IL-2RG as a possible immunotherapeutic target in CRC predicting poor prognosis and regulated by miR-7-5p and miR-26b-5p.

Despite advances in treatment strategies, colorectal cancer (CRC) continues to cause significant morbidity and mortality, with mounting evidence a close link between immune system dysfunctions issued. Interleukin-2 receptor gamma (IL-2RG) plays a pivotal role as a common subunit receptor in the IL-2 family cytokines and activates the JAK-STAT pathway. This study delves into the role of Interleukin-2 receptor gamma (IL-2RG) within the tumor microenvironment and investigates potential microRNAs (miRNAs) that directly inhibit IL-2RG, aiming to discern their impact on CRC clinical outcomes. Bioinformatics analysis revealed a significant upregulation of IL-2RG mRNA in TCGA-COAD samples and showed strong correlations with the infiltration of various lymphocytes. Single-cell analysis corroborated these findings, highlighting IL-2RG expression in critical immune cell subsets. To explore miRNA involvement in IL-2RG dysregulation, mRNA was isolated from the tumor tissues and lymphocytes of 258 CRC patients and 30 healthy controls, and IL-2RG was cloned into the pcDNA3.1/CT-GFP-TOPO vector. Human embryonic kidney cell lines (HEK-293T) were transfected with this construct. Our research involved a comprehensive analysis of miRPathDB, miRWalk, and Targetscan databases to identify the miRNAs associated with the 3' UTR of human IL-2RG. The human microRNA (miRNA) molecules, hsa-miR-7-5p and hsa-miR-26b-5p, have been identified as potent suppressors of IL-2RG expression in CRC patients. Specifically, the downregulation of hsa-miR-7-5p and hsa-miR-26b-5p has been shown to result in the upregulation of IL-2RG mRNA expression in these patients. Prognostic evaluation of IL-2RG, hsa-miR-7-5p, and hsa-miR-26b-5p, using TCGA-COAD data and patient samples, established that higher IL-2RG expression and lower expression of both miRNAs were associated with poorer outcomes. Additionally, this study identified several long non-coding RNAs (LncRNAs), such as ZFAS1, SOX21-AS1, SNHG11, SNHG16, SNHG1, DLX6-AS1, GAS5, SNHG6, and MALAT1, which may act as competing endogenous RNA molecules for IL2RG by sequestering shared hsa-miR-7-5p and hsa-miR-26b-5p. In summary, this investigation underscores the potential utility of IL-2RG, hsa-miR-7-5p, and hsa-miR-26b-5p as serum and tissue biomarkers for predicting CRC patient prognosis while also offering promise as targets for immunotherapy in CRC management.

Platelet-derived microparticles provoke chronic lymphocytic leukemia malignancy through metabolic reprogramming.

Background: It is well established that inflammation and platelets promote multiple processes of cancer malignancy. Recently, platelets have received attention for their role in carcinogenesis through the production of microvesicles or platelet-derived microparticles (PMPs), which transfer their biological content to cancer cells. We have previously characterized a new subpopulation of these microparticles (termed mito-microparticles), which package functional mitochondria. The potential of mitochondria transfer to cancer cells is particularly impactful as many aspects of mitochondrial biology (i.e., cell growth, apoptosis inhibition, and drug resistance) coincide with cancer hallmarks and disease progression. These metabolic aspects are particularly notable in chronic lymphocytic leukemia (CLL), which is characterized by a relentless accumulation of proliferating, immunologically dysfunctional, mature B-lymphocytes that fail to undergo apoptosis. The present study aimed to investigate the role of PMPs on CLL metabolic plasticity leading to cancer cell phenotypic changes. Methods: CLL cell lines were co-incubated with different concentrations of human PMPs, and their impact on cell proliferation, mitochondrial DNA copy number, OCR level, ATP production, and ROS content was evaluated. Essential genes involved in metabolic-reprogramming were identified using the bioinformatics tools, examined between patients with early and advanced CLL stages, and then validated in PMP-recipient CLLs. Finally, the impact of the induced metabolic reprogramming on CLLs' growth, survival, mobility, and invasiveness was tested against anti-cancer drugs Cytarabine, Venetoclax, and Plumbagin. Results: The data demonstrated the potency of PMPs in inducing tumoral growth and invasiveness in CLLs through mitochondrial internalization and OXPHOS stimulation which was in line with metabolic shift reported in CLL patients from early to advanced stages. This metabolic rewiring also improved CLL cells' resistance to Cytarabine, Venetoclax, and Plumbagin chemo drugs. Conclusion: Altogether, these findings depict a new platelet-mediated pathway of cancer pathogenesis. We also highlight the impact of PMPs in CLL metabolic reprogramming and disease progression.

