A methylation levels on CARMN. Subsequently, YTHDF2/YTHDF3 recognise and degrade m6A-modified CARMN. Concurrently, overexpressing CARMN significantly suppressed colorectal cancer progression in vitro and in vivo. Additionally, miR-5683 was identified as a direct downstream target of lncRNA CARMN, exerting an antitumour effect by cooperatively downregulating FGF2 expression. Our findings revealed the regulator and functional mechanism of CARMN in colorectal cancer with mutant p53, potentially offering insights into demethylation-based strategies for cancer diagnosis and therapy. The m6A methylation of CARMN that is prime for mutant p53 gain-of-function-induced malignant progression of colorectal cancer, identifying a promising approach for cancer therapy.

NXPH4 can be used as a biomarker for pan-cancer and promotes colon cancer progression.

NXPH4 promotes cancer proliferation and invasion. However, its specific role and mechanism in cancer remain unclear. Transcriptome and clinical data for pan-cancer were derived from the TCGA database. K-M survival curve and univariate Cox were used for prognostic analysis. CIBERSORT and TIMER algorithms were employed to calculate immune cell infiltration. Gene set enrichment analysis (GSEA) was employed for investigating the function of NXPH4. Western blot verified differential expression of NXPH4 in colon cancer. Functional assays (CCK-8, plate clonogenicity assay, wound healing assay, and Transwell assay) confirmed the impact of NXPH4 on proliferation, invasion, and migration of colon cancer cells. Dysregulation of NXPH4 in pan-cancer suggests its potential as a diagnostic and prognostic marker for certain cancers, including colon and liver cancer. High expression of NXPH4 in pan-cancer might be associated with the increase in copy number and hypomethylation. NXPH4 expression in pan-cancer is substantially linked to immune cell infiltration in the immune microenvironment. NXPH4 expression is associated with the susceptibility to immunotherapy and chemotherapy. Western blot further confirmed the higher expression of NXPH4 in colon cancer. Knockdown of NXPH4 significantly suppresses proliferation, invasion, and migration of colon cancer cell lines HT-29 and HCT116, as validated by functional assays.

Abstract 2897: Cancer associated fibroblasts targeted photoimmunotherapy improves drug delivery

Abstract Introduction: Desmoplasia is often associated with adverse outcomes in cancer, particularly in esophageal cancers characterized by stroma-rich environments and poor prognoses. The dense extracellular matrix (ECM) within the stroma hinders effective drug delivery, primarily due to elevated interstitial pressure. Cancer-associated fibroblasts (CAFs) of the tumor microenvironment deteriorate this issue by promoting cancer proliferation and ECM production. We have developed a photoimmunotherapy targeting these CAFs via fibroblast activation protein (FAP-PIT), which has demonstrated notable antitumor efficacy in CAF-rich tumors that are typically chemoresistant. This study explores the potential of FAP-PIT to enhance drug delivery in CAF-rich tumors. Methods: We have conjugated Sibrotuzumab, a clinically-trialed FAP antibody with the photosensitizer IR700DX for application in FAP-PIT. We compared the protein expression levels of ECM components—collagen I (Col I) and hyaluronic acid (HA)—between CAFs and normal fibroblasts (NFs) in vitro. Spheroid models, both CAF-poor (solely cancer cells) and CAF-rich (cancer cells plus fibroblasts), were employed to assess drug uptake. Additionally, 140 esophageal cancer cases from 2008 to 2010 were retrospectively analyzed. In vivo, we induced bilateral subcutaneous tumors to apply FAP-PIT, monitoring drug uptake using in vivo imaging systems (IVIS) and immunohistochemistry (IHC). Results: Initially, we determined how significantly CAFs impede the drug delivery. In vitro, CAFs secreted higher levels of Col I and HA compared to NFs. Clinically, the expression of Col I and HA consistently showed a positive correlation with α-smooth muscle actin (αSMA) and FAP. CAF-rich spheroids exhibited significantly reduced uptake of fluorescently labeled chemotherapy compared to CAF-poor spheroids. Conversely, FAP-PIT enhanced drug penetration in CAF-rich spheroids through CAFs depletion and ECM remodeling. In vivo studies further demonstrated that FAP-PIT significantly increased Panitumumab and nab-Paclitaxel accumulation in CAF-rich tumors, leading to superior antitumor efficacy. IHC analysis validated the significant reduction of CAFs and Col I. Conclusion: The ECM, including CAFs, poses a barrier to drug delivery. Our findings indicate that FAP-PIT can effectively enhance drug penetration in CAF-rich tumors, presenting a novel therapeutic strategy to overcome drug resistance. Citation Format: Seitaro Nishimura, Kazuhiro Noma, Tasuku Matsumoto, Tatsuya Takahashi, Yasushige Takeda, Hijiri Matsumoto, Tomoyoshi Kunitomo, Kento Kawasaki, Hajime Kashima, Takuya Kato, Satoru Kikuchi, Toshiaki Ohara, Hiroshi Tazawa, Toshiyoshi Fujiwara. Cancer associated fibroblasts targeted photoimmunotherapy improves drug delivery [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2897.

