Alternative splicing of TIA-1 in human colon cancer regulates VEGF isoform expression, angiogenesis, tumour growth and bevacizumab resistance

© 2014 The Authors. The angiogenic capability of colorectal carcinomas (CRC), and their susceptibility to anti-angiogenic therapy, is determined by expression of vascular endothelial growth factor (VEGF) isoforms. The intracellular protein T-cell Intracellular Antigen (TIA-1) alters post-transcriptional RNA processing and binds VEGF-A mRNA. We therefore tested the hypothesis that TIA-1 could regulate VEGF-A isoform expression in colorectal cancers. TIA-1 and VEGF-A isoform expression was measured in colorectal cancers and cell lines. We discovered that an endogenous splice variant of TIA-1 encoding a truncated protein, short TIA-1 (sTIA-1) was expressed in CRC tissues and invasive K-Ras mutant colon cancer cells and tissues but not in adenoma cell lines. sTIA-1 was more highly expressed in CRC than in normal tissues and increased with tumour stage. Knockdown of sTIA-1 or over-expression of full length TIA-1 (flTIA-1) induced expression of the anti-angiogenic VEGF isoform VEGF-A 165 b. Whereas flTIA-1 selectively bound VEGF-A 165 mRNA and increased translation of VEGF-A 165 b, sTIA-1 prevented this binding. In nude mice, xenografted colon cancer cells over-expressing flTIA-1 formed smaller, less vascular tumours than those expressing sTIA-1, but flTIA-1 expression inhibited the effect of anti-VEGF antibodies. These results indicate that alternative splicing of an RNA binding protein can regulate isoform specific expression of VEGF providing an added layer of complexity to the angiogenic profile of colorectal cancer and their resistance to anti-angiogenic therapy

Common variation near CDKN1A, POLD3 and SHROOM2 influences colorectal cancer risk

We performed a meta-analysis of five genome-wide association studies to identify common variants influencing colorectal cancer (CRC) risk comprising 8,682 cases and 9,649 controls. Replication analysis was performed in case-control sets totaling 21,096 cases and 19,555 controls. We identified three new CRC risk loci at 6p21 (rs1321311, near CDKN1A; P = 1.14 × 10(-10)), 11q13.4 (rs3824999, intronic to POLD3; P = 3.65 × 10(-10)) and Xp22.2 (rs5934683, near SHROOM2; P = 7.30 × 10(-10)) This brings the number of independent loci associated with CRC risk to 20 and provides further insight into the genetic architecture of inherited susceptibility to CRC.Swedish Research Council et al.Manuscrip

Tumour-infiltrating CD8+ lymphocytes and colorectal cancer recurrence by tumour and nodal stage

Background Intratumoural T-cell infiltrate intensity cortes wrelaith clinical outcome in stage II/III colorectal cancer (CRC). We aimed to determine whether this association varies across this heterogeneous group. Methods We performed a pooled analysis of 1804 CRCs from the QUASAR2 and VICTOR trials. Intratumoural CD8+ and CD3+ densities were quantified by immunohistochemistry in tissue microarray (TMA) cores, and their association with clinical outcome analysed by Cox regression. We validated our results using publicly available gene expression data in a pooled analysis of 1375 CRCs from seven independent series. Results In QUASAR2, intratumoural CD8+ was a stronger predictor of CRC recurrence than CD3+ and showed similar discriminative ability to both markers in combination. Pooled multivariable analysis of both trials showed increasing CD8+ density was associated with reduced recurrence risk independent of confounders including DNA mismatch repair deficiency, POLE mutation and chromosomal instability (multivariable hazard ratio [HR] for each two-fold increase = 0.92, 95%CI = 0.87–0.97, P = 3.6 × 10−3). This association was not uniform across risk strata defined by tumour and nodal stage: absent in low-risk (pT3,N0) cases (HR = 1.03, 95%CI = 0.87–1.21, P = 0.75), modest in intermediate-risk (pT4,N0 or pT1-3,N1-2) cases (HR = 0.92, 95%CI = 0.86–1.0, P = 0.046) and strong in high-risk (pT4,N1-2) cases (HR = 0.87, 95%CI = 0.79–0.97, P = 9.4 × 10−3); PINTERACTION = 0.090. Analysis of tumour CD8A expression in the independent validation cohort revealed similar variation in prognostic value across risk strata (PINTERACTION = 0.048). Conclusions The prognostic value of intratumoural CD8+ cell infiltration in stage II/III CRC varies across tumour and nodal risk strata. </p

