MALAT1 in Liquid Biopsy: The Diagnostic and Prognostic Promise for Colorectal Cancer and Adenomas?

Introduction: The development of colorectal cancer (CRC) is a multistep process accompanied by the accumulation of mutations that start from specific precancerous lesion - colorectal adenomas (CA). CRC incidence and mortality can be reduced by the early identification of these neoplasm. Colonoscopy is the most widely used screening method for CRC identification. Nowadays, clinical research interest is shifting to the use of liquid biopsy that may help with the early diagnosis of CA and CRC. In our previous study, we identified long non-coding RNA MALAT1 gene amplification associated with the development of CA. Methods: This study aimed to describe the potential of MALAT1 expression levels in the adenoma tissue of patients used in the previous study by real-time qPCR. Furthermore, we analysed the plasma samples of an independent group of patients with CA (n=97), CRC (n=101), and cancer-free individuals (CFI, n=48). Results: There was no difference in the MALAT1 expression level between CA patients with or without MALAT1 amplification. However, the plasma MALAT1 expression levels were significantly upregulated in patients with CRC and CA compared to CFI (for both p<0.001). Moreover, a correlation between MALAT1 expression and histological types of adenomas was identified- high-CRC-risk adenomas also displayed the highest MALAT1 expression levels. Furthermore, in CRC patients, MALAT1 levels were associated with a response to therapy. Conclusion: MALAT1 expression levels could serve as a promising circulating biomarker for early CA and CRC diagnosis, and even as a predictor of therapy response in CRC patients.

A Fecal MicroRNA Signature by Small RNA Sequencing Accurately Distinguishes Colorectal Cancers: Results From a Multicenter Study.

BACKGROUND & AIMS: Fecal tests currently used for colorectal cancer (CRC) screening show limited accuracy in detecting early tumors or precancerous lesions. In this respect, we comprehensively evaluated stool microRNA (miRNA) profiles as biomarkers for noninvasive CRC diagnosis. METHODS: A total of 1273 small RNA sequencing experiments were performed in multiple biospecimens. In a cross-sectional study, miRNA profiles were investigated in fecal samples from an Italian and a Czech cohort (155 CRCs, 87 adenomas, 96 other intestinal diseases, 141 colonoscopy-negative controls). A predictive miRNA signature for cancer detection was defined by a machine learning strategy and tested in additional fecal samples from 141 CRC patients and 80 healthy volunteers. miRNA profiles were compared with those of 132 tumors/adenomas paired with adjacent mucosa, 210 plasma extracellular vesicle samples, and 185 fecal immunochemical test leftover samples. RESULTS: Twenty-five miRNAs showed altered levels in the stool of CRC patients in both cohorts (adjusted P < .05). A 5-miRNA signature, including miR-149-3p, miR-607-5p, miR-1246, miR-4488, and miR-6777-5p, distinguished patients from control individuals (area under the curve [AUC], 0.86; 95% confidence interval [CI], 0.79-0.94) and was validated in an independent cohort (AUC, 0.96; 95% CI, 0.92-1.00). The signature classified control individuals from patients with low-/high-stage tumors and advanced adenomas (AUC, 0.82; 95% CI, 0.71-0.97). Tissue miRNA profiles mirrored those of stool samples, and fecal profiles of different gastrointestinal diseases highlighted miRNAs specifically dysregulated in CRC. miRNA profiles in fecal immunochemical test leftover samples showed good correlation with those of stool collected in preservative buffer, and their alterations could be detected in adenoma or CRC patients. CONCLUSIONS: Our comprehensive fecal miRNome analysis identified a signature accurately discriminating cancer aimed at improving noninvasive diagnosis and screening strategies.

MiR-140 leads to MRE11 downregulation and ameliorates oxaliplatin treatment and therapy response in colorectal cancer patients.