Anti‑tumor properties of FoxO1 in YD‑9 oral squamous cell carcinoma cells.

Oral squamous cell carcinoma (OSCC) is a tumor with a poor prognosis and a high recurrence rate. Despite its high annual incidence worldwide, appropriate therapeutic strategies have not yet been developed. Consequently, the 5‑year survival rate for OSCC is low when advanced stages or recurrence is diagnosed. Forkhead transcriptional factor O1 (FoxO1) is a key mediator for maintaining cellular homeostasis. FoxO1 can function as a tumor suppressor as well as an oncogene depending on the cancer type. Therefore, the precise molecular functions of FoxO1 need to be validated, considering intracellular factors and the extracellular environment. To the best of our knowledge, however, the roles of FoxO1 in OSCC have not yet been defined. The present study examined FoxO1 levels under pathological conditions (oral lichen planus and oral cancer) and selected an appropriate OSCC cell line (YD‑9). Crispr/Cas9 was used to generate FoxO1‑deficient YD‑9 cells in which the protein levels of phospho ERK and phospho STAT3 were upregulated, promoting cancer proliferation and migration. In addition, FoxO1 reduction increased the levels of the cell proliferation markers phospho H3 (Ser10) and PCNA. FoxO1 loss significantly reduced cellular ROS levels and apoptosis in YD‑9 cells. Collectively, the present study demonstrated that FoxO1 exerted an anti‑tumor effect by suppressing proliferation and migration/invasion but promoting oxidative stress‑linked cell death in YD‑9 OSCC cells.

Hepatocellular carcinoma progression promoted by 5-lipoxygenase activity in CD163(+) tumor-associated macrophages

Arachidonic acid 5-lipoxygenase (5-LOX), an enzyme that synthesizes leukotrienes (LTs), is involved in cancer development including proliferation, invasion, metastasis and drug resistance. However, the functional role of 5-LOX in hepatocellular carcinoma (HCC) remains to be elucidated. In this study, we analyzed the contribution of 5-LOX in HCC progression and investigated the potential of targeted therapy. Analysis of 86 resected HCC specimens and the clinical data of 362 cases of liver cancer from The Cancer Genome Atlas Liver Hepatocellular Carcinoma dataset, showed that 5-LOX expression was associated with postoperative survival. The cancer proliferative and stem cell potential were correlated with the levels of 5-LOX in CD163(+) tumor-associated macrophages (TAMs). In an HCC mouse model, CD163(+) TAMs expressed 5-LOX and produced LTB4 and LTC/D/E4; the 5-LOX inhibitor, zileuton, suppressed HCC progression. LTB4 and LTC/D/E4 promoted cancer proliferation and stem cell capacity via phosphorylation of extracellular signal-regulated kinase 1/2 and stem cell-associated genes. Taken together, we identified a novel mechanism of HCC progression in which CD163(+) TAMs express 5-LOX and produce LTB4 and LTC/D/E4, thereby enhancing the proliferative and stem cell potential of HCC cells. Furthermore, inhibition of 5-LOX activity regulates HCC progression, suggesting it has potential as a new therapeutic target.