Circular RNA Expression Signatures Provide Promising Diagnostic and Therapeutic Biomarkers for Chronic Lymphocytic Leukemia.

Chronic lymphocytic leukemia (CLL) is a known hematologic malignancy associated with a growing incidence and post-treatment relapse. Hence, finding a reliable diagnostic biomarker for CLL is crucial. Circular RNAs (circRNAs) represent a new class of RNA involved in many biological processes and diseases. This study aimed to define a circRNA-based panel for the early diagnosis of CLL. To this point, the list of the most deregulated circRNAs in CLL cell models was retrieved using bioinformatic algorithms and applied to the verified CLL patients' online datasets as the training cohort (n = 100). The diagnostic performance of potential biomarkers represented in individual and discriminating panels, was then analyzed between CLL Binet stages and validated in individual sample sets I (n = 220) and II (n = 251). We also estimated the 5-year overall survival (OS), introduced the cancer-related signaling pathways regulated by the announced circRNAs, and provided a list of possible therapeutic compounds to control the CLL. These findings show that the detected circRNA biomarkers exhibit better predictive performance compared to current validated clinical risk scales, and are applicable for the early detection and treatment of CLL.

Characterization of exogenous αSN response genes and their relation to Parkinson's disease using network analyses.

Despite extensive research, the molecular mechanisms underlying the toxicity of αSN in Parkinson's disease (PD) pathology are still poorly understood. To address this, we used a microarray dataset to identify genes that are induced and differentially expressed after exposure to toxic αSN aggregates, which we call exogenous αSN response (EASR) genes. Using systems biology approaches, we then determined, at multiple levels of analysis, how these EASR genes could be related to PD pathology. A key result was the identification of functional connections between EASR genes and previously identified PD-related genes by employing the proteins' interactions networks and 9 brain region-specific co-expression networks. In each brain region, co-expression modules of EASR genes were enriched for gene sets whose expression are altered by SARS-CoV-2 infection, leading to the hypothesis that EASR co-expression genes may explain the observed links between COVID-19 and PD. An examination of the expression pattern of EASR genes in different non-neurological healthy brain regions revealed that regions with lower mean expression of the upregulated EASR genes, such as substantia nigra, are more vulnerable to αSN aggregates and lose their neurological functions during PD progression. Gene Set Enrichment Analysis of healthy and PD samples from substantia nigra revealed that a specific co-expression network, "TNF-α signaling via NF-κB", is an upregulated pathway associated with the PD phenotype. Inhibitors of the "TNF-α signaling via NF-κB" pathway may, therefore, decrease the activity level of this pathway and thereby provide therapeutic benefits for PD patients. We virtually screened FDA-approved drugs against these upregulated genes (NR4A1, DUSP1, and FOS) using docking-based drug discovery and identified several promising drugs. Altogether, our study provides a better understanding of αSN toxicity mechanisms in PD and identifies potential therapeutic targets and small molecules for treatment of PD.

The Pleiotropy of PAX5 Gene Products and Function.

PAX5, a member of the Paired Box (PAX) transcription factor family, is an essential factor for B-lineage identity during lymphoid differentiation. Mechanistically, PAX5 controls gene expression profiles, which are pivotal to cellular processes such as viability, proliferation, and differentiation. Given its crucial function in B-cell development, PAX5 aberrant expression also correlates with hallmark cancer processes leading to hematological and other types of cancer lesions. Despite the well-established association of PAX5 in the development, maintenance, and progression of cancer disease, the use of PAX5 as a cancer biomarker or therapeutic target has yet to be implemented. This may be partly due to the assortment of PAX5 expressed products, which layers the complexity of their function and role in various regulatory networks and biological processes. In this review, we provide an overview of the reported data describing PAX5 products, their regulation, and function in cellular processes, cellular biology, and neoplasm.

