Increased Lymph Node Metastasis Research Articles

AMIGO2 characterizes cancer-associated fibroblasts in metastatic colon cancer and induces the release of paracrine active tumorigenic secretomes.

Secretomes of cancer-associated fibroblasts (CAFs) in colorectal cancer (CRC) contribute to malignancy. Detailed knowledge is available on the components and functions of CAF secretomes. Little is known about the regulation of CAF secretomes. Here, we searched for receptor-like membrane-bound molecules in CAFs, which may regulate the production and release of tumor-activating secretomes. The adhesion molecule with Ig-like domain 2 (AMIGO2) was significantly upregulated in cultivated CAFs compared to normal tissue-associated fibroblasts (NAFs), and this was confirmed in patient-derived tissues. AMIGO2 expression was low or absent in healthy colon, significantly increased in fibroblasts of primary CRC, and highest in the stromal tissues of CRC-derived liver metastases. AMIGO2 expression in CAFs correlated with a higher T-category, increased lymph node metastasis, progressed tumor stages and was associated with reduced survival in different cohorts of CRC patients. Interestingly, AMIGO2 expression was induced by transforming growth factor-β and higher in female patients, who exhibit a more aggressive disease course. In functional studies, conditioned media of NAFs with experimentally induced AMIGO2 overexpression enhanced proliferation and migration of different CRC tumor cells, while siRNA-mediated inhibition of AMIGO2 in CAFs attenuated these effects. Accordingly, therapeutic inhibition of the receptor-like AMIGO2 protein in CRC CAFs could prevent tumorigenic secretomes in CRC. © 2024 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

High chromosomal instability is associated with higher 10-year risks of recurrence for hormone receptor-positive, human epidermal growth factor receptor 2-negative breast cancer patients: clinical evidence from a large-scale, multiple-site, retrospective study.

Long-term survival varies among hormone receptor-positive (HR+) and human epidermal growth factor receptor 2-negative (HER2-) breast cancer patients and is seriously impaired by metastasis. Chromosomal instability (CIN) was one of the key drivers of breast cancer metastasis. Here we evaluate CIN and 10-year invasive disease-free survival (iDFS) and overall survival (OS) in HR+/HER2-- breast cancer. In this large-scale, multiple-site, retrospective study, 354 HR+/HER2- breast cancer patients were recruited. Of these, 204 patients were used for internal training, 70 for external validation, and 80 for cross-validation. All medical records were carefully reviewed to obtain the disease recurrence information. Formalin-fixed paraffin-embedded tissue samples were collected, followed by low-pass whole-genome sequencing with a median genome coverage of 1.86X using minimal 1 ng DNA input. CIN was then assessed using a customized bioinformatics workflow. Three or more instances of CIN per sample was defined as high CIN and the frequency was 42.2% (86/204) in the internal cohort. High CIN correlated significantly with increased lymph node metastasis, vascular invasion, progesterone receptor negative status, HER2 low, worse pathological type, and performed as an independent prognostic factor for HR+/- breast cancer. Patients with high CIN had shorter iDFS and OS than those with low CIN [10-year iDFS 11.1% versus 82.2%, hazard ratio (HR) = 11.12, p < 0.01; 10-year OS 45.7% versus 94.3%, HR = 14.17, p < 0.01]. These findings were validated in two external cohorts with 70 breast cancer patients. Moreover, high CIN could predict the prognosis more accurately than Adjuvant! Online score (10-year iDFS 11.1% versus 48.6%, HR = 2.71, p < 0.01). Cross-validation analysis found that high consistency (83.8%) was observed between CIN and MammaPrint score, while only 45% between CIN and Adjuvant! Online score. In conclusion, high CIN is an independent prognostic indicator for HR+/HER2- breast cancer with shorter iDFS and OS and holds promise for predicting recurrence and metastasis.

Clinical and prognostic significance of Hec1 expression in patients with Cervical Cancer.