Exosomal lncRNA HCG18 contributes to cholangiocarcinoma growth and metastasis through mediating miR-424-5p/SOX9 axis through PI3K/AKT pathway

Cholangiocarcinoma is a highly aggressive malignant tumor disease with the increasing incidence and mortality. It’s urgent to identify specific biomarkers for cholangiocarcinoma treatment and diagnosis. Recent studies have noted the importance of lncRNAs in cancer and the following downstream mechanism with miRNAs network has been a hotspot. This work aimed to discover the role of lncRNA HCG18 and its possible downstream mechanism in cholangiocarcinoma tumor progression. Initially, through bioinformatics tools, we observed abnormal expression of lncRNA HCG18 in cholangiocarcinoma. In vitro experiments like (CCK-8, EdU, colony formation, flow cytometry, transwell, wound healing assays) and animal study confirmed that lncRNA HCG18 served as a cancer-promoting gene, promoted cancer proliferation, migration and invasion abilities. Besides, we found cancer cell-secreted exosomes transitted HCG18 to surrounding tumor cells and accelerated tumor growth and metastasis. After that, we confirmed HCG18 directly interacted with miR-424-5p through FISH, RIP and dual luciferase reporter assays with negative modulation. The inhibition of miR-424-5p reversed the HCG18 knockdown induced suppression on cholangiocarcinoma cancer cells. More specific, miR-424-5p targeted to SOX9 contributed to cholangiocarcinoma growth and metastasis through mediating PI3K/AKT pathway. In conclusion, these findings provide solid evidence of lncRNAs/miRNAs regulation in cholangiocarcinoma progression.

Downregulation of T-cell cytotoxic marker IL18R1 promotes cancer proliferation and migration and is associated with dismal prognosis and immunity in lung squamous cell carcinoma.

Immunotherapy can improve the survival of patients with advanced lung squamous cell carcinoma (LUSC). T cytotoxic cells are one of the main members of the immune microenvironment. Herein, we aimed to identify the roles of T-cell cytotoxic markers interleukin 18 (IL18) receptor 1 (IL18R1) in the LUSC progression using bioinformatics, clinical tissue specimen, and cell experiment. We assessed the association between the IL18R1 expression and immune infiltration and IL18R1-related competing RNA network. The IL18R1 expression was downregulated in the LUSC tissues. The IL18R1 expression downregulation was associated with diagnosis and short overall survival and disease-specific survival, and it was also an independent risk factor for dismal survival time in LUSC. IL18R1-related nomograms predicted the survival time of patients with LUSC. IL18R1 overexpression inhibited the proliferation, migration, and invasion of LUSC cells. The IL18R1 expression was significantly associated with the microenvironment (stromal, immune, and estimate scores), immune cells (such as the T cells, cytotoxic cells, CD8 T cells), and immune cell markers (such as the CD8A, PD-1, and CTLA4) in LUSC. AC091563.1 and RBPMS-AS1 downregulation was positively associated with the IL18R1 expression, negatively associated with the miR-128-3p expression, and associated with short disease-specific survival and progression in LUSC. In conclusion, IL18R1 was significantly downregulated and associated with the prognosis and immune microenvironment. IL18R1 overexpression inhibits the growth and migration of cancer cells in LUSC. Furthermore, AC091563.1 and RBPMS-AS1 might compete with IL18R1 to bind miR-128-3p for participating in LUSC progression. These results showed that IL18R1 is a biomarker for evaluating the prognosis of patients with LUSC.