Can hypoxia-inducible factor-1α overexpression discriminate human colorectal cancers with different microsatellite instability?

Clinicopathological features of high-frequency microsatellite instability (MSI-H) colorectal cancers (CRCs) are different from low-frequency MSI (MSI-L) and microsatellite stable (MSS) CRCs. The clinical features of MSI-L cases are unknown, and although the tumors usually show instability for dinucleotide markers, evaluation based on dinucleotides alone could lead to the misclassification of MSI-L or MSS as MSI-H. In this research, we investigated the usefulness of hypoxia-inducible factor-1α (HIF-1α) expression to discriminate MSI-L from MSS and MSI-H in human CRC. Tumor tissue from 94 CRC patients was used to determine the expression level of HIF-1α mRNA and HIF-1α protein using quantitative real-time PCR and immunohistochemistry analyses, respectively. The results indicated that HIF-1α mRNA and HIF-1α protein levels were upregulated in CRC patients compared with controls (P < 0.0001). Average HIF-1α expression in tissues with advanced stages and grades was also higher than that in earlier stages and grades. Expression of HIF-1α mRNA varied between CRC patients with different types of microsatellite instability (MSS, MSI-L and MSI-H). Taken together, our findings provide preliminary evidence that HIF-1α expression level in CRC tumors correlates with different MSI categories. HIF-1α expression may therefore represent a novel marker to separate the MSI-L group from the MSS and MSI-H groups.

Pax-5 Protein Expression Is Regulated by Transcriptional 3'UTR Editing.

The Pax-5 gene encodes a transcription factor that is essential for B-cell commitment and maturation. However, Pax-5 deregulation is associated with various cancer lesions, notably hematopoietic cancers. Mechanistically, studies have characterized genetic alterations within the Pax-5 locus that result in either dominant oncogenic function or haploinsufficiency-inducing mutations leading to oncogenesis. Apart from these mutations, some examples of aberrant Pax-5 expression cannot be associated with genetic alterations. In the present study, we set out to elucidate potential alterations in post-transcriptional regulation of Pax-5 expression and establish that Pax-5 transcript editing represents an important means to aberrant expression. Upon the profiling of Pax-5 mRNA in leukemic cells, we found that the 3'end of the Pax-5 transcript is submitted to alternative polyadenylation (APA) and alternative splicing events. Using rapid amplification of cDNA ends (3'RACE) from polysomal fractions, we found that Pax-5 3' untranslated region (UTR) shortening correlates with increased ribosomal occupancy for translation. These observations were also validated using reporter gene assays with truncated 3'UTR regions cloned downstream of a luciferase gene. We also showed that Pax-5 3'UTR editing has direct repercussions on regulatory elements such as miRNAs, which in turn impact Pax-5 protein expression. More importantly, we found that advanced staging of various hematopoietic cancer lesions relates to shorter Pax-5 3'UTRs. Altogether, our findings identify novel molecular mechanisms that account for aberrant expression and function of the Pax-5 oncogene in cancer cells. These findings also present new avenues for strategic intervention in Pax-5-mediated cancers.

Identification of a stool long non-coding RNAs panel as a potential biomarker for early detection of colorectal cancer.