Cancer therapy failure is a fundamental challenge in cancer treatment. One of the most common reasons for therapy failure is the development of acquired resistance of cancer cells. DNA-damaging agents are frequently used in first-line chemotherapy regimens and DNA damage response, and DNA repair pathways are significantly involved in the mechanisms of chemoresistance. MRE11, a part of the MRN complex involved in double-strand break (DSB) repair, is connected to colorectal cancer (CRC) patients' prognosis. Our previous results showed that single-nucleotide polymorphisms (SNPs) in the 3' untranslated region (3'UTR) microRNA (miRNA) binding sites of MRE11 gene are associated with decreased cancer risk but with shorter survival of CRC patients, which implies the role of miRNA regulation in CRC. The therapy of colorectal cancer utilizes oxaliplatin (oxalato(trans-l-1,2-diaminocyclohexane)platinum), which is often compromised by chemoresistance development. There is, therefore, a crucial clinical need to understand the cellular processes associated with drug resistance and improve treatment responses by applying efficient combination therapies. The main aim of this study was to investigate the effect of miRNAs on the oxaliplatin therapy response of CRC patients. By the in silico analysis, miR-140 was predicted to target MRE11 and modulate CRC prognosis. The lower expression of miR-140 was associated with the metastatic phenotype (p < 0.05) and poor progression-free survival (odds ratio (OR) = 0.4, p < 0.05). In the in vitro analysis, we used miRNA mimics to increase the level of miR-140 in the CRC cell line. This resulted in decreased proliferation of CRC cells (p < 0.05). Increased levels of miR-140 also led to increased sensitivity of cancer cells to oxaliplatin (p < 0.05) and to the accumulation of DNA damage. Our results, both in vitro and in vivo, suggest that miR-140 may act as a tumor suppressor and plays an important role in DSB DNA repair and, consequently, CRC therapy response.

Monitoring of telomere dynamics in peripheral blood leukocytes in relation to colorectal cancer patients' outcomes.

We investigated the possible associations between leukocyte telomere length, therapy outcomes, and clinicopathological features in patients with colorectal cancer. Additionally, telomerase reverse transcriptase (TERT) expression was evaluated. Telomere length was measured using singleplex qPCR in 478 consecutive leukocyte DNA samples from 198 patients. Blood was drawn at diagnosis prior to any therapy and then at 6-month intervals for 18 months. Following diagnosis, the telomeres gradually shortened during the course of the treatment regardless of the patient's age. The most pronounced decrease was observed 12 months after the diagnosis (p < 0.0001). Based on tumor localization, the decrease in telomere length one year after the diagnosis followed different trajectories (p = 0.03). In patients treated with adjuvant therapy, telomere length correlated with the time elapsed after completion of therapy (p = 0.03). TERT expression did not correlate with the telomere length; however, it was higher in women than men (1.35-fold, 95% CI 1.11-1.65, p = 0.003) and in smokers than non-smokers (1.27-fold, 95% CI 1.01-1.61, p = 0.04). Leukocyte telomere length declines naturally during aging, but the accelerated shortening observed in our patients was age-independent. Telomere length manifestly reflected chemotherapy impact and could be linked to therapy toxicity.

The Associations of Selenoprotein Genetic Variants with the Risks of Colorectal Adenoma and Colorectal Cancer: Case-Control Studies in Irish and Czech Populations.