Dysfunctional tRNA reprogramming and codon-biased translation in cancer.

Many cancers hijack translation to increase the synthesis of tumor-driving proteins, the messenger mRNAs of which have specific codon usage patterns. Termed 'codon-biased translation' and originally identified in stress response regulation, this mechanism is supported by diverse studies demonstrating how the 50 RNA modifications of the epitranscriptome, specific tRNAs, and codon-biased mRNAs are used by oncogenic programs to promote proliferation and chemoresistance. The epitranscriptome writers METTL1-WDR4, Elongator complex protein (ELP)1-6, CTU1-2, and ALKBH8-TRM112 illustrate the principal mechanism of codon-biased translation, with gene amplifications, increased RNA modifications, and enhanced tRNA stability promoting cancer proliferation. Furthermore, systems-level analyses of 34 tRNA writers and 493 tRNA genes highlight the theme of tRNA epitranscriptome dysregulation in many cancers and identify candidate tRNA writers, tRNA modifications, and tRNA molecules as drivers of pathological codon-biased translation.

Zinc Finger Proteins: Functions and Mechanisms in Colon Cancer.

According to the global cancer burden data for 2020 issued by the World Health Organization (WHO), colorectal cancer has risen to be the third-most frequent cancer globally after breast and lung cancer. Despite advances in surgical treatment and chemoradiotherapy for colon cancer, individuals with extensive liver metastases still have depressing prognoses. Numerous studies suggest ZFPs are crucial to the development of colon cancer. The ZFP family is encoded by more than 2% of the human genome sequence and is the largest transcriptional family, all with finger-like structural domains that could combine with Zn2+. In this review, we summarize the functions, molecular mechanisms and recent advances of ZFPs in colon cancer. We also discuss how these proteins control the development and progression of colon cancer by regulating cell proliferation, EMT, invasion and metastasis, inflammation, apoptosis, the cell cycle, drug resistance, cancer stem cells and DNA methylation. Additionally, several investigations have demonstrated that Myeloid zinc finger 1 (MZF1) has dual functions in colon cancer, which may both promote cancer proliferation and inhibit cancer progression through apoptosis. Generally, a comprehensive understanding of the action mechanisms of ZFPs in colon cancer will not only shed light on the discovery of new diagnostic and prognosis indicators but will also facilitate the design of novel targeted therapies.

The mechanism of branched-chain amino acid transferases in different diseases: Research progress and future prospects.

It is well known that the enzyme catalyzes the first step of branched-chain amino acid (BCAA) catabolism is branched-chain amino transferase (BCAT), which is involved in the synthesis and degradation of leucine, isoleucine and valine. There are two main subtypes of human branched chain amino transferase (hBCAT), including cytoplasmic BCAT (BCAT1) and mitochondrial BCAT (BCAT2). In recent years, the role of BCAT in tumors has attracted the attention of scientists, and there have been continuous research reports that BCAT plays a role in the tumor, Alzheimer’s disease, myeloid leukaemia and other diseases. It plays a significant role in the growth and development of diseases, and new discoveries about this gene in some diseases are made every year. BCAT usually promotes cancer proliferation and invasion by activating the phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin pathway and activating Wnt/β-catenin signal transduction. This article reviews the role and mechanism of BCAT in different diseases, as well as the recent biomedical research progress. This review aims to make a comprehensive summary of the role and mechanism of BCAT in different diseases and to provide new research ideas for the treatment, prognosis and prevention of certain diseases.