An endogenous nanomineral chaperones luminal antigen and peptidoglycan to intestinal immune cells.

In humans and other mammals it is known that calcium and phosphate ions are secreted from the distal small intestine into the lumen. However, why this secretion occurs is unclear. Here, we show that the process leads to the formation of amorphous magnesium-substituted calcium phosphate nanoparticles that trap soluble macromolecules, such as bacterial peptidoglycan and orally fed protein antigens, in the lumen and transport them to immune cells of the intestinal tissue. The macromolecule-containing nanoparticles utilize epithelial M cells to enter Peyer's patches, small areas of the intestine concentrated with particle-scavenging immune cells. In wild-type mice, intestinal immune cells containing these naturally formed nanoparticles expressed the immune tolerance-associated molecule 'programmed death-ligand 1', whereas in NOD1/2 double knockout mice, which cannot recognize peptidoglycan, programmed death-ligand 1 was undetected. Our results explain a role for constitutively formed calcium phosphate nanoparticles in the gut lumen and show how this helps to shape intestinal immune homeostasis

VICTOR : a phase III placebo-controlled trial of rofecoxib in colorectal cancer patients following surgical resection

Background: The cyclo-oxygenase-2 inhibitor, rofecoxib (R) was hypothesised to improve survival in cancer patients who had undergone surgery for colorectal cancer (CRC). This trial recruited from April 2002 until September 2004 when R was withdrawn over concerns about its cardiovascular safety (CVS). This report provides preliminary efficacy results. Methods: Recruited patients had undergone Ro resection of a stage II/III CRC and completion of adjuvant therapy (radiotherapy/chemotherapy/both/neither) less than 12 weeks previously. Excluded patients were those with active peptic ulceration, gastro-intestinal bleeding and those receiving long-term NSAID therapy (except low dose aspirin). 7,000 patients were planned to receive 25mg R daily or an identical placebo (P) for 2 or 5 years, with the goal of detecting a reduction in risk of death - hazard ratio (HR) 0.82. After the trial's premature closure, a modified protocol of the post- treatment follow-up phase and revised statistical analysis plan permitted the detection of a reduction (HR=0.75) in risk of death with 87% power, type I error 0.05, with one pre-planned event-driven interim analysis. Overall survival (OS) and disease-free survival (DFS) were both measured from randomisation, with DFS defined as the time to recurrence or death from any cause. Results: 1,167 of 1,217 patients randomised to R and 1,160 0f 1,217 randomised to P received treatment with median durations of 7.4 months and 8.2 months respectively. Median follow-up was 3.0 and 3.1 years in the two arms (R vs P), with 177 vs 191 deaths and 291 vs 316 DFS events. This pre-planned Kaplan-Meier and log- rank analysis demonstrated that the R patients had slightly longer OS than the P patients, HR 0.94 (95% CI 0.77-1.16; p=0.57). Similarly DFS was slightly higher in the R patients, HR 0.91 (95% CI 0.78-1.07; p=0.25). 19 patients in each arm died without recurrence of CRC. Conclusions: In this study of short treatment duration treatment with R is unlikely to result in a substantial improvement in OS but a small protective effect against recurrence is suggested

VICTOR : a phase III placebo-controlled trial of rofecoxib in colorectal cancer patients following surgical resection