BACKGROUND: Selenium manifests its biological effects through its incorporation into selenoproteins, which play several roles in countering oxidative and inflammatory responses implicated in colorectal carcinogenesis. Selenoprotein genetic variants may contribute to colorectal cancer (CRC) development, as we previously observed for SNP variants in a large European prospective study and a Czech case-control cohort. METHODS: We tested if significantly associated selenoprotein gene SNPs from these studies were also associated with CRC risk in case-control studies from Ireland (colorectal neoplasia, i.e., cancer and adenoma cases: 450, controls: 461) and the Czech Republic (CRC cases: 718, controls: 646). Genotyping of 23 SNPs (20 in the Irish and 13 in the Czechs) was performed by competitive specific allele-specific PCR (KASPar). Multivariable adjusted logistic regression was used to assess the associations with CRC development. RESULTS: We found significant associations with an increased CRC risk for rs5859 (SELENOF) and rs2972994 (SELENOP) in the Irish cohort but only with rs4802034 (SELENOV) in the Czechs. Significant associations were observed for rs5859 (SELENOF), rs4659382 (SELENON), rs2972994 (SELENOP), rs34713741 (SELENOS), and the related Se metabolism gene variant rs2275129 (SEPHS1) with advanced colorectal neoplasia development. However, none of these findings retained significance after multiple testing corrections. CONCLUSIONS: Several SNPs previously associated with CRC risk were also associated with CRC or colorectal neoplasia development in either the Irish or Czech cohorts. Selenoprotein gene variation may modify CRC risk across diverse European populations, although the specific variants may differ.

Association of circulating short chain fatty acid levels with colorectal adenomas and colorectal cancer.

BACKGROUND & AIMS: Short chain fatty acid (SCFAs) are bacterially derived metabolites suggested to have protective roles against colorectal cancer (CRC) development. However, there is sparse evidence from epidemiological studies in this context. Here, we assessed whether circulating SCFA concentrations varied in patients with colorectal adenomas (CRA) and CRC. METHODS: Levels of seven SCFAs were extracted from plasma samples and determined by gas chromatography for 213 individuals from Ireland and the Czech Republic (CRC, n = 84; CRA, n = 66; controls, n = 63). RESULTS: In the Irish CRA/CRC cohort, only levels of 2-MethylButyric acid were significantly higher in cancers compared to the adenoma and control groups (p-values = 0.016 and 0.043). Using regression analysis, we observed that levels of Acetic and Propionic acid were associated with an increased CRC risk in the Czech cohort (Odd Ratio (OR): 1.02; 95% Confidence interval (CI): 1.00-1.03; OR: 1.29; 95% CI: 1.05-1.59, respectively), while i-Valeric and Valeric acid levels were associated with a decreased cancer risk (OR: 0.92; 95% CI: 0.86-0.99; OR: 0.67; 95% CI: 0.44-1.00). In the Irish cohort, levels of SCFAs were not associated with CRC risk. CONCLUSIONS: The association with colorectal neoplasia varied between the studied SCFAs. Future studies need to confirm these findings and address the mechanism of how these acids may promote or prevent colorectal carcinogenesis.

Analysis of MicroRNA Expression Changes During the Course of Therapy In Rectal Cancer Patients.

MicroRNAs (miRNAs) regulate gene expression in a tissue-specific manner. However, little is known about the miRNA expression changes induced by the therapy in rectal cancer (RC) patients. We evaluated miRNA expression levels before and after therapy and identified specific miRNA signatures reflecting disease course and treatment responses of RC patients. First, miRNA expression levels were assessed by next-generation sequencing in two plasma samplings (at the time of diagnosis and a year after) from 20 RC patients. MiR-122-5p and miR-142-5p were classified for subsequent validation in plasma and plasma extracellular vesicles (EVs) on an independent group of RC patients (n=107). Due to the intrinsic high differences in miRNA expression levels between samplings, cancer-free individuals (n=51) were included in the validation phase to determine the baseline expression levels of the selected miRNAs. Expression levels of these miRNAs were significantly different between RC patients and controls (for all p <0.001). A year after diagnosis, miRNA expression profiles were significantly modified in patients responding to treatment and were no longer different from those measured in cancer-free individuals. On the other hand, patients not responding to therapy maintained low expression levels in their second sampling (miR-122-5p: plasma: p=0.05, EVs: p=0.007; miR-142-5p: plasma: p=0.008). Besides, overexpression of miR-122-5p and miR-142-5p in RC cell lines inhibited cell growth and survival. This study provides novel evidence that circulating miR-122-5p and miR-142-5p have a high potential for RC screening and early detection as well as for the assessment of patients' outcomes and the effectiveness of treatment schedule.