CircARID1A binds to IGF2BP3 in gastric cancer and promotes cancer proliferation by forming a circARID1A-IGF2BP3-SLC7A5 RNA–protein ternary complex

BackgroundGastric cancer (GC) is one of the most common malignant tumors in China. Circular RNAs (circRNAs) are novel non-coding RNAs with important regulatory roles in cancer progression. IGF2BP3 has been found to play oncogenic roles in various cancers including GC, while the exact mechanism of IGF2BP3 is largely unknown.MethodsThe expression of IGF2BP3 in GC was evaluated by Western Blot and bioinformatics analysis. CircRNA expression profiles were screened via IGF2BP3 RIP-seq in GC. Sanger sequencing, RNase R digestion, nucleo-plasmic separation and RNA-FISH assays were used to detect the existence and expression of circARID1A. RNA ISH assay was employed to test the expression of circARID1A in paraffin-embedded GC tissues. Moreover, the function of circARID1A on cellular proliferation was assessed by CCK-8, plate colony formation, EdU assays and GC xenograft mouse model in vivo. Furthermore, the location or binding of circARID1A, IGF2BP3 protein and SLC7A5 in GC was evaluated by RNA-FISH/IF or RNA pull-down assays.ResultsWe identified a novel circRNA, circARID1A, that can bind to IGF2BP3 protein. CircARID1A was significantly upregulated in GC tissues compared with noncancerous tissues and positively correlated with tumor length, tumor volume, and TNM stage. CircARID1A knockdown inhibited the proliferation of GC cells in vitro and in vivo and circARID1A played an important role in the oncogenic function of IGF2BP3. Mechanistically, circARID1A served as a scaffold to facilitate the interaction between IGF2BP3 and SLC7A5 mRNA, finally increasing SLC7A5 mRNA stability. Additionally, circARID1A was able to directly bind SLC7A5 mRNA through complementary base-pairing and then formed the circARID1A-IGF2BP3-SLC7A5 RNA–protein ternary complex and promoted the proliferation of GC via regulating AKT/mTOR pathway.ConclusionsAltogether, our data suggest that circARID1A is involved in the function of IGF2BP3 and GC proliferation, and the circARID1A-IGF2BP3-SLC7A5 axis has the potential to serve as a novel therapeutic target for GC.

TAZ promotes vasculogenic mimicry in gastric cancer through the upregulation of TEAD4.

Vasculogenic mimicry (VM) is a unique blood supply pattern in malignant tumors that is closely associated with metastasis and poor prognosis. The Hippo signaling effector TAZ is upregulated in several cancers, promoting cancer proliferation and metastasis. This study aimed to identify the function of TAZ and its regulatory mechanism in promoting VM in gastric cancer (GC). The expression of TAZ and TEAD4 and their correlations with overall survival and VM-related markers were analyzed in 228 cases of GC. The regulatory mechanism of TAZ and its interaction with TEAD4 in epithelial-mesenchymal transition (EMT) and VM were investigated in vitro and in vivo. TAZ was highly expressed in GC samples and was associated with shorter patient survival time. TAZ expression was positively correlated with TEAD4 and VM in patients with GC. TAZ enhanced the migration and invasion capacity of GC cells through EMT in vitro and upregulated the expression of VM-associated proteins, including VE-cadherin, MMP2, and MMP9, thus promoting VM formation. Overexpression of TAZ accelerated the growth of subcutaneous xenograft and promoted VM formation in vivo. Co-immunoprecipitation assays showed that TAZ can directly bind to TEAD4, and in vitro experiments showed that this binding mediates the function of TAZ in regulating EMT and VM formation in GC. TAZ promotes GC metastasis and VM by upregulating TEAD4 expression. Our findings expand the role of TAZ in VM and provide new theoretical support for the use of antiangiogenic therapy in the treatment of advanced GC.

