Systematic review of the old and new concepts in the epithelial-mesenchymal transition of colorectal cancer

Epithelial-to-mesenchymal transition(EMT) is defined as the transformation of an epithelial cell into a spindle cell with the loss of membrane E-cadherin expression and the gain of mesenchymal markers positivity. In the field of colorectal cancer(CRC), first data about EMT was published in 1995 and more than 400 papers had been written up to March 2016. Most of them are focused on the molecular pathways and experimentally-proved chemoresistance. In the present article, an update in the field of EMT in CRC based on the review of the literature and personal experience of the authors is presented. The information about the molecular and immunohistochemical(IHC) particularities of these processes and their possible role in the prognosis of CRC were also up-dated. This article focuses on the IHC quantification of the EMT, the immunoprofile of tumor buds and on the relation between EMT, angiogenesis, and stem cells activation. The EMT-induced chemoresistance vs chemotherapyor radiotherapy-induced EMT and cellular senescence was also synthesized for both conventional and targeted therapy. As a future perspective, the EMTangiogenesis-stemness link could be used as a possible valuable parameter for clinical follow-up and targeted therapeutic oncologic management of patients with CRC. Association of dexamethasone and angiotensin converting enzyme inhibitors combined with conventional chemotherapies could have clinical benefits in patients with CRC. The main conclusion is that, although many studies have been published, the EMT features are still incompletely elucidated and newly discovered EMT markers provide confusing data in understanding this complicated process, which might have significant clinical impact.

Paradoxical expression pattern of the epithelial mesenchymal transition-related biomarkers CD44, SLUG, N-cadherin and VSIG1/Glycoprotein A34 in gastrointestinal stromal tumors

AIM To evaluate the immunohistochemical(IHC) expression of five biomarkers, commonly involved in epithelial mesenchymal/mesenchymal epithelial transition(EMT/MET), in gastrointestinal stromal tumors(GISTs). METHODS In 80 consecutive GISTs the IHC examinations were performed using the EMT-related antibodies E-cadherin,N-cadherin, SLUG, V-set and immunoglobulin domain containing 1(VSIG1) and CD44. RESULTS The positivity rate was 88.75% for SLUG, 83.75% for VSIG1, 36.25% for CD44 and 10% for N-cadherin. No correlation was noted between the examined markers and clinicopathological parameters. Nuclear positivity for SLUG and VSIG1 was observed in all cases with distant metastasis. The extra-gastrointestinal stromal tumors(e-GISTs) expressed nuclear positivity for VSIG1 and SLUG, with infrequent positivity for N-cadherin and CD44. The low overall survival was mainly dependent on VSIG1 negativity(P = 0.01) and nuclear positivity for SLUG and/or CD44. CONCLUSION GIST aggressivity may be induced by nuclear upregulation of SLUG and loss or cytoplasm-to-nuclear translocation of VSIG1. SLUG and VSIG1 may act as activated nuclear transcription factors. The CD44, but not N-cadherin, might also have an independent prognostic value in these tumors. The role of the EMT/MET-related transcription factors in the evolution of GISTs, should be revisited with a larger dataset. This is the first study exploring the IHC pattern of VSIG1 in GISTs.

Interaction of arylsulfatases A and B with maspin: A possible explanation for dysregulation of tumor cell metabolism and invasive potential of colorectal cancer

BACKGROUND Although it has been shown that arylsulfatases are lost in colorectal cancer(CRC)cell lines,their exact role in the carcinogenesis and behavior of this cancer was not elucidated.No data about the correlation between serum and immunohistochemical(IHC)level of arylsulfatases(ARSA,ARSB)in patients with CRC were published yet.AIM To evaluate the possible prognostic value of ARSA and/or ARSB in CRC,at circulating and protein levels.METHODS The present study included 45 consecutive patients who were prospectively diagnosed with CRC.For IHC stains(protein expression)ARSA,ARSB and maspin expression were quantified.For these markers,cytoplasmic expression was taken into account.For gene expression study,circulating mRNA was isolated from all patients,before surgery.A group of 45 healthy patients without inflammatory or tumor pathologies was used as control group.Reverse transcription and Taqman Gene Expression Array were used for ARSB gene expression.RESULTS The preoperative circulating RNA level of the ARSB gene was significantly decreased in patients with CRC(RQ<1),compared with the control group(RQ>1).A more significant decrease(RQ<0.5)occurred in ulcero-infiltrative maspinpositive adenocarcinomas,with a higher degree of tumor budding,diagnosed in locally advanced stages(pT3/4).ARSA/maspin immunopositivity indicated a higher risk for lymph node metastasis,while triple positivity for maspin/ARSA/ARSB and ARSB gene expression level<0.5 were indicators of CRC aggressive behavior,independent of lymph node status.CONCLUSION The significant independent negative prognostic factors of CRC are the ulceroinfiltrative aspect,high budding degree,triple positivity for maspin,ARSA and ARSB,and low ARSB gene expression.

DNA extraction from paraffin embedded colorectal carcinoma samples: A comparison study of manual vs automated methods,using four commercially kits

BACKGROUND Nucleic acid isolation from formalin-fixed, paraffin-embedded tissue(FFPET)samples is a daily routine in molecular pathology laboratories, but extraction from FFPET is not always easily achieved. Choosing the right extraction technique is key for further examinations.AIM To compare the performance of four commercially available kits used for DNA extraction in routine practice.METHODS DNA isolation was performed on 46 randomly selected formalin-fixed, paraffinembedded(FFPE) colorectal adenocarcinoma(CRC) surgical specimens. Four commercially available extraction kits were used: two for manual DNA extraction(the Pure Link Genomic DNA Mini Kit from Invitrogen and the High Pure FFPE DNA Isolation Kit from Roche) and two for automated DNA extraction(the i Prep Genomic DNA Kit from Invitrogen and the Magna Pure LC DNA Isolation Kit from Roche). The DNA concentration and quality(odds ratio) among the four systems were compared. The results were correlated with the clinicopathological aspects of CRC cases: age, gender, localization, macro-and microscopic features,lymph node metastases, and the lymph node ratio.RESULTS The highest DNA concentration was obtained using the manual kits: 157.24 ±62.99 ng/μL for the Pure Link Genomic DNA Mini Kit and 86.64 ng/μL± 43.84 for the High Pure FFPE DNA Isolation Kit(P < 0.0001). Lower concentrations were obtained with automated systems: 20.39 ± 21.19 ng/μL for the Magna Pure LC DNA Isolation Kit and 8.722 ± 6.408 ng/μL for the i Prep Genomic DNA Kit,with differences between the systems used(P < 0.0001). The comparison between age, gender, tumor localization, pT or pN stage and the lymph node ratio indicated no statistically significant difference in DNA concentration using any of the nucleic acid isolation kits. DNA concentration was influenced by the macroscopic features and grade of differentiation. A higher DNA concentration was obtained for well-differentiated polypoid colorectal adenocarcinomas(CRCs), compared with undifferentiated ulcero-infiltrative carcinomas,irrespective of the kit used.CONCLUSION For research or diagnosis that needs high DNA concentrations, manual methods of DNA isolation should be used. A higher amount of DNA can be obtained from polypoid-type differentiated CRCs. Automated systems confer comfort and a lower amount of DNA that is, however, sufficient for classic polymerase chain reaction(PCR) and real-time quantitative PCR molecular examinations. All four commercially available kits can be successfully used in daily practice.