Mutational landscape of plasma cell-free DNA identifies molecular features associated with therapeutic response in patients with colon cancer. A pilot study.

Cell-free DNA (cfDNA) has recently been used as a non-invasive diagnostic tool for detecting tumour-specific mutations. cfDNA may also be used for monitoring disease progression and treatment response, but so far researchers focused on one or few genes only. A genomic profile may provide better information on patient prognosis compared to single specific mutations. In this hypothesis-generating study, we profiled by whole exome sequencing serial plasma samples from 10 colon cancer (CC) patients collected before and after 5-fluorouracil-based therapy, and one year after diagnosis to determine alterations associated with treatment response. In parallel, genome profiling was also performed in patients' corresponding tumour tissue to ascertain the molecular landscape of resistant tumours. The mutation concordance between cfDNA and tumour tissue DNA was higher in more advanced tumour stages than in the early stages of the disease. In non-responders, a specific mutation profile was observed in tumour tissues (TPSD1 p.Ala92Thr, CPAMD8 p.Arg341Gln, OBP2A p.ArgTyr123CysHis). A pathogenic APC mutation (p.Ser1315Ter) was detected only in cfDNA of one poor responder one year after the diagnosis and after therapy termination. Another poor responder presented a likely pathogenic TP53 mutation (p.Arg110Pro) in cfDNA of all plasma samplings and in tumour tissue. In conclusion, cfDNA could be used for genetic characterisation of CC patients and might be clinically useful for non-invasive therapy response monitoring.

Local Immune Changes in Early Stages of Inflammation and Carcinogenesis Correlate with the Collagen Scaffold Changes of the Colon Mucosa.

Continuous activation of the immune system inside a tissue can lead to remodelling of the tissue structure and creation of a specific microenvironment, such as during the tumour development. Chronic inflammation is a central player in stimulating changes that alter the tissue stroma and can lead to fibrotic evolution. In the colon mucosa, regulatory mechanisms, including TGF-ß1, avoid damaging inflammation in front of the continuous challenge by the intestinal microbiome. Inducing either DSS colitis or AOM colorectal carcinogenesis in AVN-Wistar rats, we evaluated at one month after the end of each treatment whether immunological changes and remodelling of the collagen scaffold were already in development. At this time point, we found in both models a general downregulation of pro-inflammatory cytokines and even of TGF-ß1, but not of IL-6. Moreover, we demonstrated by multi-photon microscopy the simultaneously presence of pro-fibrotic remodelling of the collagen scaffold, with measurable changes in comparison to the control mucosa. The scaffold was significantly modified depending on the type of induced stimulation. These results suggest that at one month after the end of the DSS or AOM inductions, a smouldering inflammation is present in both induced conditions, since the pro-inflammatory cytokines still exceed, in proportion, the local homeostatic regulation of which TGF-ß1 is a part (inflammatory threshold). Such an inflammation appears sufficient to sustain remodelling of the collagen scaffold that may be taken as a possible pathological marker for revealing pre-neoplastic inflammation.

Expression quantitative trait loci in ABC transporters are associated with survival in 5-FU treated colorectal cancer patients.