Abstract P5-05-08: CXCL16 as a potential therapeutic target of triple-negative breast cancer

Abstract CXCL16, as a chemotactic cytokine, promotes cancer proliferation, migration, and immune cell infiltration in tumor microenvironment. However, CXCL16 has been shown to play a conflicting role in breast cancer. The aim of this study was to investigate the therapeutic potential of CXCL16 in breast cancer. Breast cancer patients (n=1097) from the TCGA dataset were divided into two group according to the CXCL16 expressions, the CXCL16low (n=548) and the CXCLhigh (n=549) groups. Mean diagnostic ages were similar between groups (59 ± 13.5 vs 58 ± 12.9 years, P=0.233). The ratio of triple-negative breast cancer (TNBC) was significantly higher in the CXCLhigh than that of the CXCL16low groups (24.8 vs 6.8%, P<0.001), while the presence of distant metastasis or overall survivals were not different between groups. Interestingly, gene scoring analysis showed that oncogenic molecules including M2-macrophage- (r=0.36, P<0.001), efferocytosis- (r=0.42, P<0.001), and chemokine- (r=0.46, P<0.001) related gene sets showed significantly strong positive relations with the CXCL16 expressions in all patients including both TNBC (n=924) and non-TNBC (n=173). Additionally, the gene set score from PI3K/Akt pathway showed positive relation with the CXCL16 expressions in the non-TNBC (r=022, P<0.001) but not in the TNBC group. Next, a xenograft mouse model of TNBC was established with MDA-MB-231 cells, and anti-mouse CXCL16 antibody (designated as aCXCL16 group) or control IgG (designated as control group) were intravenously injected (25µg/100µl), twice per week. During 3 weeks, tumor growth was significantly delayed in the aCXCL16 than the control group by 37%. In conclusion, CXCL16 is highly expressed in TNBC and positively correlates with M2-macrophage related genes. Neutralizing CXCL16 by anti-CXCL16 shows therapeutic effects on the xenograft model of TNBC, suggesting that CXCL16 can be a potential therapeutic target for breast cancer. Citation Format: Sun Wook Cho, Mi Gyeong Jang, Hyun Jin Sun, Han Sai Lee, Young Shin Song, Seong Keun Kim. CXCL16 as a potential therapeutic target of triple-negative breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-05-08.

Nerve Dependence in Colorectal Cancer.

Cancerous invasion of nerves has been reported in a list of malignant tumors as a high-risk pathological feature and marker of poor disease outcome especially in neurotrophic cancers (such as in pancreas and prostate), indicating that although once neglected, nerves could have played a pivotal role in tumorigenesis and cancer progression. In colorectal cancer, perineural invasion, a specific form of tumor-nerve interaction referring to the identification of tumor cells in proximity to the nerve, has been recognized as a strong and independent prognosis predictor; denervation of autonomic nerves and enteric nerves have shown that the existence of these nerves in the gut are accompanied by promoted cancer proliferation, further supporting that nerve is a potential accomplice to shield and nurture tumor cells. However, the precise role of nerve in CRC and the pattern of interaction between CRC cells and nerve has not been unveiled yet. Here we aim to review some basic knowledge of the importance of nerves in CRC and attempt to depict a mechanistic view of tumor-nerve interaction during CRC development.

SETD8 stabilized by USP17 epigenetically activates SREBP1 pathway to drive lipogenesis and oncogenesis of ccRCC