BACKGROUND: The feces of colorectal cancer (CRC) patients contain tumor colonocytes, which constantly shed into the lumen area. Therefore, stool evaluation can be considered as a rapid and low-risk way to directly determine the colon and rectum status. As long non-coding RNAs (lncRNAs) alterations are important in cancer cells fate regulation, we aimed to assess the level of a panel of cancer-related lncRNAs in fecal colonocytes. METHODS: The population study consisted of 150 subjects, including a training set, a validation set, and a group of 30 colon polyps. The expression levels of lncRNAs were evaluated by quantitative real-time PCR (qRT-PCR). The NPInetr and EnrichR tools were used to identify the interactions and functions of lncRNAs. RESULTS: A total of 10 significantly dysregulated lncRNAs, including CCAT1, CCAT2, H19, HOTAIR, HULC, MALAT1, PCAT1, MEG3, PTENP1, and TUSC7, were chosen for designing a predictive panel. The diagnostic performance of the panel in distinguishing CRCs from the healthy group was AUC: 0.8554 in the training set and 0.8465 in the validation set. The AUC for early CRCs (I-II TNM stages) was 0.8554 in the training set and 0.8465 in the validation set, and for advanced CRCs (III-IV TNM stages) were 0.9281 in the training set and 0.9236 in the validation set. The corresponding AUC for CRCs vs polyps were 0.9228 (I-IV TNM stages), 0.9042 (I-II TNM stages), and 0.9362 (III-IV TNM stages). CONCLUSIONS: These data represented the application of analysis of fecal colonocytes lncRNAs in early detection of CRC.

Establishment of a CALU, AURKA, and MCM2 gene panel for discrimination of metastasis from primary colon and lung cancers.

Metastasis is known as a key step in cancer recurrence and could be stimulated by multiple factors. Calumenin (CALU) is one of these factors which has a direct impact on cancer metastasis and yet, its underlined mechanisms have not been completely elucidated. The current study was aimed to identify CALU co-expressed genes, their signaling pathways, and expression status within the human cancers. To this point, CALU associated genes were visualized using the Cytoscape plugin BisoGenet and annotated with the Enrichr web-based application. The list of CALU related diseases was retrieved using the DisGenNet, and cancer datasets were downloaded from The Cancer Genome Atlas (TCGA) and analyzed with the Cufflink software. ROC curve analysis was used to estimate the diagnostic accuracy of DEGs in each cancer, and the Kaplan-Meier survival analysis was performed to plot the overall survival of patients. The protein level of the signature biomarkers was measured in 40 biopsy specimens and matched adjacent normal tissues collected from CRC and lung cancer patients. Analysis of CALU co-expressed genes network in TCGA datasets indicated that the network is markedly altered in human colon (COAD) and lung (LUAD) cancers. Diagnostic accuracy estimation of differentially expressed genes showed that a gene panel consisted of CALU, AURKA, and MCM2 was able to successfully distinguish cancer tumors from healthy samples. Cancer cases with abnormal expression of the signature genes had a significantly lower survival rate than other patients. Additionally, comparison of CALU, AURKA, and MCM2 proteins between healthy samples, early and advanced tumors showed that the level of these proteins was increased through normal-carcinoma transition in both types of cancers. These data indicate that the interactions between CALU, AURKA, and MCM2 has a pivotal role in cancer development, and thereby needs to be explored in the future.

Development and validation of a lipogenic genes panel for diagnosis and recurrence of colorectal cancer.

BACKGROUND & AIM: Accumulated evidence indicates that the elevation of lipid metabolism is an essential step in colorectal cancer (CRC) development, and analysis of the key lipogenic mediators may lead to identifying the new clinically useful prognostic gene signatures. METHODS: The expression pattern of 61 lipogenic genes was assessed between CRC tumors and matched adjacent normal tissues in a training set (n = 257) with the Mann-Whitney U test. Cox's proportional hazards model and the Kaplan-Meier method were used to identifying a lipogenic-biomarkers signature associated with the prognosis of CRC. The biomarkers signature was then confirmed in two independent validation groups, including a set of 223 CRC samples and an additional set of 203 COAD profiles retrieving from the Cancer Genome Atlas (TCGA). RESULTS: Five genes, including ACOT8, ACSL5, FASN, HMGCS2, and SCD1, were significantly enhanced in CRC tumors. Using the cutoff value 0.493, the samples were classified into high risk and low risk. The AUC of panel for discriminating of all, early (I-II stages), and advanced CRC (III-IV stages) were 0.8922, 0.8446, and 0.9162 (Training set), along with 0.8800, 0.8205, and 0.7351 (validation set I), and 0.9071, 0.8946, and 0.9107 (Validation set II), respectively. There was a reverse correlation between the high predicted point of panel and worse OS of CRC patients in training set (HR (95% CI): 0.1096 (0.07089-0.1694), P < 0.001), validation set I (HR (95% CI): 0.3350 (0.2116-0.5304), P < 0.001), and validation set II (HR (95% CI): 0.1568 (0.1090-0.2257), P < 0.001). CONCLUSION: Our study showed that the panel of ACOT8/ACSL5/FASN/HMGBCS2/SCD1 genes had a better prognostic performance than validated clinical risk scales and is applicable for early detection of CRC and tumor recurrence.