The chemotherapeutic efficacy in colorectal cancer (CRC) is limited due to the inter-individual variability in drug response and the development of tumour resistance. ATP-binding cassette (ABC) transporters are crucial in the development of resistance by the efflux of anticancer agents from cancer cells. In this study, we identified 14 single nucleotide polymorphisms (SNPs) in 11 ABC transporter genes acting as an expression of quantitative trait loci (eQTLs), i.e. whose variation influence the expression of many downstream genes. These SNPs were genotyped in a case-control study comprising 1098 cases and 1442 healthy controls and analysed in relation to CRC development risk and patient survival. Considering a strict correction for multiple tests, we did not observe any significant association between SNPs and CRC risk. The rs3819720 polymorphism in the ABCB3/TAP2 gene was statistically significantly associated with shorter overall survival (OS) in the codominant, and dominant models [GA vs. GG, hazard ratio (HR) = 1.48; P = 0.002; AA vs. GG, HR = 1.70; P = 0.004 and GA + AA vs. GG, HR = 1.52; P = 0.0006]. Additionally, GA carriers of the same SNP displayed worse OS after receiving 5-FU based chemotherapy. The variant allele of rs3819720 polymorphism statistically significantly affected the expression of 36 downstream genes. Screening for eQTL polymorphisms in relevant genes such as ABC transporters that can regulate the expression of several other genes may help to identify the genetic background involved in the individual response to the treatment of CRC patients.

Epistatic effect of TLR3 and cGAS-STING-IKKε-TBK1-IFN signaling variants on colorectal cancer risk.

OBJECTIVE: The TLR3/cGAS-STING-IFN signaling has recently been reported to be disturbed in colorectal cancer due to deregulated expression of the genes involved. Our study aimed to investigate the influence of potential regulatory variants in these genes on the risk of sporadic colorectal cancer (CRC) in a Czech cohort of 1424 CRC patients and 1114 healthy controls. METHODS: The variants in the TLR3, CGAS, TMEM173, IKBKE, and TBK1 genes were selected using various online bioinformatic tools, such as UCSC browser, HaploReg, Regulome DB, Gtex Portal, SIFT, PolyPhen2, and miRNA prediction tools. RESULTS: Logistic regression analysis adjusted for age and sex detected a nominal association between CRC risk and three variants, CGAS rs72960018 (OR: 1.68, 95% CI: 1.11-2.53, P-value = .01), CGAS rs9352000 (OR: 2.02, 95% CI: 1.07-3.84, P-value = .03) and TMEM173 rs13153461 (OR: 1.53, 95% CI: 1.03-2.27, P-value = .03). Their cumulative effect revealed a threefold increased CRC risk in carriers of 5-6 risk alleles compared to those with 0-2 risk alleles. Epistatic interactions between these genes and the previously genotyped IFNAR1, IFNAR2, IFNA, IFNB, IFNK, IFNW, IRF3, and IRF7 genes, were computed to test their effect on CRC risk. Overall, we obtained nine pair-wise interactions within and between the CGAS, TMEM173, IKBKE, and TBK1 genes. Two of them remained statistically significant after Bonferroni correction. Additional 52 interactions were observed when IFN variants were added to the analysis. CONCLUSIONS: Our data suggest that epistatic interactions and a high number of risk alleles may play an important role in CRC carcinogenesis, offering novel biological understanding for the CRC management.

Malignant Peritoneal Tumors - Introduction.

This article provides an introduction to peritoneal tumors, which are the subject of a series of review papers published in Issue 5 (2019) of Klinicka onkologie. Many malignant peritoneal tumors are characterized by production of mucinous and gelatinous masses, multiple peritoneal disability, so-called peritoneal carcinomatosis, and various grades of malignancy depending on their origin, staging, and histological type. Malignant peritoneal tumors are rare and their clinical symptomatology is nonspecific and varies according to the extent of disability. Diagnosis, particularly in the initial asymptomatic stages, is very complicated and often impossible, and tumors are often diagnosed by chance during other operations. Malignant peritoneal tumors were regarded as incurable and lethal for a long time; however, this view has changed over the past three decades. The Sugarbaker method, a combination of cytoreductive surgery and hyperthermic intraperitoneal chemotherapy, was introduced in the 1990s. Postoperative cytostatic lavage is usually performed in specific cases. Classifications for the extent of disease and completeness of cytoreduction were established. Studies repeatedly confirmed the efficacy of this treatment for peritoneal malignancy. The combination of an aggressive surgical approach and intraperitoneal chemotherapy not only enhances quality of life, but also prolongs progression-free survival and overall survival in selected patients. Specialized centers for treatment of peritoneal malignancy were established based on results from the Czech Republic and around the world. These centers provide complex care, including specific surgical interventions and follow-up, for selected patients with primary and secondary peritoneal malignancy.

