Maria Teresa Rodia

A module of inflammatory cytokines defines resistance of colorectal cancer to EGFR inhibitors

Epidermal Growth Factor Receptor (EGFR) activates a robust signalling network to which colon cancer tumours often become addicted. Cetuximab, one of the monoclonal antibodies targeting this pathway, is employed to treat patients with colorectal cancer. However, many patients are intrinsically refractory to this treatment, and those who respond develop secondary resistance along time. Mechanisms of cancer cell resistance include either acquisition of new mutations or non genomic activation of alternative signalling routes. In this study, we employed a colon cancer model to assess potential mechanisms driving resistance to cetuximab. Resistant cells displayed increased ability to grow in suspension as colonspheres and this phenotype was associated with poorly organized structures. Factors secreted from resistant cells were causally involved in sustaining resistance, indeed administration to parental cells of conditioned medium collected from resistant cells was sufficient to reduce cetuximab efficacy. Among secreted factors, we report herein that a signature of inflammatory cytokines, including IL1A, IL1B and IL8, which are produced following EGFR pathway activation, was associated with the acquisition of an unresponsive phenotype to cetuximab in vitro. This signature correlated with lack of response to EGFR targeting also in patient-derived tumour xenografts. Collectively, these results highlight the contribution of inflammatory cytokines to reduced sensitivity to EGFR blockade and suggest that inhibition of this panel of cytokines in combination with cetuximab might yield an effective treatment strategy for CRC patients refractory to anti-EGFR targeting.

Systematic large-scale meta-analysis identifies a panel of two mRNAs as blood biomarkers for colorectal cancer detection

Colorectal cancer (CRC) is the third most common cancer in the world. A significant survival rate is achieved if it is detected at an early stage. A whole blood screening test, without any attempt to isolate blood fractions, could be an important tool to improve early detection of colorectal cancer. We searched for candidate markers with a novel approach based on the Transcriptome Mapper (TRAM), aimed at identifying specific RNAs with the highest differential expression ratio between colorectal cancer tissue and normal blood samples. This tool permits a large-scale systematic meta-analysis of all available data obtained by microarray experiments. The targeting of RNA took into consideration that tumour phenotypic variation is associated with changes in the mRNA levels of genes regulating or affecting this variation. A real time quantitative reverse transcription polymerase chain reaction (qRT- PCR) was applied to the validation of candidate markers in the blood of 67 patients and 67 healthy controls. The expression of genes: TSPAN8, LGALS4, COL1A2 and CEACAM6 resulted as being statistically different. In particular ROC curves attested for TSPAN8 an AUC of 0.751 with a sensitivity of 83.6% and a specificity of 58.2% at a cut off of 10.85, while the panel of the two best genes showed an AUC of 0.861 and a sensitivity of 92.5% with a specificity of 67.2%. Our preliminary study on a total of 134 subjects showed promising results for a blood screening test to be validated in a larger cohort with the staging stratification and in patients with other gastrointestinal diseases.

Colorectal cancer susceptibility: apparent gender-related modulation by ABCB1 gene polymorphisms.

BackgroundThe ATP-binding cassette transporter B1 (ABCB1) gene codes for a membrane efflux pump localized in epithelial cells. Together with other Permeability-glycoproteins in the small and large intestine, its product represents a barrier against xenobiotics, bacterial toxins, drugs and other substances introduced with diet, including carcinogens. The aim of this investigation was to verify the possible contribution of ABCB1 single nucleotide polymorphisms (SNPs) to the genetic risk of colorectal cancer (CRC).ResultsDNA obtained from the peripheral blood of 98 CRC patients and 100 healthy controls was genotyped for the three selected SNPs: 1236C > T (rs1128503), 2677G > T/A (rs2032582), and 3435C > T (rs1045642). Molecular data were analyzed to asses allele and haplotype association with CRC.No evidence of an association between ABCB1 alleles and CRC occurrence as a whole was found. However, ABCB1 showed either association with carcinoma of the sigmoid colon, and appeared able to influence the sex ratio among CRC patients. These two effects seemed to act independently based on multivariate analysis. We showed that ABCB1 polymorphisms were able to influence CRC susceptibility related to tumor localization and patient gender.ConclusionsWe suggest that sensitivity to undetermined risk factors could depend on the genetic background of ABCB1 locus, with a mechanism that also depends on patient gender.

