Extracellular vesicles:General features and usefulness in diagnosis and therapeutic management of colorectal cancer

In the world,among all type of cancers,colorectal cancer(CRC)is the third most commonly diagnosed in males and the second in females.In most of cases,(RP1)patients’prognosis limitation with malignant tumors can be attributed to delayed diagnosis of the disease.Identification of patients with early-stage disease leads to more effective therapeutic interventions.Therefore,new screening methods and further innovative treatment approaches are mandatory as they may lead to an increase in progression-free and overall survival rates.For the last decade,the interest in extracellular vesicles(EVs)research has exponentially increased as EVs generation appears to be a universal feature of every cell that is strongly involved in many mechanisms of cell-cell communication either in physiological or pathological situations.EVs can cargo biomolecules,such as lipids,proteins,nucleic acids and generate transmission signal through the intercellular transfer of their content.By this mechanism,tumor cells can recruit and modify the adjacent and systemic microenvironment to support further invasion and dissemination.This review intends to cover the most recent literature on the role of EVs production in colorectal normal and cancer tissues.Specific attention is paid to the use of EVs for early CRC diagnosis,follow-up,and prognosis as EVs have come into the spotlight of research as a high potential source of‘liquid biopsies’.The use of EVs as new targets or nanovectors as drug delivery systems for CRC therapy is also summarized.

The ubiquitin ligase Siah2 is revealed as an accomplice of the androgen receptor in castration resistant prostate cancer

The androgen receptor （AR） remains the primary molecular target forprostate cancer （PCa） treatment and for development of novel therapies. Profiling and other analyses of human prostate adenocarcinoma have shown that the AR is still functional during latestage disease in the absence of circulating hormone fol lowing castration therapy. The molecular mechanisms that operate during this ＇cas tration resistant＇ phase are still not well understood. Qi et al. have now implicated the ubiquitin ligase Siah2 as an important mediator of AR action in castration resis tant prostate cancer （CRPC）. Siah2 was found to target repressed AR chromatin complexes for degradation, resulting in activation of ARregulated genes involved in tumor cell proliferation, cell motility and lipid metabolism. The authors show a requirement for Siah2 activity for PCa cell growth under conditions of low androgen, and also that targeting Siah2 results in tumor growth suppression under castrate conditions. These findings identify a new mechanism of AR regulation in progressing disease as well as a novel enzymatic target for therapeutic intervention. The AR is a wellstudied hormone receptor that belongs to the large nuclear receptor gene superfamily. AR is activated by androgen binding （principally 5：~dihydrotestosterone, 5DHT）, which elicits changes in AR resident cytosolic complexes, translocation of the receptor to cell nuclei, formation of multiprotein transcriptional complexes onchromatin, and activation or repression of gene expression. Experiments with in vivo model systems have provided evidence that virtually all physiologic processes affected by androgens require the AR as a mediator of molecular effects at the gene level.1 AR is widely expressed beyond the reproductive systems and is believed to play important roles in several nonreproductive tissues, including muscle and brain. The fact that PCa growth is initially dependent on the pre sence of androgens in the circulation, and that prostate tumors will regress temporarily with castration, has been recognized for over half a century. Molecular cloning was initially felt to of the AR in the 1980s be a dispositive step toward pharmacological interventions that would greatly improve treatment outcomes for PCa. However, despite many advances since that time, there is currently no effec tive therapy for disease that has become unresponsive to treatment with hormone ablation. The ＇hormone refractory＇ phase of the disease is intriguing from a bioche mical perspective because the AR still appears to play a critical role under conditions where androgen concentrations in the blood are extremely low. RNA and gene profiling of many human PCa tumors has shown that hormone suppression provides a strong selec tion pressure that results in overexpression and/or amplification of the AR during meta static dissemination.2 Molecular and bio chemical studies of the AR have revealed a bewildering level of complexity involving over 150 protein partners. The AR is also posttranslationally modified by phosphory lation, sumoylation and acetylation, and AR expression can be controlled at transcrip tional and posttranscriptional levels, adding additional layers of regulatory complexity. One important locus of AR activity lies in the ubiquitin/proteasome pathway, which controls the specificity and rate of protein degradation. AR stability was shown pre viously to be regulated by ubiquitin ligases,3 proteins that form complexes with ubiquitin conjugating enzymes to catalyze attachment of the small protein ubiquitin to lysines on a protein target, thereby directing the modified protein to the proteasome for degradation. Siahl and Siah2 are RING finger E3 ubiquitin ligases that regulate ubiquitinationmediated degradation of a range of signaling proteins, resulting in diverse biological effects such as resistance to apoptosis and effects on mito chondrial function. Qi et al.4 previously showed that knockout of the Siah2 gene in the TRAMP transgenic mouse model of PCa, a system that rapidly produces aggres sive autochthonous prostate tumors that pro gress to metastasis, resulted in suppression of tumor formation. In a recent paper in Cancer Cell,5 the same group has now gone on to uncover the mechanism of this surprising effect. Further experiments in TRAMP mice indi cated that loss of Siah2 decreased prostate size, an indication of a loss of AR signaling, and also increased the sensitivity to castra tion, suggesting the possibility that Siah2 may operate under low androgen conditions. Knockdown of Siah2 in PCa cell lines indi cated that Siah2 controls a subset of AR regulated genes, including the gene encoding the important clinical biomarker, prostate specific antigen. Global transcriptional profi ling identified a Siah2regulated gene network consisting of almost 1000 genes, about 100 of which were found to be AR regulated. The AR and Siah2dependent genes were mostly associated with lipid, sterol and cholesterol metabolism. Analysis of human PCa profiling