miR-497-5p mediates starvation-induced death in colon cancer cells by targeting acyl-CoA synthetase-5 and modulation of lipid metabolism.

Modulation of fatty acids metabolism is an appropriate strategy for starvation-induced death in tumor cancers. Colon cancer cells express a high level of acyl-CoA synthetase-5 (ACSL5), and as yet no therapeutic approach has been achieved. Herein, ACSL5-related microRNAs (miRNAs) were identified via TargetScan, and their impacts on ACSL5 and lipid content along with metabolic activity, cell cycle, migration, and invasion of colorectal cancer (CRC) cells were examined, and subsequently compared with transcriptome for better visualization of intracellular-signaling networks. In vivo analysis was performed using BALB/c mice xenograft model of CRC injected with target miRNA. Clinical significances were also evaluated in 80 CRC tumors and matched adjacent normal tissues. There was a reverse correlation between ACSL5 and miR-497-5p, which miR-497-5p overexpression modulated CRC cell proliferation and development. A similar observation was received from the in vivo examination in which intratumoral injection of miR-497-5p reversed the tumor growth in the CRC xenograft model. Downregulation of miR-497-5p correlated with tumor differentiation, tumor, node, and metastasis staging, lymph node metastasis, and poor survival in patients with CRC. These results suggested that miR-497-5p upregulation could be considered as a therapeutic strategy for modulation of lipid metabolism in colon cancer.

miR-298 plays a pivotal role in colon cancer invasiveness by targeting PTEN.

Evidence indicate that the miR-298 dysregulation might associate with colorectal cancer (CRC) development. Herein, we evaluated the effect of miR-298 dysregulation on colon cancer invasiveness. First, metabolic activity, cell cycle progression, apoptosis, and invasion of miR-298 overexpressed/knocked out colon cancer cells were examined and combined with their transcriptome analysis data for better visualization of miR-298 intracellular signaling networks. Interaction between miR-298 and its target was evaluated with luciferase assay and validated using western blot analysis. The proportion of abnormal miR-298 level was investigated in tumor samples, matched normal adjacent tissues, and plasmas of 100 CRC patients, and also compared with 100 normal plasma samples. The Mann-Whitney U test was performed to assess miR-298 differences among the studied groups, and the correlation between miR-298 and the risk of CRC was shown by univariate and multivariate logistic regression. The data indicate that miR-298 overexpression promoted proliferation and metastasis in CRC cells via targeting phosphatase and tensin homolog. Comparative analysis of CRC tumors, normal adjacent tissues, and plasmas indicated a significant miR-298 upregulation in tumors and plasmas (1.72-fold and 1.65-fold, respectively; p < .001). Also, the aberrant level of miR-298 contributed with CRC tumor differentiation, TNM stage and lymph node metastasis (p < .001), and independently associated with poor survival of CRC patients (p < .029; hazard ratio: 1.292; 95% confidence interval: 0.339-2.184). Collectively, these data showed that abnormal level of miR-298 correlated with cancer development and through that lowered the overall survival rate of CRC patients. Therefore, miR-298 could be considered as a therapeutic target for CRC.

Study of the Antidiabetic Activity of Punica granatum L. Fruits Aqueous Extract on the Alloxan-Diabetic Wistar Rats.