Treatment of Malignant Peritoneal Mesothelioma.

Malignant mesothelioma is a highly malignant disease that most often occurs in the pleura of the thoracic cavity, followed by the peritoneum, pericardium, or tinea vaginalis testis. Malignant peritoneal mesothelioma (MPM) accounts for 10-15% of all mesotheliomas. The most significant risk factor for MPM is exposure to asbestos. There is no specific symptomatology, and imaging (computed tomography) and histopathology are crucial for diagnosis. There are no generally accepted guidelines for radical treatment of MPM. Previously, the prognosis of MPM patients was poor, with survival of up to 1 year. However, median survival of patients who are suitable candidates for radical therapy is currently 3-5 years. A combination of cytoreductive surgery (CRS) and hyperthermic perioperative chemotherapy (HIPEC) is recommended in selected patients, while chemotherapy alone has insufficient efficacy. Systemic chemotherapy remains the only treatment option for patients who are unsuitable for CRS and HIPEC. In selected patients scheduled for or currently undergoing CRS and HIPEC, surgery may be performed in combination with systemic chemotherapy in the neoadjuvant or adjuvant setting; however, the benefit is unclear. There are no recommendations for follow-up of MPM patients after radical surgery. Existing guidelines for the pleural form (e.g., those issued by the European Society for Medical Oncology) do not specify the frequency or method of investigation. In the absence of specific serum markers, only CA 125 and mesothelin are generally available. Imaging methods include ultrasonography, computed tomography, and magnetic resonance imaging.

Fusobacterium nucleatum tumor DNA levels are associated with survival in colorectal cancer patients.

There is increasing evidence indicating a role for Fusobacterium nucleatum (F. nucleatum) in colorectal cancer (CRC) development and prognosis. This study evaluated F. nucleatum as a prognostic biomarker, by assessing its association with post-diagnosis survival from CRC. From September 2008 to April 2012 CRC patients (n = 190) were recruited from three hospitals within the Czech Republic. F. nucleatum DNA copies were measured in adjacent non-malignant and colorectal tumor tissues using quantitative real-time PCR. Cox Proportional Hazards (HR) models were applied to evaluate the association between F. nucleatum DNA and overall survival, adjusting for key confounders. Risk prediction modeling was conducted to evaluate the ability to predict survival based on F. nucleatum status. High, compared with low, levels of F. nucleatum in colorectal tumor tissues were associated with poorer overall survival (adjusted HR 1.68, 95% CI 1.02-2.77), which was slightly attenuated after additional adjustment for microsatellite instability status. However, inclusion of F. nucleatum in risk prediction models did not improve the ability to identify patients who died beyond known prognostic factors such as disease pathology staging. Although the increased presence of F. nucleatum was associated with poorer prognosis in CRC patients, this may have limited clinical relevance as a prognostic biomarker.

Correction: Single nucleotide polymorphisms within MUC4 are associated with colorectal cancer survival.

[This corrects the article DOI: 10.1371/journal.pone.0216666.].

Single nucleotide polymorphisms within MUC4 are associated with colorectal cancer survival.