Hec1 is a component of the Ndc80 kinetochore complex and is frequently upregulated in various cancers. However, the clinical significance of Hec1 in cervical cancer remains largely unknown. This study aimed to investigate the expression patterns of Hec1 in cervical cancer and its relationship with the clinicopathological characteristics of patients diagnosed with the disease. Immunohistochemistry was used to assess the expression of Hec1 in 136 cervical cancer tissue samples and 82 normal cervical tissue samples. The relationship between Hec1 protein expression and the clinicopathological characteristics of cervical cancer patients was analyzed using the Chi-square test. Additionally, the association between Hec1 protein expression and patient survival was examined using Kaplan-Meier survival curves. Independent risk factors affecting the prognosis of cervical cancer patients were analyzed using the Cox proportional hazards regression model. The positive expression rate of Hec1 protein in cervical cancer tissues was 83.82%, significantly higher than the 7.31% in normal cervical tissues. Compared to patients with negative Hec1 expression, those with positive expression exhibited significantly higher FIGO staging, increased lymph node metastasis, greater depth of tumor stromal infiltration, and larger tumor diameter. Multivariable analysis using the Cox proportional hazards regression model indicated that Hec1 positive expression was an independent risk factor for both overall survival (HR = 2.79, 95% CI: 1.65-4.05, p = 0.012) and progression-free survival (HR = 1.81, 95% CI: 1.22-3.18, p = 0.002) in cervical cancer patients. Kaplan-Meier survival curve analysis showed that patients with positive Hec1 expression experienced a lower overall survival (HR: 2.72, 95% CI: 1.15-4.52, p = 0.004) and progression-free survival (HR: 3.12, 95% CI: 1.62-5.03, p = 0.002) when compared to those with negative Hec1 expression. Hec1 is significantly upregulated in cervical cancer tissues and associated with poor prognosis in cervical cancer patients. Therefore, Hec1 could be a novel biomarker, not only for the diagnosis and treatment evaluation of cervical cancer but also as an indicator for predicting the prognosis of cervical cancer patients.

Clinicopathological and prognostic role of CEP55 expression in cancer patients: A meta‐analysis of 31 studies with 12,543 patients

AbstractBackgroundDysregulated expression of centrosomal protein 55 (CEP55) has been detected in multiple types of cancers. However, the clinical value of CEP55 expression in cancer is controversial. The current meta‐analysis quantitatively investigated the association between CEP55 expression and prognostic outcomes in cancers.MethodsA literature search was performed using the Chinese National Knowledge Infrastructure (CNKI), Wanfang databases, Web of Science, Embase, MEDLINE, PubMed, and Cochrane Library for primary studies. The pooled hazard ratios (HRs) and odds ratios (ORs) were used to investigate the association between CEP55 expression and its prognostic and clinicopathological value in cancers.ResultsA total of 12,543 patients from 31 studies were included in this meta‐analysis. High CEP55 expression was significantly associated with poor differentiation, deeper tumor invasion and increased lymph node metastasis in cancer patients. The pooled results indicated that elevated CEP55 expression can predict a poor overall survival (OS) and disease‐free survival (DFS) in cancers. These results should be interpreted with caution because of publication bias. However, the pooled HR for OS of lung cancers was 1.50 (95% CI = 1.36–1.67, p &lt; 0.01) with no heterogeneity and publication bias.ConclusionsCEP55 overexpression correlated with poor cancer differentiation, deeper tumor invasion, and increased lymph node metastasis, suggesting that CEP55 may serve as a predictive and prognostic biomarker for cancers, especially for lung cancers.

Lysine Methyltransferase 2D Regulates Immune Response and Metastasis in Head and Neck Cancer.

The most frequently altered epigenetic modifier in head and neck squamous carcinoma (HNSC) is the histone methyltransferase KMT2D. KMT2D catalyzes methylation of histone H3K4 resulting in open chromatin and the activation of target genes. Tumor-associated macrophages (TAMs) promote cancer growth by causing T lymphocyte exhaustion. C-C motif chemokine ligand 2 (CCL2) is a potent TAM chemotactic factor. In HNSC, TAMs have been associated with unfavorable patient outcomes and metastasis. The aim of this study was to determine the role of KMT2D in HNSC using genetically engineered in vivo models. KMT2D protein expression was correlated with lymph node metastasis in human HNSC using immunohistochemistry. Genetically engineered KMT2D and CCL2 knockout models of HNSC were created in vivo. HNSC was characterized using qRT-PCR, histopathology, and immunohistochemistry/immunofluorescence microscopy. We also analyzed the effects of KMT2D expression on the proliferation and migration of human HNSC lines. The regulation of the CCL2 gene by KMT2D was characterized using chromatin immunoprecipitation-sequencing assay of transposase accessible chromatin-sequencing, and chromatin conformation capture-sequencing. Human HNSC cases with high KMT2D expression exhibited significantly increased lymph node metastasis. Reduced KMT2D expression in our genetically engineered model correlated with reduced lymph node metastasis, longer latency, and slow tumor growth. CCL2 expression was decreased in KMT2D deficient HNSC, which correlated with a reduced TAM gene expression signature. Genomic experiments demonstrated that KMT2D directly targeted the CCL2 gene. A new genetically engineered in vivo model of CCL2-null HNSC was created, recapitulating the KMT2D deficient phenotype and showing a decreased T lymphocyte exhaustion signature. KMT2D regulates CCL2-mediated immune response and metastasis in HNSC.

