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The transient build-up of DNA supercoiling during the translocation of replication forks threatens genome stability and is controlled by DNA topoisomerases (TOPs). This crucial process has been exploited with TOP poisons for cancer chemotherapy. However, pinpointing cellular determinants of the best clinical response to TOP poisons still remains enigmatic. Here, we present an integrated approach and demonstrate that endogenous and exogenous expression of the oncofetal high-mobility group AT-hook 2 (HMGA2) protein exhibited broad protection against the formation of hydroxyurea-induced DNA breaks in various cancer cells, thus corroborating our previously proposed model in which HMGA2 functions as a replication fork chaperone that forms a protective DNA scaffold at or close to stalled replication forks. We now further demonstrate that high levels of HMGA2 also protected cancer cells against DNA breaks triggered by the clinically important TOP1 poison irinotecan. This protection is most likely due to the recently identified DNA supercoil constraining function of HMGA2 in combination with exclusion of TOP1 from binding to supercoiled substrate DNA. In contrast, low to moderate HMGA2 protein levels surprisingly potentiated the formation of irinotecan-induced genotoxic covalent TOP1-DNA cleavage complexes. Our data from cell-based and several in vitro assays indicate that, mechanistically, this potentiating role involves enhanced drug-target interactions mediated by HMGA2 in ternary complexes with supercoiled DNA. Subtelomeric regions were found to be extraordinarily vulnerable to these genotoxic challenges induced by TOP1 poisoning, pointing at strong DNA topological barriers located at human telomeres. These findings were corroborated by an increased irinotecan sensitivity of patient-derived xenografts of colorectal cancers exhibiting low to moderate HMGA2 levels. Collectively, we uncovered a therapeutically important control mechanism of transient changes in chromosomal DNA topology that ultimately leads to enhanced human subtelomere stability.

KRAS is a gatekeeper gene in human colorectal tumorigenesis. KRAS is 'undruggable'; hence, efforts have been diverted to inhibit downstream RAF/MEK/ERK and PI3K/Akt signaling. Nevertheless, none of these inhibitors has progressed to clinical use despite extensive trials. We examined levels of phospho‐ERK1/2(T202/Y204) and phospho‐Akt1/2/3(S473) in human colorectal tumor compared to matched mucosa with semi‐quantitative near‐infrared western blot and confocal fluorescence immunohistochemistry imaging. Surprisingly, 75.5% (25/33) of tumors had lower or equivalent phospho‐ERK1/2 and 96.9% (31/32) of tumors had lower phospho‐Akt1/2/3 compared to matched mucosa, irrespective of KRAS mutation status. In contrast, we discovered KRAS‐dependent SOX9 upregulation in 28 of the 31 (90.3%) tumors. These observations were substantiated by analysis of the public domain transcriptomics The Cancer Genome Atlas (TCGA) and NCBI Gene Expression Omnibus (GEO) datasets and proteomics Clinical Proteomic Tumor Analysis Consortium (CPTAC) dataset. These data suggest that RAF/MEK/ERK and PI3K/Akt signaling are unlikely to be activated in most human colorectal cancer. [ABSTRACT FROM AUTHOR]

Introduction: Successful stenting of an obstructing colorectal tumor can avert upfront emergency surgery in malnourished obstructed patients with metastatic disease and poor physiological condition. This study aims to evaluate the outcomes of stenting followed by primary colorectal tumor resection among patients with obstructed stage IV colorectal cancer at presentation, over a 10‐year period. Methods: From 2007 to 2016, a cohort comprising 25 consecutive patients were retrospectively reviewed from a prospectively collected database. The durability of palliation of bowel obstruction, oncological outcomes and factors influencing overall survival were analyzed. Results: No re‐interventions were required for bowel obstruction during the study period. The overall perioperative morbidity rate was 16%, with no postoperative 90‐day mortality. Laparoscopic resection rate was 52% and stoma formation rate was 8%. The median overall survival was 24 months for the entire cohort, and the 1‐, 3‐ and 5‐year survival rates were 80%, 35% and 23.33% respectively. More than one site of distant metastases, peritoneal involvement, and elevated carcinoembryonic antigen levels were significantly associated with poorer survival outcomes. Patients with peritoneal‐only metastasis had worse outcomes, with a median survival of 7 months and no patients surviving beyond 18 months. Conclusion: Stenting followed by resection of the primary obstructing colorectal cancer provides durable palliation among patients with stage IV disease, with low perioperative morbidity and stoma formation rates. Superior survival was observed among patients with single‐site, non‐peritoneal distant metastases. [ABSTRACT FROM AUTHOR]