Pancreatic ß-cells dysfunction and impairment of insulin action usually leads to hyperglycemia. Punica granatum L. is a well-known traditional herbal remedy in Iran due to its positive effects on ameliorating blood glucose homeostasis. In this study, Alloxan-diabetic male Wistar rats were administrated with pomegranate fruits aqueous extract (PE) in different doses of 100, 200, and 350 mg/kg bw (PE+Da, PE+Db and PE+Dc, respectively), and the effects of PE polyphenols content on glucose metabolism in treated groups were examined using oral glucose tolerance test (OGTT), short-term and long-term PE consumption periods models followed by evaluation of plasma insulin, free fatty acids, and triglycerides levels and tissues contents of glycogen and triglycerides; compared with diabetic control (DC) and healthy control (NC) groups. By using Real-time PCR, the possibility of modulations of the Insulin receptor substrate 1 (IRS-1), Protein kinase B (Akt), Glucose transporter 2 and 4 (Glut-2, 4) mRNAs expression levels in PE treated rats were investigated. The obtained data showed noticeable reduction in fasting blood glucose (FBG) by 28.1% and 67.9% in short-term and long-term treatment models, respectively, in PE + Dc group. Also, there existed marked increase in the mRNAs expression levels of IRS-1, Akt, Glut-2, and Glut-4, which results in improvement of glucose uptake and promotes its storage. Taking together, it is suggested that PE administration contributes to the modulation of both hyperglycemia and hyperlipidemia in Alloxan-diabetic Wistar rats.

Investigating the diagnostic performance of HOTTIP, PVT1, and UCA1 long noncoding RNAs as a predictive panel for the screening of colorectal cancer patients with lymph node metastasis.

Like other noncoding RNAs (ncRNAs), dysregulation of long ncRNAs (lncRNAs) has been associated with various clinicopathological features of colorectal cancer (CRC) patients such as lymph node metastasis (LNM). Recently, three aberrant expressed oncogenic lncRNA (onco-lncRNAs), including HOXA transcript at the distal tip (HOTTIP), plasmacytoma variant translocation 1 (PVT1), and urothelial carcinoma associated 1 (UCA1) have been reported in LNM. Herein, we compared the diagnostic performance of these lncRNAs as individual biomarkers and as a discriminating panel between LNM CRC patients, nonmetastatic lymph nodes (NLN) and normal healthy subjects. The lncRNAs expression level was measured by quantitative real-time PCR and analyzed by the Mann-Whitney U test. The receiver operating characteristic (ROC) curve analysis was applied to evaluate the diagnostic power. The Kaplan-Meier survival analysis was performed to outline the overall survival (OS) of CRC patients with an abnormal level of lncRNAs. The area under the ROC curve (AUC) of the overexpressed HOTTIP (0.7817; 95% CI, 0.6809-0.8824), PVT1 (0.8559; 95% CI, 0.7737-0.9382), and UCA1 (0.8135; 95% CI, 0.722-0.9051) introduced them as individual CRC biomarkers. As a predictive panel, the AUC values of the HOTTIP, PVT1, and UCA1 for training set were 0.9256 (95% CI, 0.8634-0.9879; all CRCs), 0.8708 (95% CI, 0.7709-0.9378; NLN) and 0.9804 (95% CI, 0.9585-0.9998; LNM), and for validation set were 0.9286 (95% CI, 0.8752-0.9820; all CRCs), 0.8911 (95% CI, 0.8238-0.9585; NLN), and 0.9833 (95% CI, 0.9642-1.002; LNM), respectively. Also, HOTTIP/PVT1/UCA1 panel dysregulation had a marked correlation with patient's OS in training set (logrank test P = 0.0121; hazard ratio [HR], 0.1225; 95% confidence interval [CI], 0.02376-0.6312), and in validation set (logrank test P < 0.0001, HR, 0.2003; 95% CI, 0.08942-0.4486). These data showed that the combination of HOTTIP, PVT1, and UCA1 as a predictive panel, has a better diagnostic performance than each of these lncRNAs individually, and could be used for the screening of patients with advanced CRC.

A detailed image of rutin underlying intracellular signaling pathways in human SW480 colorectal cancer cells based on miRNAs-lncRNAs-mRNAs-TFs interactions.