Quantifying spatial CXCL9 distribution with image analysis predicts improved prognosis of triple-negative breast cancer.

Background: The C-X-C motif chemokine ligand 9 (CXCL9) plays a pivotal role in tumor immunity by recruiting and activating immune cells. However, the relationship between CXCL9 expression and prognosis in triple-negative breast cancer (TNBC) is unclear. Methods: We investigated CXCL9 mRNA expression, clinicopathological features, and prognosis in TNBC patients. We also used computational image analysis to quantify and assess the distribution of CXCL9 protein in the tumor core (TC) and invasive margin (IM). Results: CXCL9 mRNA expression was significantly higher in TNBC tumors compared to normal tissue (p < 0.001) and was associated with smaller tumors (p = 0.022) and earlier stages (p = 0.033). High CXCL9 mRNA expression was correlated with improved overall survival (OS) in three independent cohorts (all p < 0.05). In a separate analysis, low CXCL9 protein expression was associated with increased lymph node metastasis (p = 0.018 and p = 0.036). High CXCL9 protein expression in the TC, IM, or both was associated with prolonged OS (all p < 0.001). Conclusion: High CXCL9 expression, at both the mRNA and protein levels, is associated with improved prognosis in TNBC patients. CXCL9 expression in the TC and/or IM may be an independent prognostic factor.

Circ_0000592 facilitates the progression of esophageal squamous cell carcinoma via miR-155-5p/FZD5 axis.

Esophageal squamous cell carcinoma (ESCC) is one of the gastrointestinal malignancies with high prevalence and poor prognosis. Previous reports suggested that circular ribose nucleic acidsmight exert regulatory functions in ESCC. This study aims to explore the role of circ_0000592 in ESCC progression, providing novel insights into the diagnosis and therapeutic avenues for ESCC. The GSE131969 data set was utilized to assess circ_0000592 expression in ESCC. The validation was performed in the tumorous tissues of ESCC patients (n = 80) and human-immortalized ESCC cell lines. The correlation between circ_0000592 expression and prognosis was analyzed. The impact of circ_0000592 on ESCC cell activity was evaluated through downregulating circ_0000592, as well as encompassing cell viability, migration, and invasion abilities. The downstream pathway of circ_0000592 was explored by binding site prediction from the TargetScan database, followed by in vitro and in vivo experiments. An in vivo xenograft tumor model was established to highlight the role of circ_0000592 in ESCC. Patients with ESCC exhibited higher circ_0000592 expression levels compared to noncancerous patients, which were associated with reduced survival time, higher TNM stage, and increased lymph node metastasis. The circ_0000592 downregulation suppressed cell viability, migration, and invasion abilities in vitro. Mechanistically, circ_0000592 countered the inhibitory effects on the target gene Frizzled 5 (FZD5) through interactions with miR-155-5p. The overexpression of miR-155-5p curtailed the luciferase activity of circ_0000592 in ESCC cells, inhibiting downstream molecule FZD5 protein expression and subsequently mitigating the detrimental consequences of escalated circ_0000592 expression in ESCC cells. Consistently, circ_0000592 downregulation curbed proliferation and metastasis of ESCC tumors in vivo. In summary, circ_0000592 promoted the progress of ESCC by counteracting the inhibitory impact on FZD5 through its interaction with miR-155-5p. Together, our findings highlighted circ_0000592 as a prospective therapeutic target for ESCC.

The interaction of PRDX1 with Cofilin promotes oral squamous cell carcinoma metastasis.

Peroxiredoxin 1 (PRDX1) is an important member of the peroxiredoxin family (PRDX) and is upregulated in a variety of tumors. Previous studies have found that high PRDX1 expression is closely related to the metastasis of oral squamous cell carcinoma (OSCC), but the specific molecular mechanism is elusive. To elucidate the role of PRDX1 in the metastasis process of OSCC, we evaluated the expression of PRDX1 in OSCC clinical specimens and its impact on the prognosis of OSCC patients. Then, the effect of PRDX1 on OSCC metastasis and cytoskeletal reconstruction was explored in vitro and in nude mouse tongue cancer models, and the molecular mechanisms were also investigated. PRDX1 can directly interact with the actin-binding protein Cofilin, inhibiting the phosphorylation of its Ser3 site, accelerating the depolymerization and turnover of actin, promoting OSCC cell movement, and aggravating the invasion and metastasis of OSCC. In clinical samples and mouse tongue cancer models, PRDX1 also increased lymph node metastasis of OSCC and was negatively correlated with the phosphorylation of Cofilin; PRDX1 also reduced the overall survival rate of OSCC patients. In summary, our study identified that PRDX1 may be a potential therapeutic target to inhibit OSCC metastasis.