Recently, epigenetic modifications, including DNA methylation, histone modification and noncoding RNA (ncRNA)-associated gene silencing, have received increasing attention from the scientific community. Many studies have demonstrated that epigenetic regulation can render dynamic alterations in the transcriptional potential of a cell, which then affects the cell's biological function. The initiation and development of clear cell renal cell carcinoma (ccRCC), the most common subtype of renal cell cancer (RCC), is also closely related to genomic alterations by epigenetic modification. For ccRCC, lipid accumulation is one of the most typical characteristics. In other words, dysregulation of lipid uptake and synthesis occurs in ccRCC, which inversely promotes cancer proliferation and progression. However, the link among epigenetic alterations, lipid biosynthesis and renal cancer progression remains unclear. SETD8 is a histone methyltransferase and plays pivotal roles in cell cycle regulation and oncogenesis of various cancers, but its role in RCC is not well understood. In this study, we discovered that SETD8 was significantly overexpressed in RCC tumors, which was positively related to lipid storage and correlated with advanced tumor grade and stage and poor patient prognosis. Depletion of SETD8 by siRNAs or inhibitor UNC0379 diminished fatty acid (FA) de novo synthesis, cell proliferation and metastasis in ccRCC cells. Mechanistically, SETD8, which was posttranslationally stabilized by USP17, could transcriptionally modulate sterol regulatory element-binding protein 1 (SREBP1), a key transcription factor in fatty acid biosynthesis and lipogenesis, by monomethylating the 20th lysine of the H4 histone, elevating lipid biosynthesis and accumulation in RCC and further promoting cancer progression and metastasis. Taken together, the USP17/SETD8/SREBP1 signaling pathway plays a pivotal role in promoting RCC progression. SETD8 might be a novel biomarker and potential therapeutic target for treating RCC.

Administration of P2X7 Receptor Blockers in Oncological Experimental Models.

The tumor microenvironment is rich in components that strongly influence cancer cell survival. One of the pivotal molecules present at the tumor bed is ATP, which has an essential role in promoting cancer proliferation and metastasis and immune responses via its receptor P2X7. Several studies have proved the efficacy of P2X7 pharmacological blockade in inhibiting primary and metastatic tumor growth in preclinical models. Here we describe the experimental procedures that we optimized to test P2X7 roles in carcinogenesis by antagonist administration. Special attention is paid to their concentrations and routes of administration. The depicted in vitro models include cell count and viability assays, which are useful to test P2X7 roles in cell proliferation and vitality, and the soft agar colony formation test that allows investigation of the transforming and invading abilities of tumor cells. We also describe systemic and intramass administration of P2X7 blockers in murine models of melanoma and leukemia. Both xenotransplant and syngeneic experimental tumor models are detailed.

Hsa_circ_0023305 Enhances Laryngeal Squamous Cell Carcinoma Progression and Modulates TRPM7 via miR-218-5p Sponging.

Recently, circular RNAs have been shown to function as critical regulators of many human cancers. However, the circRNA mechanism in laryngeal squamous cell carcinoma (LSCC) remains elusive. Recent investigations using bioinformatics analysis revealed high expression of hsa_circ_0023305 in LSCC tissues compared to normal tissues. Furthermore, we discovered that hsa_circ_0023305 expression level was positively correlated to tumor/node/metastasis (TNM) stage as well as lymph node metastasis in LSCC. Moreover, higher hsa_circ_0023305 levels were correlated to poorer LSCC patient outcomes. Knockdown of hsa_circ_0023305 significantly inhibited LSCC cell proliferation, invasion, and migration abilities. Our team validated that hsa_circ_0023305 functioned as a miR-218-5p sponge from a mechanistic perspective, targeting the melastatin-related transient receptor potential 7 (TRPM7) in LSCC cells. TRPM7 regulates a nonselective cation channel and promotes cancer proliferation and metastasis. Our data demonstrated that miR-218-5p was downregulated in LSCC and that miR-218-5p upregulation repressed LSCC proliferation and invasion both in vivo and in vitro. Additionally, we found that hsa_circ_0023305-mediated upregulation of TRPM7 inhibited miR-218-5p and contributed to LSCC migration, proliferation, and invasion. In summary, these data propose a new mechanism by which the hsa_circ_0023305/miR-218-5p/TRPM7 network enhances LSCC progression.