Background: Laparoscopic low anterior resection for rectal cancer has superior short‐term benefits compared to open surgery. When operative conditions do not favour a totally‐laparoscopic (TL) approach, a hybrid operation can be performed. In this laparoscopic‐assisted (LA) approach, mobilization and vessel ligation are performed laparoscopically, with total mesorectal excision and distal transection performed either partially or totally in an extra‐corporeal fashion. We compared short‐term post‐operative and oncological outcomes of both approaches. Methods: A prospectively collected database of patients who underwent laparoscopic low anterior resection for rectal cancer between January 2009 and December 2014 was retrospectively analysed. Demographics, post‐operative and oncological outcomes were compared. Results: Of 174 patients, 97 were completed by TL, 62 by LA and the remaining 15 were converted to open. Baseline demographics were similar. LA group compared to TL group had bulkier rectal cancers (6.75 cm3 versus 4.50 cm3, P = 0.04) which were lower (6 cm versus 7 cm from anal verge, P = 0.02). They were of a more advanced tumour grade and had greater incidence of lymphovascular invasion. Yet, post‐operative outcomes such as time to diet, pain scores, hospitalization duration, wound‐related and anastomotic complications, 30‐day morbidity and mortality were similar. There was no difference in oncological adequacy, including circumferential resection margins, distal margins, lymph node harvest and 2‐year local recurrence rates. Conclusion: Laparoscopic‐assisted low anterior resection enables minimally invasive rectal surgery to be performed despite unfavourable tumour factors and technical challenges; and compares favourably with TL approach in terms of short‐term outcomes and oncological safety. [ABSTRACT FROM AUTHOR]

Familial adenomatous polyposis (FAP) is an autosomal-dominantly inherited form of colorectal cancer (CRC) caused by mutation in the adenomatous polyposis coli (APC) gene. Our ability to exhaustively screen for APC mutations identify microsatellite-stable and APC-mutation negative familial CRC patients, enabling us to search for novel genes. We performed genome-wide scan on two affected siblings of one family and 88 ethnicity- and gender-matched healthy controls to identify deletions shared by the siblings. Combined loss of heterozygosity, copy number and allelic-specific copy number analysis uncovered 5 shared deletions. Long-range polymerase chain reaction (PCR) confirmed chromosome 19q13 deletion, which was subsequently found in one other family. The 32 kb deleted region harbors the CYP2A7 gene and was enriched with enhancer, repressor and insulator sites. The wildtype allele was lost in the polyps of the proband. Further, real-time RT-PCR assays showed that expressions of MIA and MIA-RAB4B located 35 kb upstream of the deletion, were up-regulated in the polyps compared to the matched mucosa of the proband. MIA-RAB4B, the read-through long non-coding RNA (lncRNA), RAB4B, PIM2 and TAOK1 share common binding site of a microRNA, miR-24, in their 3’UTRs. PIM2 and TAOK1, two target oncogenes of miR-24, were co-ordinately up-regulated with MIA-RAB4B in the polyps, suggesting that MIA-RAB4B could function as competitive endogenous RNA to titrate miR-24 away from its other targets. The data suggest that the 19.13 deletion disrupted chromatin boundary, leading to altered expression of several genes and lncRNA, could contribute to colorectal cancer via novel genetic and epigenetic mechanisms. [ABSTRACT FROM AUTHOR]

Germline mutation in the adenomatous polyposis coli (APC) gene causes the majority (80%) of familial adenomatous polyposis (FAP), an autosomal dominantly inherited form of colorectal cancer (CRC). Mutation in 5′end of exon 9 of APC usually results in an attenuated form of FAP (aFAP), characterized by later age of onset and fewer polyps. The presence of exon 9a, an in-frame isoform with exon 8 spliced to 3′end of exon 9, modulates any deleterious effect of the mutation. A third lowly expressed isoform that completely skips exon 9 is present in both healthy individuals and FAP patients. We report here an interesting case of a proband with an APC mutation in 5′end of exon 9 that presented with six synchronous advanced CRCs at age 37. The novel insertion-deletion (indel) at codon 409, c.1226-1229delTTTTinsAAA, caused upregulation of the 'skip exon 9' isoform, r934-1312del, resulting in a premature stop codon at exon 10 and a truncated protein that removed all of the β-catenin (CTNNB1) binding motifs, thus activating the downstream T-cell transcription factor (Tcf) pathway. Exon 9a isoform was concomitantly downregulated. This finding emphasizes the necessity of examining the various isoforms of exon 9 to avoid clinical mismanagement and counseling based on just the mutation site by genomic DNA sequencing alone. [ABSTRACT FROM AUTHOR]