RBMS1 Coordinates with the m6A Reader YTHDF1 to Promote NSCLC Metastasis through Stimulating S100P Translation.

Metastasis is the leading cause for the high mortality of lung cancer, however, effective anti-metastatic drugs are still limited. Here it is reported that the RNA-binding protein RBMS1 is positively associated with increased lymph node metastasis in non-small cell lung cancer (NSCLC). Depletion of RBMS1 suppresses cancer cell migration and invasion in vitro and inhibits cancer cell metastasis in vivo. Mechanistically, RBMS1 interacts with YTHDF1 to promote the translation of S100P, thereby accelerating NSCLC cell metastasis. The RRM2 motif of RBMS1 and the YTH domain of YTHDF1 are required for the binding of RBMS1 and YTHDF1. RBMS1 ablation inhibits the translation of S100P and suppresses tumor metastasis. Targeting RBMS1 with NTP, a small molecular chemical inhibitor of RBMS1, attenuates tumor metastasis in a mouse lung metastasis model. Correlation studies in lung cancer patients further validate the clinical relevance of the findings. Collectively, the study provides insight into the molecular mechanism by which RBMS1 promotes NSCLC metastasis and offers a therapeutic strategy for metastatic NSCLC.

Prognostic Impact of Malnutrition Diagnosed by the GLIM Criteria for Resected Extrahepatic Cholangiocarcinoma.

Recently, the Global Leadership Initiative on Malnutrition (GLIM), which includes the world's leading clinical nutrition societies, proposed the first global diagnostic criteria for malnutrition. However, the association between malnutrition diagnosed by the GLIM criteria and prognosis in patients with resected extrahepatic cholangiocarcinoma (ECC) remains unknown. This study aimed to investigate the predictive validity of the GLIM criteria for the prognosis of patients with resected ECC. Between 2000 and 2020, 166 patients who underwent curative-intent resection for ECC were retrospectively analyzed. Prognostic significance of preoperative malnutrition diagnosed by the GLIM criteria was investigated using a multivariate Cox proportional hazards model. Eighty-five (51.2%) and 46 (27.7%) patients were diagnosed with moderate and severe malnutrition, respectively. Increased malnutrition severity tended to be correlated with increased lymph node metastasis rate (p-for-trend=0.0381). The severe malnutrition group had worse 1-, 3-, and 5-year overall survival rates than the normal (without malnutrition) group (82.2% vs. 91.2%, 45.6% vs. 65.1%, 29.3% vs. 61.5%, respectively, p=0.0159). In multivariate analysis, preoperative severe malnutrition was an independent predictor for poor prognosis (hazard ratio=1.68, 95% confidence interval=1.06-2.66, p=0.0282), along with intraoperative blood loss >1,000 ml, lymph node metastasis, perineural invasion, and curability. Severe preoperative malnutrition diagnosed by the GLIM criteria was associated with poor prognosis in patients who underwent curative-intent resection for ECC.

Androgen Receptor Activation Induces Senescence in Thyroid Cancer Cells.

Thyroid cancer (TC) is the most common endocrine malignancy, with an approximately three-fold higher incidence in women. TCGA data indicate that androgen receptor (AR) RNA is significantly downregulated in PTC. In this study, AR-expressing 8505C (anaplastic TC) (84E7) and K1 (papillary TC) cells experienced an 80% decrease in proliferation over 6 days of exposure to physiological levels of 5α-dihydrotestosterone (DHT). In 84E7, continuous AR activation resulted in G1 growth arrest, accompanied by a flattened, vacuolized cell morphology, with enlargement of the cell and the nuclear area, which is indicative of senescence; this was substantiated by an increase in senescence-associated β-galactosidase activity, total RNA and protein content, and reactive oxygen species. Additionally, the expression of tumor suppressor proteins p16, p21, and p27 was significantly increased. A non-inflammatory senescence-associated secretory profile was induced, significantly decreasing inflammatory cytokines and chemokines such as IL-6, IL-8, TNF, RANTES, and MCP-1; this is consistent with the lower incidence of thyroid inflammation and cancer in men. Migration increased six-fold, which is consistent with the clinical observation of increased lymph node metastasis in men. Proteolytic invasion potential was not significantly altered, which is consistent with unchanged MMP/TIMP expression. Our studies provide evidence that the induction of senescence is a novel function of AR activation in thyroid cancer cells, and may underlie the protective role of AR activation in the decreased incidence of TC in men.

