bluuebunny/crossref_metadata_2025 huggingface dataset:
Blood-based screening for methylation changes in colorectal cancer patients using novel nanotechnologies.
Authors: Ahuja Nita , Kwak Ruby , Keeley Brian , Stark Alejandro , Guzzetta Angela Anna , Wolfgang Christopher Lee , Herman James Gordon , Iacobuzio-Donahue Christine A , Wang Tza Huei
Published: February 2017

384

Background: Identification of blood-based biomarkers for cancer screening is essential in order to develop novel and minimally invasive methods for colorectal cancer screening. Our lab has successfully applied a novel nanotechnology that allows us to detect and amplify a single tumor DNA fragment in a plasma sample. This DNA is tested for methylation of several genes including TFPI2 which has shown to be highly sensitive and specific for the detection colorectal cancer in stool. Methods: Whole blood was obtained from 18 colorectal cancer patients and plasma was isolated. Plasma was processed using Methylation On Beads nanotechnology (MOB) and bisulfate treated. Methylation status was determined via quantitative PCR method. Results: Two genes, TFPI2 and IGFBP3, were detected with a high sensitivity. TFPI2, demonstrated a methylation frequency of 94.4%, which is concordant with the TFPI2 methylation frequency of 99% in primary colorectal cancer tissues. IGFBP3 showed the methylation frequency of 61.1%, which corresponds with the methylation frequency of 52% in retrospective colorectal cancer tissues in previous studies. Quantification using standard curves indicated a single copy level of DNA found in plasma. Conclusions: Blood-based screening is challenging due to extremely low quantities of circulating DNA in blood. Utilizing a novel nanotechnology that detects DNA at a single copy level, the methylation changes in colorectal cancer were successfully detected in plasmas at similar frequencies as in tissue samples. This study has demonstrated the feasablility and applicability to blood-based screening. Future studies will focus on improving the sensitivity and determining the specificity of this method.
Prediction of clinical outcome in stage II and III colon cancer by a common gene variant in AXIN2.
Authors: Szkandera Joanna , Absenger Gudrun , Weissmueller Melanie , Pichler Martin , Stotz Michael , Lax Sigurd , Leitner Gerhard , Asslaber Martin , Renner Wilfried , Gerger Armin
Published: February 2017

387

Background: Recent evidence suggests that the Wnt and Notch signaling pathways are involved in colon cancer progression and tumor recurrence. There is substantial germline genetic variability in these pathways, including single nucleotide polymorphisms (SNPs). SNPs may alter transcription, translation or splicing, thereby causing inter-individual differences in a patient’s tumor recurrence capacity and chemoresistance. We hypothesized that SNPs analyzed in a comprehensive panel of Wnt and Notch pathway genes predict clinical outcome in patients with colon cancer. Methods: A total of 815 patients with stage II and III colon cancer treated at the Medical University of Graz were included in this retrospective study. FFPE tissue specimens from normal tissue adjacent to the tumor samples were available from 599 patients. 18 SNPs in Wnt and Notch pathway genes (SFRP, DKK2, DKK3, Axin2, APC, MYC, TCF7L2 and NOTCH-2) were determined by 5’-exonuclease assay (TaqMan). The primary endpoint of the study was disease-free survival (DFS). Results: The homozygous mutant variant of AXIN2 rs2240308 G>A was associated with a significantly increased median DFS (HR 0.638, 95% CI 0.432-0.942, p=0.024) in univariate analysis. Patients carrying at least one G allele in AXIN2 rs2240308 G>A showed a median DFS of 114 months. In contrast, patients with homozygous A/A showed a median DFS of 133 months. After Cox proportional hazards model adjustment for known prognostic markers this result remained significant (HR 0.671, 95% CI 0.453-0.992, p=0.046). Conclusions: To the best of our knowledge, this is the first study identifying a common genetic variant in AXIN2 as an independent prognostic marker in stage II and III colon cancer. Larger prospective trials are warranted to confirm these findings.
Plasma levels of TIMP-1 in chemo-naive patients with metastatic colorectal cancer treated with first-line FLOX with or without cetuximab: Results from the Nordic VII Study.
Authors: Tarpgaard Line Schmidt , Guren Tormod Kyrre , Glimelius Bengt , Sorbye Halfdan , Ikdahl Tone , Christensen Ib Jarle , Qvortrup Camilla , Kure Elin , Pfeiffer Per , Tveit Kjell Magne , Nielsen Hans J. , Brünner Nils
Published: February 2017