Natural dietary ingredients like flavonoids are important for body improvement against diseases. The flavonol rutin is widely found in fruits and vegetables and shows significant anticancer properties. However, the underlined signaling pathways have not been elucidated yet. In this study, the impacts of various doses of rutin (400-700 mM/ml) have been examined on human colon cancer SW480 cells metabolism, cell cycle, and apoptosis. The transcriptome was analyzed by bioinformatics tools and the interactions between rutin modulated microRNAs (miRNAs), long noncoding RNAs (lncRNAs), messenger RNAs (mRNAs), and transcription factors (TFs) were built, filtered and enriched. A dose of 600 mM of rutin significantly decreased cells metabolic activity, halved the population and arrested the cell cycle at the sub-G1 phase. The enrichment analysis of miRNAs-lncRNAs-mRNAs-TFs network showed that these effects were mediated through alteration of glucose, lipid, and protein metabolism, modulating endoplasmic reticulum stress responses, negative regulation of cell cycle process, and inducing the extrinsic and intrinsic apoptotic signaling pathways. Additionally, the key parent nodes of each annotation were illustrated. These findings create a detailed image of rutin underlying intracellular signaling pathways in CRC and also help us to better understand the role of dietary natural compounds in cancer treatment.

HER2+ mCRC patients with exon 20 R784G substitution mutation do not respond to the cetuximab therapy.

The human epidermal growth factor 2 (HER2) gene undergoes various mutations that could alter its activity or respond to the antibody therapies. Cetuximab, a known anti-EGFR monoclonal antibody (mAB), is widely administered in metastatic colorectal cancer (mCRC) cases. Here we identified mCRC patients who did not respond to cetuximab (500 mg/m2 , q2w) after fluoropyrimidine/oxaliplatin regimen failure. Tumor samples were examined with immunohistochemistry for protein distribution, polymerase chain reaction (PCR) sequencing for mutation detection and real-time PCR for mRNA expression pattern analysis between cetuximab sensitive and resistance patients. The conformational differences of normal and mutated protein structures were predicted by bioinformatics analysis. The 5-year survival rates of target groups were estimated using the Kaplan-Meier method. Immunohistochemistry showed that all cases had high level of HER2 protein. No K-Ras or B-Raf mutation was observed among the study population; however, cetuximab resistance patients harbored a somatic mutation R784G at the exon 20 region of HER2 coding sequence. According to bioinformatics analysis, this mutation caused a notable misfold in protein conformation. Meanwhile, survival analysis showed R784G mutated mCRC patients had shortened survival rate compared with the mCRC cases with wild-type HER2. Collectively, these data report a new mechanism of resistance to cetuximab and might be applicable in modifying new therapeutic strategies for HER2 involved cancers.

Intratumoral infiltrating lymphocytes correlate with improved survival in colorectal cancer patients: Independent of oncogenetic features.

BACKGROUND: The clinical relevance and prognostic value of tumor-infiltrating lymphocytes (TILs) as an interplay between malignant cells and immune function has been known for decades. On contrary, this potential may be different by T lymphocytes subsets endowed with a different function. Colorectal cancer (CRC) is a heterogeneous disease with different suggested prognostic biomarkers. So, this study was conducted to examine the prognostic value of CD8+ TILs on the survival rate of CRC as an independent factor of oncogenetic tumor features. METHODS: With respect to this, 281 formalin-fixed, paraffin-embedded tissue samples of Iranian CRC patients were evaluated for clinical features including tumor location, tumor stage, differentiation grade, and mucinous characteristics. Then, using the standard immunohistochemical technique, tumor sections were examined, and CD8+ TILs were counted and identified in two regions of the tumor, including intratumoral (ITCIL TILs) and stromal (S TILs). The prognostic value of CD8+ TILs was determined by comparing with parameters, such as diagnostic age, tumor stage, adjuvant therapy, microsatellite instability (MSI) status, KRAS and BRAF mutations, family history, and survival. RESULTS: The presence of intratumoral tumor cell-infiltrating lymphocytes (ITCIL) CD8+ lymphocytes are significantly associated with differentiation (p = 0.004), tumor, node, and metastases (TNM) stage (p = 0.001), and MSI (p = 0.001). Meanwhile, based on the level of stromal infiltrating lymphocytes (SIL) infiltration, analysis of CRC patients was statistically associated with a location (p = 0.002), TNM stage (p < 0.001), metastasis (p < 0.001), and KRAS mutation (p = 0.031). Also, tumors with severe ITCIL CD8+ lymphocytes have a good prognosis compared with tumors with poor or moderate ITCIL CD8+ lymphocytes. CONCLUSIONS: These results suggest that intratumor cell-infiltrating CD8- T lymphocytes as an independent prognostic factor that have an antitumor activity as judged by their favorable effect on patients' survival and could potentially be exploited in the treatment of CRC.