Background: Germline defects of mismatch repair (MMR) genes underlie Lynch Syndrome (LS). We aimed to gain comprehensive genetic and epigenetic profiles of LS families in Singapore, which will facilitate efficient molecular diagnosis of LS in Singapore and the region. Methods: Fifty nine unrelated families were studied. Mutations in exons, splice-site junctions and promoters of five MMR genes were scanned by high resolution melting assay followed by DNA sequencing, large fragment deletions/duplications and promoter methylation in MLH1, MSH2, MSH6 and PMS2 were evaluated by multiplex ligation-dependent probe amplification. Tumor microsatellite instability (MSI) was assessed with five mononucleotide markers and immunohistochemical staining (IHC) was also performed. Results: Pathogenic defects, all confined to MLH1 and MSH2, were identified in 17 out of 59 (28.8%) families. The mutational spectrum was highly heterogeneous and 28 novel variants were identified. One recurrent mutation in MLH1 (c.793C>T) was also observed. 92.9% sensitivity for indication of germline mutations conferred by IHC surpassed 64.3% sensitivity by MSI. Furthermore, 15.6% patients with MSS tumors harbored pathogenic mutations. Conclusions: Among major ethnic groups in Singapore, all pathogenic germline defects were confined to MLH1 and MSH2. Caution should be applied when the Amsterdam criteria and consensus microsatellite marker panel recommended in the revised Bethesda guidelines are applied to the local context. We recommend a screening strategy for the local LS by starting with tumor IHC and the hotspot mutation testing at MLH1 c.793C>T followed by comprehensive mutation scanning in MLH1 and MSH2 prior to proceeding to other MMR genes. [ABSTRACT FROM AUTHOR]

Objective: To identify methylated genes in serum with diagnostic potentials for early colorectal cancer (CRC). Methods: Serum methylation levels of up to 12 genes were measured in two sets of serum samples with the second set from 26 stage I CRC patients and 26 age/gender-matched controls. Results: Serum methylation levels of TAC1, SEPT9, and EYA4 were significant discriminants between stage I CRC and healthy controls. Combination of TAC1 and SEPT9 rendered 73.1% sensitivity with 92.3% specificity. Conclusion: Serum methylation levels of TAC1. SEPT9 and EYA4 may be useful biomarkers for early detection of CRC though a validation study is necessary. [ABSTRACT FROM AUTHOR]

Introduction: The use of faecal immunochemical occult blood test (FIT) has been reported to decrease mortality from colorectal cancer. The Singapore Cancer Society (SCS) gives out FIT kits to encourage opportunistic screening of colorectal cancer. Any Singapore citizen or permanent resident aged ≥ 50 years is eligible to receive two FIT kits. Participants with at least one positive FIT are referred for further evaluation. We aimed to analyse the results of SCS data from the year 2008.Methods: The factors evaluated included compliance, positive test rate (PR) and positive predictive value (PPV) of FIT.Results: 20,989 participants received 41,978 kits in 2008. Compliance was 38.9%, with 8,156 participants returning at least one kit. 8% of participants tested positive, and 75% of these test-positive participants agreed to undergo further investigations. 33 participants had colorectal cancers, 45 had advanced polyps (≥ 1 cm) and 90 had polyps < 1 cm. Histologically, 114 polyps were adenomatous, 20 were hyperplastic and 1 was serrated. PPV of colorectal neoplasia for those who underwent further colonoscopy was 34%. Over half of the participants who had only one positive test had colorectal neoplasia.Conclusion: PR and PPV of FIT in our study were comparable to that in the literature. However, compliance was low and a quarter of all participants who tested positive refused further investigations. Extensive population education programmes are required to improve compliance and tackle inhibitions among the masses. It is also important to take steps to enhance the cost effectiveness of future screening programmes. [ABSTRACT FROM AUTHOR]