Mucins and their glycosylation have been suggested to play an important role in colorectal carcinogenesis. We examined potentially functional genetic variants in the mucin genes or genes involved in their glycosylation with respect to colorectal cancer (CRC) risk and clinical outcome. We genotyped 23 single nucleotide polymorphisms (SNPs) covering 123 SNPs through pairwise linkage disequilibrium (r2>0.80) in the MUC1, MUC2, MUC4, MUC5AC, MUC6, and B3GNT6 genes in a hospital-based case-control study of 1532 CRC cases and 1108 healthy controls from the Czech Republic. We also analyzed these SNPs in relation to overall survival and event-free survival in a subgroup of 672 patients. Among patients without distant metastasis at the time of diagnosis, two MUC4 SNPs, rs3107764 and rs842225, showed association with overall survival (HR 1.40, 95%CI 1.08-1.82, additive model, log-rank p = 0.004 and HR 0.64, 95%CI 0.42-0.99, recessive model, log-rank p = 0.01, respectively) and event-free survival (HR 1.31, 95%CI 1.03-1.68, log-rank p = 0.004 and HR 0.64, 95%CI 0.42-0.96, log-rank p = 0.006, respectively) after adjustment for age, sex and TNM stage. Our data suggest that genetic variation especially in the transmembrane mucin gene MUC4 may play a role in the survival of CRC and further studies are warranted.

Significance of postoperative follow-up of patients with metastatic colorectal cancer using circulating tumor DNA.

BACKGROUND: One of the most notable applications for circulating tumor DNA (ctDNA) detection in peripheral blood of patients with metastatic colorectal cancer (mCRC) is a long-term postoperative follow-up. Sometimes referred to as a "liquid (re)biopsy" it is a minimally invasive procedure and can be performed repeatedly at relatively short intervals (months or even weeks). The presence of the disease and the actual extent of the tumor burden (tumor mass) within the patient's body can be monitored. This is of particular importance, especially when evaluating radicality of surgical treatment as well as for early detection of disease progression or recurrence. AIM: To confirm the radicality of surgery using ctDNA and compare available methods for detection of recurrence in metastatic colorectal cancer. METHODS: A total of 47 patients with detected ctDNA and indications for resection of mCRC were enrolled in the multicenter study involving three surgical centers. Standard postoperative follow-ups using imaging techniques and the determination of tumor markers were supplemented by ctDNA sampling. In addition to the baseline ctDNA testing prior to surgery, a postoperative observation was conducted by evaluating ctDNA presence up to a week after surgery and subsequently at approximately three-month intervals. The presence of ctDNA was correlated with radicality of surgical treatment and the actual clinical status of the patient. RESULTS: Among the monitored patients, the R0 (curative) resection correlated with postoperative ctDNA negativity in 26 out of 28 cases of surgical procedures (26/28, 93%). In the remaining cases of R0 surgeries that displayed ctDNA, both patients were diagnosed with a recurrence of the disease after 6 months. In 7 patients who underwent an R1 resection, 4 ctDNA positivities (4/7, 57%) were detected after surgery and associated with the confirmation of early disease recurrence (after 3 to 7 months). All 15 patients (15/15, 100%) undergoing R2 resection remained constantly ctDNA positive during the entire follow-up period. In 22 cases of recurrence, ctDNA positivity was detected 22 times (22/22, 100%) compared to 16 positives (16/22, 73%) by imaging methods and 15 cases (15/22, 68%) of elevated tumor markers. CONCLUSION: ctDNA detection in patients with mCRC is a viable tool for early detection of disease recurrence as well as for confirmation of the radicality of surgical treatment.

Functional Polymorphisms in DNA Repair Genes Are Associated with Sporadic Colorectal Cancer Susceptibility and Clinical Outcome.

DNA repair processes are involved in both the onset and treatment efficacy of colorectal cancer (CRC). A change of a single nucleotide causing an amino acid substitution in the corresponding protein may alter the efficiency of DNA repair, thus modifying the CRC susceptibility and clinical outcome. We performed a candidate gene approach in order to analyze the association of non-synonymous single nucleotide polymorphisms (nsSNPs) in the genes covering the main DNA repair pathways with CRC risk and clinical outcome modifications. Our candidate polymorphisms were selected according to the foremost genomic and functional prediction databases. Sixteen nsSNPs in 12 DNA repair genes were evaluated in cohorts from the Czech Republic and Austria. Apart from the tumor-node-metastasis (TNM) stage, which occurred as the main prognostic factor in all of the performed analyses, we observed several significant associations of different nsSNPs with survival and clinical outcomes in both cohorts. However, only some of the genes (REV3L, POLQ, and NEIL3) were prominently defined as prediction factors in the classification and regression tree analysis; therefore, the study suggests their association for patient survival. In summary, we provide observational and bioinformatics evidence that even subtle alterations in specific proteins of the DNA repair pathways may contribute to CRC susceptibility and clinical outcome.