SLC34A2 promotes cancer proliferation and cell cycle progression by targeting TMPRSS3 in colorectal cancer

Solute carrier family 34 member 2 (SLC34A2), a family member of sodium-driven phosphate cotransporters, has been reported to facilitate cell proliferation and tumor growth. However, the functional mechanism by which SLC34A2 promotes cell growth and cell cycle progression remains poorly understood. Here, we reported that SLC34A2 was overexpressed in CRC by analysis of TCGA and GEO datasets. A total of 45 differentially expressed genes (DEGs) were identified from comparing SLC34A2-high or –low groups and functional enrichment analysis of these DEGs demonstrated that cell cycle pathway was enriched. Interestingly, we found a positive correlation between TMPRSS3 (transmembrane serine protease 3) and SLC34A2, which was confirmed by RT-qPCR and western blotting. Furthermore, TMPRSS3 was also upregulated in CRC tumor tissues compared to normal tissues. Patients with high TMPRSS3 expression had poor prognosis. Functionally, TMPRSS3 deficiency inhibited cell proliferation and colony formation in CRC cells. TMPRSS3 overexpression in SLC34A2-deficient cells antagonized siSLC34A2-mediated cell cycle inhibition by promoting cyclin E, cyclin A protein expression. Based on these results, our study suggests that SLC34A2 promotes cancer proliferation and cell cycle progression by targeting TMPRSS3 in colorectal cancer cells.

Identification of Circulating Tumor Cells and Chemokine Receptor CCR6 as Noninvasive Biomarkers in Egyptian Patients with Hepatocellular Carcinoma

Background: Circulating tumor cells (CTCs) are cancer cells in the circulation that are derived from the original tumor or metastatic foci. Chemokines and their receptors have major roles in hepatocellular carcinoma (HCC) metastasis. They also represent a very promising group of markers to detect CTCs in HCC patients. The CCL20-CCR6 axis promotes cancer proliferation, migration, and tumor remodeling through immune cell control. Objective: This study aimed to identify the role of CTC and CCR6 in hepatocellular carcinoma patients. Patients and methods: The study included 91 subjects; 71 HCC patients and 20 normal individuals. Routine laboratory investigations included CBC, PT and INR, ALT, AST, bilirubin, albumin, alpha fetoprotein, and creatinine in addition to detection of chemokine receptor 6 (CCR6) RNA by real time PCR and CTCs by flow cytometry as CD45¯CK19+ cells were performed for all subjects enrolled in this study. Results: There was significant difference between the studied groups as regards liver function tests. Also, high significant difference was recorded regarding CCR6 RNA expression, CTCs count and hepatic focal lesion detection by ultra sound. No significant difference between both groups regarding kidney function tests. Correlation studies between CTCs and other variables showed positive correlations with liver function tests (ALT, AST, BIL, Alb, AFP, and INR) and negative correlations with albumin and platelets. Conclusion: CCR6 mRNA concentration was significantly increased in HCC patients and its elevation was correlated with CTCs percentage. Therefore, it can be concluded that the combined assessment of CTCs and CCR6 could be considered as noninvasive biomarkers for HCC patients.

Multiple myeloma driving factor WHSC1 is a transcription target of oncogene HMGA2 that facilitates colon cancer proliferation and metastasis

Colon cancer is a common human cancer worldwide. The survival rate of late staged or metastatic colon cancer patients remains low even though the effectiveness of treatment in colon cancer has greatly improved. Research on tumorigenesis mechanisms and discovery of novel molecular target for treating colon cancer is critical.The promotion roles of WHSC1 in multiple myeloma have been demonstrated previously, yet, the regulation of WHSC1 in other cancers is largely unknown, especially in colon cancer. Here, in this study, we analyzed and identified WHSC1 while studying the genetic regulations of HMGA2 in colon cancer cells by microarray analysis, and investigated the HMGA2-WHSC1 interaction. We then applied CRISPR technology to establish stable WHSC1 knockout cells, to address the functional regulation of WHSC1 in colon cancer. In summary, our results for the first time identified the HMGA2-WHSC1 interaction in colon cancer. Moreover, we discovered that WHSC1 promotes cancer proliferation, facilitates resistance of chemotherapy agent, and promotes metastatic capacity of colon cancer.