Differential expression of PDL-1 and tumour-associated macrophages in N0 and N+ penile cancer.

12 Background: Penile squamous cell carcinoma (PSCC) is rare, representing 0.4-0.6% of all male malignancies in the global-north, as compared with 2-10% in the global-south. Overall survival (OS) dramatically decreases with increased lymph node metastases (LNM). Increased prevalence of pro-tumorigenic M2 CD163+ tumour associated macrophages (TAM) is associated with poor PSCC OS, whereas increased prevalence of the generic M1/2 CD68+ TAMs is associated with better PSCC OS. PDL-1 expression has been noted in 40-60% of PSCC primary lesions. However, there is a paucity of evidence on the overall tumour immune microenvironment (TME) within penile LNM disease. This study describes the relationship between PDL-1 expression and TAMs within the TME of the primary tumour and paired LNMs, with the goal of predicting the utility of PD-1 inhibitors in clinical trials in PSCC. Methods: The Ventana Discovery ULTRA platform was used to optimise a 7-marker multiplex immunofluorescence panel on whole FFPE sections of primary and LNM lesions. Multispectral images were captured using a Vectra 3 microscope. Single cell spatial analysis was performed using HALO software, with statistical analysis of data using R 4.2.1. Results: 52 cases with differing LN stages were identified: N0 (N=20); N1/N2 (N=21); N3 (N=11). Increased expression of PDL-1 was noted at the primary lesion invasive margin (IM) of N+, compared with N0 patients. High expression in inguinal LNMs (iLN+) and pelvic LNMs (pLN+) was also observed, with no expression detected in microscopically normal, tumour free LNs (LN-) in the N0 cohort. More CD163+ M2 TAMs were observed at the IM of N+ compared with N0. However, no significant difference was observed between the three LN groups. M1/2 CD68+ TAMs were more prevalent within pLN+ vs iLN+ or LN-. There was no significant difference in M1/2 CD68+ TAMs number at the IM. Conclusions: Our data highlights increased prevalence of M2 CD163+ TAMs within the IM is associated with LNM. Increased presence of CD68+ M1/2 TAMs in pLN+ may represent active inflammation at the leading microscopic edge of metastatic deposits. A significant difference in the expression of PDL-1 between localised and metastatic disease at both primary and LN location is noted. High PDL-1 expression at the IM of the primary lesion is associated with increased LNMs, where PDL-1 expression is significantly higher as compared to LN-. This study supports the rationale for the use of anti-PD-1/PDL-1 therapies in patients with LNM disease.

MiR-155 rs767649 T>A gene polymorphism is associated with downregulation of miR-155 expression, suppressor of cytokine signaling-1 overexpression, and low probability of metastatic tumor at the time of breast cancer diagnosis.

MicroRNA-155 is a key player in inflammatory reactions, carcinogenesis, and tumor development. In this study, polymorphism of miRNA-155 rs767649 T>A and its gene and suppressor of cytokine signaling-1 (SOCS-1) expression were investigated in relation to cancer susceptibility and development in breast cancer (BC) patients. Polymorphism of miRNA-155 rs767649 T>A was evaluated between a population of 174 patients with BC and 129 controls using restriction fragment length polymorphism and the expression of miR-155 and SOCS-1 were examined in peripheral blood mononuclear cells (PBMCs) by real-time polymerase chain reaction. TT genotype of miR-155 rs767649 T>A was associated with higher level of miR-155 in PBMCs of BC patients relative to AT and AA genotypes (21.76 ± 4.4, 4.046 ± 1.35, 2.56 ± 0.81, respectively; P < 0.001) and increased lymph node metastasis (r = 0.292, P = 0.001), not BC susceptibility (P = 0.402 and P = 0.535; respectively). TT genotype of miR-155 rs767649 T>A was associated with less gene expression of SOCS-1 in PBMCs of BC patients compared to AT and AA genotypes (1.173 ± 0.57, 0.92 ± 0.827, 5.512 ± 0.92, respectively; P = 0.003). This study demonstrated for the first time the association between the T allele of the rs767649 T>A polymorphism in the pre-MIR155 gene and higher expression of miR-155, lower expression of SOCS-1, and swift latent progression in newly diagnosed BC patients. Thus, miR-155 may play a critical role in BC pathogenesis.