Expression profile of miR-17/92 cluster is predictive of treatment response in rectal cancer.

MicroRNA (miRNA) profiling represents a promising source of cancer-related biomarkers. miRNA signatures are specific for each cancer type and subgroups of patients with diverse treatment sensitivity. Yet this miRNA potential has not been satisfactorily explored in rectal cancer (RC). The aim of the study was to identify the specific miRNA signature with clinical and therapeutic relevance for RC. Expressions of 2555 miRNA were examined in 20 pairs of rectal tumors and matched non-malignant tissues by 3D-Gene Toray microarray. Candidate miRNAs were validated in an independent cohort of 100 paired rectal tissues and in whole plasma and exosomes of 100 RC patients. To study the association of miRNA profile with therapeutic outcomes, plasma samples were taken repeatedly over a time period of 1 year reflecting thus patients' treatment responses. Finally, the most prominent miRNAs were investigated in vitro for their involvement in cell growth. We identified RC-specific miRNA signature that distinguishes responders from non-responders to adjuvant chemotherapy. A predominant part of identified miRNAs was represented by the members of miR-17/92 cluster. Upregulation of miRNA-17, -18a, -18b, -19a, -19b, -20a, -20b and -106a in tumor was associated with higher risk of tumor relapse and their overexpression in RC cell lines stimulated cellular proliferation. Examination of these miRNAs in plasma exosomes showed that their levels differed between RC patients and healthy controls and correlated with patient's treatment response. miRNAs from miR-17/92 cluster represent a non-invasive biomarker to predict posttreatment prognosis in RC patients.

Base excision repair capacity as a determinant of prognosis and therapy response in colon cancer patients.

The DNA-damaging agent 5-fluorouracil represents the most commonly used chemotherapeutic drug for colorectal cancer patients. DNA lesions associated with 5-fluorouracil therapy are primarily repaired by base excision repair (BER) and mismatch repair (MMR) pathways. Published evidence suggests that the individual DNA repair capacity (DRC) may affect a patient's prognosis and response to chemotherapy. With this in mind, we designed a prospective study of which the main aim was to investigate BER-DRC in relation to 5-fluorouracil response as potential predictive and/or prognostic biomarker. BER-DRC was supplemented by a microsatellite instability (MSI) analysis which represents an indirect marker of MMR activity in the tumor. All parameters were measured in paired samples of tumor tissue and non-malignant adjacent mucosa of 123 incident colon cancer patients. Our results indicate that BER-DRC in non-malignant adjacent mucosa was positively associated with overall survival (P = 0.007) and relapse-free survival (P = 0.04). Additionally, in multivariate analysis, good therapy responders in TNM stage II and III with an elevated BER-DRC in mucosa exhibited better overall survival. Moreover, the overall survival of these patients was even better in the presence of a decreased BER-DRC in tumor tissue. The ratio of BER-DRC in tumor tissue over BER-DRC in mucosa positively correlated with advanced tumor stage (P = 0.003). With respect to MSI, we observed that MSI-high tumors were mostly localized in proximal colon; however, in our cohort, the MSI status affected neither patients' prognosis nor survival. In summary, the results of the present study suggest that the level of BER-DRC is associated with patients' survival. BER-DRC represents a potential prognostic biomarker, applicable for prediction of therapy response and useful for individual approach to patients.