Possible Gender-Related Modulation by the ROCK1 Gene in Colorectal Cancer Susceptibility

Aim: In view of accumulating evidence supporting a pivotal role of the Rho/ROCK pathway in cancer, we investigated Rho-kinase polymorphisms as potential susceptibility factors in colorectal cancer (CRC) in a representative sample of the Italian population. Methods: DNA obtained from the peripheral blood samples of 137 CRC patients and 141 healthy controls was genotyped for four ROCK1 (rs35996865; rs73963110; rs2127958; rs288980) and five ROCK2 (rs12692437; rs7563468; rs35768389; rs17463896; rs16857265) selected single nucleotide polymorphisms. Results: None of the allelic variants of the nine selected markers was associated with the occurrence of CRC or with the development of regional lymph node metastasis. By contrast, the ROCK1 rs35996865 G variant allele was significantly more frequent in male patients (p = 0.028) than in the control group. Conclusion: This finding is, at present, the first that points to a possible gender-related modulation by the ROCK1 gene in CRC susceptibility.

LGALS4, CEACAM6, TSPAN8, and COL1A2: Blood Markers for Colorectal Cancer—Validation in a Cohort of Subjects With Positive Fecal Immunochemical Test Result

Background A noninvasive blood test for the early detection of colorectal cancer (CRC) is highly required. We evaluated a panel of 4 mRNAs as putative markers of CRC. Materials and Methods We tested LGALS4, CEACAM6, TSPAN8, and COL1A2, referred to as the CELTiC panel, using quantitative reverse transcription polymerase chain reaction, on subjects with positive fecal immunochemical test (FIT) results and undergoing colonoscopy. Using a nonparametric test and multinomial logistic model, FIT‐positive subjects were compared with CRC patients and healthy individuals. Results All the genes of the CELTiC panel displayed statistically significant differences between the healthy subjects (n = 67), both low‐risk (n = 36) and high‐risk/CRC (n = 92) subjects, and those in the negative‐colonoscopy, FIT‐positive group (n = 36). The multinomial logistic model revealed LGALS4 was the most powerful marker discriminating the 4 groups. When assessing the diagnostic values by analysis of the areas under the receiver operating characteristic curves (AUCs), the CELTiC panel reached an AUC of 0.91 (sensitivity, 79%; specificity, 94%) comparing normal subjects to low‐risk subjects, and 0.88 (sensitivity, 75%; specificity, 87%) comparing normal and high‐risk/CRC subjects. The comparison between the normal subjects and the negative‐colonoscopy, FIT‐positive group revealed an AUC of 0.93 (sensitivity, 82%; specificity, 97%). Conclusion The CELTiC panel could represent a useful tool for discriminating subjects with positive FIT findings and for the early detection of precancerous adenomatous lesions and CRC. Micro‐Abstract To implement the effectiveness of early detection for colorectal cancer, we searched for mRNA blood markers (CELTiC panel) in a total of 231 blood samples from healthy subjects, colorectal cancer patients (CRC), and subjects with negative colonoscopy but positive fecal immunochemical test (FIT) results. Our data suggest that the CELTiC panel might represent a useful tool for discriminating subjects with positive FIT results and for early detection of precancerous adenomatous lesions and CRC.

Contents Vol. 82, 2015

Founded 1938 as ‘Schweizerische Zeitschrift für allgemeine Pathologie und Bakteriologie’ by A. v. Albertini, A. Grumbach and H. Mooser, continued as ‘Pathologia et Microbiologia’ (1960–1975) and ‘Experimental Cell Biology’ (1976–1989); incorporating ‘Pathology and Immunopathology Research’, founded 1982 as ‘Survey and Synthesis of Pathology Research’ by J.M. Cruse and R.E. Lewis, continued as ‘Pathobiology’, edited by J.M. Cruse and R.E. Lewis (1990–1998) Continued by Ch. Wittekind (1999–2004)

EGFR positive feedback loops and βeta Catenin driven miR-17-92 cluster converge to regulate EMT and drug resistance

Epidermal growth factor receptor (EGFR)-targeted cancer drug represents a mile- stone in oncology. Nevertheless the responses are invariably limited by the emer- gence of secondary drug-resistance (Misale, Di Nicolantonio et al. 2014). We found that drug-treated ‘‘EGFR-addicted’’ cancer cells engage a positive feedback loop lead- ing to NF-KB/βCatenin axis activation (Lauriola, Enuka et al. 2014), consequently promoting cell survival and limiting overall drug response. Specifically, secondary activation of βCatenin drives the production of an oncogenic cluster of microRNAs 17-92 (Lauriola, Donghwa et al. 2015) implicated in EMT transformation and resist- ance in colon clones. Hence βCatenin and EGFR combination pharmacological inhi- bition overcome the colon spheres growth and enhance tumor regression. These findings suggest that inhibition of EGFR feedback loop along with NF-kB/βCatenin axis may increase the response to a broad spectrum of drugs that target pathways of oncogene addiction.