The Application of Gene Expression Profiling in Predictions of Occult Lymph Node Metastasis in Colorectal Cancer Patients.

A key factor in determining the likely outcome for a patient with colorectal cancer is whether or not the tumour has metastasised to the lymph nodes-information which is also important in assessing any possibilities of lymph node resection so as to improve survival. In this review we perform a wide-range assessment of literature relating to recent developments in gene expression profiling (GEP) of the primary tumour, to determine their utility in assessing node status. A set of characteristic genes seems to be involved in the prediction of lymph node metastasis (LNM) in colorectal patients. Hence, GEP is applicable in personalised/individualised/tailored therapies and provides insights into developing novel therapeutic targets. Not only is GEP useful in prediction of LNM, but it also allows classification based on differences such as sample size, target gene expression, and examination method.

Punica granatum L. Fruit Aqueous Extract Suppresses Reactive Oxygen Species-Mediated p53/p65/miR-145 Expressions followed by Elevated Levels of irs-1 in Alloxan-Diabetic Rats.

OBJECTIVES: Reactive oxygen species (ROS) is an apoptosis inducer in pancreatic ß-cells that stimulates p53/p65 mediated microRNA (miR)-145 expression. Punica granatum L. (pomegranate) is an antioxidant fruit that attenuates ROS generation. This study examines the effects of pomegranate fruit aqueous extract (PGE) on the levels of ROS, p53, p65, miR-145, and its target insulin receptor substrate 1 (irs-1) mRNA in Alloxan-diabetic male Wistar rats. MATERIALS AND METHODS: In this experimental study, diabetic rats received different doses of PGE. The effects of the PGE polyphenols were examined through a long-term PGE treatment period model, followed by an evaluation of the plasma and tissue contents of free fatty acids (FFAs), triglycerides (TG), and glycogen compared with diabetic controls (DC) and normal controls (NC). We used real-time polymerase chain reaction (PCR) to investigate the modulation of p53, p65, miR-145, and irs-1 expression levels. RESULTS: There was a noticeable reduction in fasting blood glucose (FBG) and ROS generation compared to DC. We observed marked decreases in p53, p65, miR-145 expression levels followed by an elevated level of irs-1, which contributed to improvement in insulin sensitivity. CONCLUSIONS: PGE administration downregulated miR-145 levels in Alloxan-diabetic Wistar rats by suppression of ROS-mediated p53 and p65 overexpression.

Coexistence of KRAS and BRAF Mutations in Colorectal Cancer: A Case Report Supporting The Concept of Tumoral Heterogeneity.

The detection of KRAS and BRAF mutations is a crucial step for the correct therapeutic approach and predicting the epidermal growth factor receptor (EGFR)-targeted therapy resistance of colorectal carcinomas. The concomitant KRAS and BRAF mutations occur rarely in the colorectal cancers (CRCs) with the prevalence of less than 0.001% of the cases. In patients with KRAS-mutant tumors, BRAF mutations should not regularly be tested unless the patient is participating in a clinical trial enriching for the presence of KRAS or BRAF-mutated tumor. The current report demonstrates a case with advanced adenocarcinoma of the colon showing the coexistence of KRAS and BRAF mutations and may have profound clinical implications for disease progression and therapeutic responses.