The Effect of the Mucinous Component Presence on the Clinical Outcomes of Colorectal Cancer

Background. The effect of colorectal cancer (CRC) histological subtypes on the prognosis is still a controversial issue. We aimed to compare clinical findings, histopathologic data, and survival outcomes in CRC patients with classical and mucinous subtypes.&#x0D; Methods. Patients who were operated on for CRC between 2010 and 2017 were included in the study. Patients were classified into two groups according to the presence of a mucinous component: mucinous adenocarcinoma (MAC) - mucinous component &gt; 50% and classical adenocarcinoma (CAC). Clinical and histopathologic findings, recurrence, metastasis, and survival rates were compared.&#x0D; Results. Data of the 484 CRC patients were documented. Sixty-nine patients (14.3%) were in the MAC group and 415 (85.7%) patients were in the CAC group. The mean age of patients with MAC and CAC was 63.4 ± 13.5 and 68.5 ± 12.7 years, respectively (p = 0.002). Proximal colon localization was found in 30 (43.5%) MAC patients and 123 (29.6%) CAC patients (p = 0.029). The number of patients with metastatic lymph nodes was higher in the MAC group (58% vs. 41.2%, p = 0.03). Nevertheless, there was no significant difference between the CAC and MAC groups in terms of disease-free survival (63.1% vs. 69.6%, p = 0.37) and disease-related mortality (23.6% vs. 23.2%, p = 0.94) over the follow-up period. Multivariate analysis showed that the presence of perineural invasion, patient’s age, and disease stage were associated with mortality in CRC patients.&#x0D; Conclusions. MACs occurred at a younger age than CACs and were more likely localized in the proximal colon as compared to CACs. Despite increased lymph node metastasis in MAC patients, no statistical significance was detected in overall survival or disease-free survival. Multivariate analysis revealed that age, perineural invasion, and disease stage were relevant to mortality in CRC patients.

The oncogenic role of TFAP2A in bladder urothelial carcinoma via a novel long noncoding RNA TPRG1-AS1/DNMT3A/CRTAC1 axis

BackgroundOverexpression of TFAP2A has been linked to increased lymph node metastasis in basal-squamous bladder cancer. However, its downstream targets in bladder urothelial carcinoma (BLCA), the most malignant cancer of the urinary tract, remain unclear. In the current study, we aim to explore the function and mechanism of TFAP2A in BLCA. MethodsTFAP2A expression and the prognostic significance in BLCA was analyzed using TCGA and GTEX projects. TFAP2A was knocked-down in BLCA cells to study its impact on glucose uptake, lactate and ATP production, expression of HK2, and the number of vascular meshes formed by HUVEC. The target long noncoding RNAs (lncRNAs) of TFAP2A were predicted by bioinformatics tools, followed by ChIP-qPCR and luciferase assays. The downstream targets of TPRG1-AS1 were analyzed by microarray analysis. Rescue experiments were conducted for validation. ResultsTFAP2A upregulation in BLCA predicted dismal survival of patients. Loss of TFAP2A inhibited glycolysis (as evidenced by reduced glucose uptake, lactate, ATP production, and the expression of HK2) and angiogenesis (decreased number of vascular meshes formed by HUVEC). TFAP2A promoted the transcription of TPRG1-AS1. TPRG1-AS1 reversed the inhibitory effect of TFAP2A knockdown on glycolysis and angiogenesis in BLCA cells. TPRG1-AS1 inhibited the transcription of CRTAC1 by recruiting a DNA methyltransferase to the promoter of CRTAC1 and increasing the DNA methylation of its promoter. CRTAC1 inhibited glycolysis and angiogenesis in BLCA cells. TFAP2A silencing curbed tumor growth in vivo via the TPRG1-AS1/CRTAC1 axis. ConclusionTFAP2A reduces CRTAC1 expression by promoting TPRG1-AS1 transcription, thereby expediting BLCA glycolysis and angiogenesis.