Relationship between Fusobacterium nucleatum,inflammatory mediators and microRNAs in colorectal carcinogenesis

AIM To examine the effect of Fusobacterium nucleatum(F. nucleatum) on the microenvironment of colonic neoplasms and the expression of inflammatory mediators and microRNAs(miRNAs).METHODS Levels of F. nucleatum DNA, cytokine gene mRNA(TLR2, TLR4, NFKB1, TNF, IL1 B, IL6 and IL8), and potentially interacting miRNAs(miR-21-3p, miR-22-3p, mi R-28-5p, miR-34a-5p, miR-135b-5p) were measured by quantitative polymerase chain reaction(qPCR) TaqMan? assays in DNA and/or RNA extracted from the disease and adjacent normal fresh tissues of 27 colorectal adenoma(CRA) and 43 colorectal cancer(CRC) patients. KRAS mutations were detected by direct sequencing and microsatellite instability(MSI) status by multiplex PCR. Cytoscape v3.1.1 was used to construct the postulated miRNA:mRNA interaction network.RESULTS Overabundance of F. nucleatum in neoplastic tissue compared to matched normal tissue was detected in CRA(51.8%) and more markedly in CRC(72.1%). We observed significantly greater expression of TLR4, IL1 B, IL8, and miR-135 b in CRA lesions and TLR2, IL1 B, IL6, IL8, mi R-34 a and miR-135 b in CRC tumours compared to their respective normal tissues. Only two transcripts for miR-22 and miR-28 were exclusively downregulated in CRC tumour samples. The mRNA expression of IL1 B, IL6, IL8 and miR-22 was positively correlated with F. nucleatum quantification in CRC tumours. The mRNA expression of miR-135 b and TNF was inversely correlated. The miRNA:mRNA interaction network suggested that the upregulation of miR-34 a in CRC proceeds via a TLR2/TLR4-dependent response to F. nucleatum. Finally, KRAS mutations were more frequently observed in CRC samples infected with F. nucleatum and were associated with greater expression of miR-21 in CRA, while IL8 was upregulated in MSI-high CRC.CONCLUSION Our findings indicate that F. nucleatum is a risk factor for CRC by increasing the expression of inflammatory mediators through a possible mi RNA-mediated activation of TLR2/TLR4.

Anticancer effect of Psidium guajava(Guava) leaf extracts against colorectal cancer through inhibition of angiogenesis

Objective:To evaluate the anti-angiogenic and anticancer activities of Psidium guajava leaf extracts against angiogenesis-dependent colorectal cancer.Methods:Three extracts were produced using distilled water,ethanol,and n-hexane as solvents.The extracts were physically characterised through gas chromatography–mass spectrometry,ultraviolet–visible spectroscopy,and Fourier transform infrared spectroscopy.Their antioxidant activity was evaluated using the 2,2-diphenyl-1-picrylhydrazyl,total phenolic content,and total flavonoid content assays.To assess their anti-angiogenic activity,cell viability and rat aortic ring assays were conducted,while cell migration,tube formation,colony formation,and VEGF ELISA assays were conducted to elucidate their effects on different aspects of angiogenesis.Molecular docking was used to assess the antiangiogenic potential of some possible compounds in the extracts.Tumour spheroid assay was used to assess the extracts’potential as a treatment for colorectal cancer.Results:The ethanol extract showed the best antioxidant activity.The distilled water and ethanol extracts exhibited more inhibitory activity against EA.hy926 cell viability and aortic ring microvessel growth.In addition,the ethanol extract performed significantly better than the distilled water extract against cell migration and colony formation,and VEGF expression of the cells was suppressed by the ethanol extract.Both the distilled water and ethanol extracts showed significant inhibitory effect on EA.hy926 tube formation and tumour spheroids consisting of EA.hy926 and HCT116 cells.The ethanol extract containedβ-caryophyllene andβ-elemene by phytochemical analysis and subsequent docking studies,which may contribute to its anti-angiogenic activity.Conclusions:The ethanol extract of Psidium guajava has potential in the treatment of colorectal cancer through the inhibition of angiogenesis.