Background: The cyclo-oxygenase-2 inhibitor, rofecoxib (R) was hypothesised to improve survival in cancer patients who had undergone surgery for colorectal cancer (CRC). This trial recruited from April 2002 until September 2004 when R was withdrawn over concerns about its cardiovascular safety (CVS). This report provides preliminary efficacy results. Methods: Recruited patients had undergone Ro resection of a stage II/III CRC and completion of adjuvant therapy (radiotherapy/chemotherapy/both/neither) less than 12 weeks previously. Excluded patients were those with active peptic ulceration, gastro-intestinal bleeding and those receiving long-term NSAID therapy (except low dose aspirin). 7,000 patients were planned to receive 25mg R daily or an identical placebo (P) for 2 or 5 years, with the goal of detecting a reduction in risk of death - hazard ratio (HR) 0.82. After the trial's premature closure, a modified protocol of the post- treatment follow-up phase and revised statistical analysis plan permitted the detection of a reduction (HR=0.75) in risk of death with 87% power, type I error 0.05, with one pre-planned event-driven interim analysis. Overall survival (OS) and disease-free survival (DFS) were both measured from randomisation, with DFS defined as the time to recurrence or death from any cause. Results: 1,167 of 1,217 patients randomised to R and 1,160 0f 1,217 randomised to P received treatment with median durations of 7.4 months and 8.2 months respectively. Median follow-up was 3.0 and 3.1 years in the two arms (R vs P), with 177 vs 191 deaths and 291 vs 316 DFS events. This pre-planned Kaplan-Meier and log- rank analysis demonstrated that the R patients had slightly longer OS than the P patients, HR 0.94 (95% CI 0.77-1.16; p=0.57). Similarly DFS was slightly higher in the R patients, HR 0.91 (95% CI 0.78-1.07; p=0.25). 19 patients in each arm died without recurrence of CRC. Conclusions: In this study of short treatment duration treatment with R is unlikely to result in a substantial improvement in OS but a small protective effect against recurrence is suggested

Phase III randomized trial assessing rofecoxib in the adjuvant setting of colorectal cancer: final results of the VICTOR trial

Purpose Laboratory and case-control studies suggest a pivotal role for the cyclooxygenase-2 ( COX-2) pathway in colorectal carcinogenesis. The purpose of this study was to test whether the COX-2 inhibitor rofecoxib could reduce recurrence and improve survival when administered in the adjuvant setting of colorectal cancer (CRC). Patients and Methods Patients who had undergone potentially curative surgery and completion of adjuvant therapy for stage II and III CRC were randomly assigned to receive rofecoxib (20 mg daily) or placebo. The primary end point was overall survival (OS). Where formalin-fixed paraffin-embedded tumor tissue samples were available, COX-2 expression was evaluated by immunohistochemistry and correlated with clinical outcome. Results Two thousand four hundred thirty-four patients were entered onto the study. The trial was terminated early because of the worldwide withdrawal of rofecoxib. At this point, 1,167 patients had received rofecoxib and 1,160 patients had received placebo for median treatment durations of 7.4 and 8.2 months, respectively. For the rofecoxib and placebo arms, median follow-up times were 4.84 and 4.85 years, with 241 and 246 deaths and 297 and 329 recurrences, respectively. No difference was demonstrated in OS (hazard ratio [HR] = 0.97; 95% CI, 0.81 to 1.16; P = .75) or recurrence (HR = 0.89; 95% CI, 0.76 to 1.04; P = .15) comparing the two groups. Tumor COX-2 expression by immunohistochemistry was assessed for 871 patients, but neither prognostic nor predictive effects were observed. Conclusion In this study of abbreviated therapy in the adjuvant setting of CRC, rofecoxib did not improve OS or protect from recurrence in unselected patients. In addition, COX-2 expression did not correlate with prognosis overall or predict effectiveness of COX-2 inhibitors