GATA6 regulates expression of annexin A10 (ANXA10) associated with epithelial–mesenchymal transition of oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) can disturb oral function and quality of life and is associated with poor survival, likely due to the development of cervical lymph node metastases. Epithelial-mesenchymal transition (EMT) is a process in which cells acquire molecular alterations that facilitate cell motility and invasion, and has been associated with tumor metastasis. EMT changes also play important roles in the induction of lymph node metastasis in OSCC. GATA6 is known as the earliest marker of the primitive endoderm lineages. GATA6 inhibits de-differentiation and EMT in human pancreatic ductal adenocarcinoma cells and promotes EMT. However, in OSCC, the expression and function of GATA6 in EMT and lymph node metastasis remains unclear. Therefore, this study aimed to clarify the targets of GATA6 in OSCC cells and whether the change in GATA6 expression affects EMT in OSCC cells, as well as the association between GATA6 and lymph node metastasis. The results showed that GATA6 knockdown OSCC cells promoted EMT and increased lymph node metastasis compared with control cells, whereas the overexpression of GATA6 inhibited the induction of EMT and reduced lymph node metastasis. In addition, annexin A10 (ANXA10) which is the largest type of Ca2+-regulated phospholipid-binding protein in eukaryotic cells was detected as a target gene for GATA6 and ANXA10 suppressed Vimentin expression in EMT in OSCC. Therefore, the GATA6/ANXA10 cascade may be a potential therapeutic approach for the treatment of lymph node metastases in OSCC patients.

Prognostic Value of TRPM7 Expression and Factor XIIIa-Expressing Tumor-Associated Macrophages in Gastric Cancer.

Purpose TRPM7 is known to play a key role in tumor progression by regulating cellular proliferation, migration, and invasion in various cancer cell lines. However, there are no comprehensive clinical studies about the effect of TRPM7 expression on gastric cancer (GC) prognosis. In this study, it was aimed at investigating the effect of TRPM7 expression on prognosis in GC patients. Additionally, for the first time, it was investigated whether the density of Factor XIIIa-expressing tumor-associated macrophages (TAMs) in GC has an effect on the biological behaviour of the tumor. Methods TRPM7 expression and Factor XIIIa-expressing TAM density were immunohistochemically evaluated in paraffin-embedded tumor tissues of 204 GC patients undergoing surgery at a single institution. Results Tumor size was clearly higher in cases with high TRPM7 expression than those with low expression (p < 0.001, Mann-Whitney U). TRPM7 overexpression was closely related to high depth of tumor invasion (p < 0.001, ANOVA), increased lymph node metastasis (p < 0.001, ANOVA), and high distant metastasis rate (p < 0.001, Mann-Whitney U). These findings exposed that high TRPM7 expression is effective in the progression and aggressiveness of GC. In addition, while high CD8+ TIL density affects the prognosis positively, it was determined that high Factor XIIIa+ TAM density negatively affects the prognosis of patients with GC. Furthermore, multivariate analyses revealed TRPM7 overexpression was independently related with short overall (HR 9.64, 95% CI 5.74–16.19, p < 0.001) and disease-free survival (HR 5.67, 95% CI 3.61-8.92, p < 0.001) in GC patients. Conclusions Our data suggest that high TRPM7 expression is closely related to progressive tumor behaviour in GC and independently negatively affects survival in patients. In addition, it was determined that a high density of Factor XIIIa+ TAMs negatively affects the prognosis of patients with GC.

Tetranucleotide and Low Microsatellite Instability Are Inversely Associated with the CpG Island Methylator Phenotype in Colorectal Cancer.

Simple SummaryA type of DNA mismatch repair defect known as “elevated microsatellite alterations at selected tetranucleotide repeats” (EMAST) is found across many different cancers. Tetranucleotide microsatellite instability, which is caused by MSH3 mismatch repair gene/protein loss-of-function, shares a molecular basis with “low microsatellite instability” (MSI-L) in colorectal cancer. Tetranucleotide microsatellite instability is also a byproduct of “high microsatellite instability” (MSI-H) that arises from deficiency of mismatch repair due to MSH2, MSH6, MLH1 or PMS2 gene alterations. MSH3-related EMAST is emerging as a biomarker of poor prognosis in colorectal cancer and needs to be clearly differentiated from MSI-H. Here, we show that tumours with non-MSI-H-related EMAST or MSI-L rarely show concordant promoter methylation of multiple marker genes. Colorectal tumours that are positive for a single (1/5) tetranucleotide repeat marker are an important subset of the EMAST spectrum.MSH3 gene or protein deficiency or loss-of-function in colorectal cancer can cause a DNA mismatch repair defect known as “elevated microsatellite alterations at selected tetranucleotide repeats” (EMAST). A high percentage of MSI-H tumors exhibit EMAST, while MSI-L is also linked with EMAST. However, the distribution of CpG island methylator phenotype (CIMP) within the EMAST spectrum is not known. Five tetranucleotide repeat and five MSI markers were used to classify 100 sporadic colorectal tumours for EMAST, MSI-H and MSI-L according to the number of unstable markers detected. Promoter methylation was determined using methylation-specific PCR for MSH3, MCC, CDKN2A (p16) and five CIMP marker genes. EMAST was found in 55% of sporadic colorectal carcinomas. Carcinomas with only one positive marker (EMAST-1/5, 26%) were associated with advanced tumour stage, increased lymph node metastasis, MSI-L and lack of CIMP-H. EMAST-2/5 (16%) carcinomas displayed some methylation but MSI was rare. Carcinomas with ≥3 positive EMAST markers (13%) were more likely to have a proximal colon location and be MSI-H and CIMP-H. Our study suggests that EMAST/MSI-L is a valuable prognostic and predictive marker for colorectal carcinomas that do not display the high methylation phenotype CIMP-H.