FOLFIRI3-aflibercept in previously treated patients with metastatic colorectal cancer

AIM To evaluate the efficacy and safety of the modified FOLFIRI3-aflibercept as second-line therapy in patients with metastatic colorectal cancer.METHODS This is a retrospective multicenter cohort, evaluating the efficacy and safety of the association of aflibercept with FOLFIRI3(day 1: aflibercept 4 mg/kg, folinic acid 400 mg/m^2, irinotecan 90 mg/m^2, 5-fluorouracil infusion 2400 mg/m^2 per 46 h; day 3: irinotecan 90 mg/m^2) in patients with previously treated metastatic colorectal cancer. The primary endpoint was overall response rate(ORR). Secondary endpoints were disease control rate(DCR), progression-free survival(PFS), overall survival(OS), and safety.RESULTS Among 74 patients treated in four French centers, nine were excluded due to prior use of aflibercept(n = 3), more than one prior treatment line in irinotecanna?ve patients(n = 3), and inadequate liver function(n = 3). In the "irinotecan-na?ve" patients(n = 30), ORR was 43.3% and DCR was 76.7%. Median PFS and OS were 11.3 mo(95%CI: 6.1-29.0) and 17.0 mo(95%CI: 13.0-17.3), respectively. The most common(> 5%) grade 3-4 adverse events were diarrhea(37.9%), neutropenia(14.3%), stomatitis and anemia(10.4%), and hypertension(6.7%). In the "pre-exposed irinotecan" patients(n = 35), 20(57.1%) received ≥ 2 prior lines of treatment. ORR was 34.3% and DCR was 60.0%. Median PFS and OS were 5.7 mo(95%CI: 3.9-10.4) and 14.3 mo(95%CI: 12.8-19.5), respectively.CONCLUSION Minimally modified FOLFIRI has improvement dramatically the FOLFIRI3-aflibercept efficacy, whatever prior use of irinotecan. A prospective randomized trial is warranted to compare FOLFIRI-aflibercept to FOLFIRI3-aflibercept.

Anti-tumour activity and toxicological studies of combination treatment of Orthosiphon stamineus and gemcitabine on pancreatic xenograft model

BACKGROUND Pancreatic cancer is the most aggressive cancer type.Gemcitabine is the first line chemo-drug used for pancreatic cancer but exerts a broad spectrum of organ toxicities and adverse effects in patients.AIM To evaluate the anti-tumour activity and toxicological effects of Orthosiphon stamineus extract formulation(ID:C5EOSEW5050ESA trademarked as Nuvastatic^(TM)),and gemcitabine combination on pancreatic xenograft model.METHODS Mice were randomly divided into six groups of 6 mice each(n=6)and given different treatments for 28 d.The study design consisted of a 2 x 3 factorial treatment structure,with gemcitabine(yes/no)by oral(at 1200 and 400 mg/kg per day).Human pancreatic cancer cells were injected subcutaneously into the flanks of athymic nude mice.C5EOSEW5050ESA(200 or 400 mg/kg per day)was administered orally,while gemcitabine(10 mg/kg per 3 d)was given intraperitoneally either alone or in combination treatment.Histopathological analyses of vital organs,tumour tissues,and incidence of lethality were analysed.Analyses of tumour necrosis and proliferation were determined by haematoxylin-eosin staining and immunohistochemistry for Ki-67,respectively.RESULTS No signs of toxicity or damage to vital organs were observed in all treatment groups compared to the untreated group.C5EOSEW5050ESA at 200 mg/kg and gemcitabine combination had no additive antitumor effects compared to a single treatment.Remarkably,a comparably greater response in a reduction in tumour growth,Ki-67 protein expression,and necrosis was demonstrated by 400 mg/kg of C5EOSEW5050ESA and gemcitabine combination than that of the individual agents.CONCLUSION These results highlighted the synergistic activity of C5EOSEW5050ESA with gemcitabine to reduce pancreatic tumour growth in mice compared to a single treatment.Thus,this study provides valuable insights into using C5EOSEW5050ESA as a complementary treatment with gemcitabine for pancreatic cancer.

Trading in your spindles for blebs： the amoeboid tumor cell phenotype in prostate cancer

Prostate cancer （PCa） remains a principal cause ofmortalityin developed countries.Because no clinical interventions overcome resistance to androgen ablation therapy, management of castration resistance and metastatic disease remains largely untreatable. Metastasis is a multistep process in which tumor cells lose cell-cell contacts, egress from the primary tumor, intravasate, survive shear stress within the vasculature and extravasate into tissues to colonize ectopic sites. Tumor ceils reestablish migratory behaviors employed during nonneoplastic processes such as embryonic development, leukocyte trafficking and wound healing. While mesenchymal motility is an established paradigm of dissemination, an alternate, ＇amoeboid＇ phenotype is increasingly appreciated as relevant to human cancer.

Perspective: plasticity, the enemy of the good

Plasticity is an important feature of modern cancer research.However,the level at which we should consider it remains an open question.Such debate is not new in the field of cancer and can be exemplified by the different models explaining carcinogenesis.Those models mostly explain cell transformation through the deregulation of the internal circuitry.In the last years,those models dramatically increased our knowledge and led to a series of short-term successes in terms of therapeutics.However,cancer drug resistance inevitably arises.Recently,studies on the so-called tumor microenvironment enriched the cell-centered perspective but it also enlarged the complexity of cancer etiology in particular for advanced diseases.Here,we suggest that the plastic and multi-sites specific nature of cancer combined with our incapacity to promise cure should push towards a new perspective where early clinical actions,instead of late ones,should be heralded as the priority of cancer research and care.