The frequency and clinicopathological significance of NRAS mutations in primary cutaneous nodular melanoma in Indonesia.

BackgroundMelanoma is a lethal skin malignancy with a high risk of metastasis, which prompts a need for research on treatment targets and prognostic factors. Recent studies show that the presence of neuroblastoma RAS viral oncogene homolog (NRAS) mutation can influence cell growth in melanomas. The NRAS mutation, which stimulates the mitogen‐activated protein kinase (MAPK) signaling pathway, is associated with a lower survival rate. However, evidence from Indonesia population is still very rare. Further understanding of the role of NRAS mutations in Indonesian melanoma cases will be crucial in developing new management strategies for melanoma patients with NRAS mutations.AimsTo explore the frequency of NRAS mutations and their clinicopathological associations in patients with primary nodular cutaneous melanoma in Central Java and Yogyakarta, Indonesia.Methods and resultsFifty‐one paraffin‐embedded tissue samples were collected from primary nodular skin melanoma cases between 2011 and 2019 from the two largest referral hospitals in Yogyakarta and Central Java, Indonesia. The NRAS mutation status was evaluated using qualitative real‐time polymerase chain reaction (qRT‐PCR). The association of NRAS mutation was analyzed with the following: age, gender, location, lymph node metastasis, ulceration, mitotic index, tumor‐infiltrating lymphocytes (TILs), necrosis, tumor thickness, lymphovascular invasion (LVI), and tumor size. NRAS mutations were detected in 10 (19.6%) samples and predominantly observed (60%) in exon 2 (G12). These mutations were significantly correlated with lymph node metastases (p = .000); however, they were not associated with other variables analyzed in this study.ConclusionsThe prevalence of NRAS mutations in primary nodular cutaneous melanoma cases from Indonesia is consistent with previous studies and is significantly associated with increased lymph node metastases. However, the predominant mutation detected in exon 2 (G12) is different from previous studies conducted in other countries. This suggests that melanoma cases in Javanese people have different characteristics from other ethnicities.

E3 Ubiquitin Ligase UBR5 Promotes the Metastasis of Pancreatic Cancer via Destabilizing F-Actin Capping Protein CAPZA1.

The ubiquitin-proteasome system (UPS) is a regulated mechanism of intracellular protein degradation and turnover, and its dysfunction is associated with various diseases including cancer. UBR5, an E3 ubiquitin ligase, is emerging as an important regulator of the UPS in cancers, but its role in pancreatic cancer is poorly understood. Here, we show that UBR5 is significantly upregulated in pancreatic cancer tissues. High UBR5 expression is correlated with increased lymph node metastasis and poor survival of patients. The loss-of-function and gain-of-function studies demonstrated that UBR5 substantially enhanced the in vitro migratory and invasive ability of pancreatic cancer cells. UBR5 knockdown also markedly inhibited in vivo cancer metastasis in the liver metastatic model of pancreatic cancer in nude mice, suggesting UBR5 as a potent metastatic promoter in pancreatic cancer. Furthermore, using co-immunoprecipitation combined with mass spectrometry analyses, CAPZA1, a member of F-actin capping protein α subunit family, was identified as a novel substrate of UBR5. UBR5 overexpression could promote the degradation of CAPZA1 via the UPS and induce the accumulation of F-actin, which has been described as an essential molecular event during the process of CAPZA1 deficiency-induced cancer cells migration and invasion. UBR5 knockdown significantly increased the intracellular level of CAPZA1 and CAPZA1 downregulation largely reversed the UBR5 knockdown-induced suppression of cell migration and invasion in pancreatic cancer cells. Collectively, our findings unveil UBR5 as a novel and critical regulator of pancreatic cancer metastasis and highlight the potential for UBR5-CAPZA1 axis as a therapeutic target for preventing metastasis in pancreatic cancer patients, especially in those with increased UBR5 expression.