392

Background: KRAS status is presently the best known biomarker to select patients with metastatic colorectal cancer (mCRC) for therapy with EGFR inhibition. However, in NORDIC VII a survival benefit of adding cetuximab to the Nordic FLOX regimen could not be confirmed. Plasma Tissue Inhibitor of MetalloProteinases-1 (TIMP-1) is a promising biomarker in patients with mCRC, and has multiple actions including regulation of apoptosis, angiogenesis, cell growth and differentiation. High levels of TIMP-1 associate with reduced sensitivity to irinotecan-based treatment in patients with mCRC. We investigated whether high plasma TIMP-1 also correlates with response rate, progression-free survival (PFS) and overall survival (OS) in patients with mCRC treated in NORDIC VII. Methods: In NORCIC VII 571 patients with mCRC were randomized to: (A) Nordic FLOX; (B) Nordic FLOX + cetuximab or (C) Nordic FLOX intermittently + cetuximab continuously. Baseline plasma samples for TIMP-1 analysis were available from 426 patients (79%). Plasma TIMP-1 was determined using the MAC15 antibody in-house validated kinetic Enzyme Linked Immunosorbent Assay. Results: Median plasma TIMP-1 was 296 ng/mL. The tumor was KRAS mutated in 149 patients (35%). Best response did not correlate to TIMP-1 values (OR=1.16, 0.91-1.49, p=0.22). High plasma TIMP-1 was associated with short PFS (HR=1.22, 1.07-1.39, p=0.003) and OS (HR=1.55, 1.36-1.80, p<0.0001). Multivariate analysis (TIMP-1, age, gender, KRAS, ALP, WBC, platelets, CRP, CEA, WHO PS, no. of metastatic sites) demonstrated that high plasma TIMP-1 was not an independent biomarker of PFS (HR=0.95, 0.76-1.18, p=0.65) or OS (HR=1.17, 0.91-1.50, p=0.22). Multivariate analysis retained CRP, PS and no. of metastatic sites in the final model for PFS and alkaline phosphates, PS and no. of metastatic sites for OS. Conclusions: Plasma TIMP-1 is a new prognostic biomarker in patients with mCRC treated with 1st line Nordic FLOX +/- cetuximab. Due to the design of NORDIC VII, a predictive effect of plasma TIMP-1 could only be estimated for cetuximab treatment and no significant treatment by marker intervention was observed.
Demonstration of subpopulations with differing cancer stem cell phenotypes in xenograft and in vitro models of colorectal liver metastases.
Authors: Sanford Dominic E. , Giorgi Andrew , Goetz Brian D. , Panni Roheena Z. , Hawkins William G. , Linehan David , Goedegebuure Peter S , Fields Ryan C.
Published: February 2017

394

Background: Tumors are composed of heterogeneous cell populations, some of which demonstrate enhanced tumor-forming capabilities (so-called tumor initiating cells [TIC] or cancer stem cells). In colorectal cancer (CRC), CD133, 44, and 24 are cell surface markers that identify TIC. Therefore, we sought to determine if CRC liver metastases (CRC-LM) form xenografts (in vivo) and cell cultures (in vitro) with TIC markers. Methods: CRC-LM were grafted in NOD/SCID mice and passaged serially. Xenografts were mechanically dissociated and cultured under sphere forming conditions. Flow cytometry was performed for TIC phenotype. Results: 16 of 18 (89%) CRC-LM specimens formed tumors in mice. Xenografts formed EpCAM+ tumors and spheres. The frequency of CD133+, CD44+, and CD133+/CD44+ tumor cells were 55%, 33%, and 23%, respectively. There was a subpopulation of CD133+/CD44+ cells with elevated CD44 expression(CD44hi). This CD133+/CD44hi population was also CD24+; representing 5% of cells. Eight of eleven (73%) xenografts formed spheres in vitro. The frequency of CD133+, CD44+, and CD133+/CD44+ cells were 63%, 47%, and 26%, respectively. CD133+/CD44+/CD24+ cells made up 8% of sphere-forming cells. There was a non-significant trend towards increased frequency of CD133+, CD44+, and CD133/CD44 positive cells in the spheres compared to the xenografts. However, the percentage of CD133+/CD44+/CD24+ cells was significantly increased in spheres relative to xenografts (8% vs. 5%, respectively; p<0.05) (see Table). Conclusions: CRC-LM derived xenografts and spheres are composed of distinct cell populations with differing levels of TIC/cancer stem cells. Sphere cultures may enhance for the most enriched TIC population. Thus, xenografts and sphere cultures are important model systems to further study the importance of cancer stem cells in CRC progression and metastases. [Table: see text]
Association of CD133 expression levels with the k-ras mutation status in rectal cancers before and after preoperative radiochemotherapy.
Authors: Sprenger Thilo , Gaedcke Jochen , Conradi Lena-Christin , Jo Peter , Jung Klaus , Beissbarth Tim , Homayounfar Kia , Ghadimi B. Michael , Liersch Torsten
Published: February 2017

400

Background: The role of the potential stem cell marker CD133 as a predictive or prognostic marker in multimodal rectal cancer treatment is currently under debate. While CD133 has been identified as a prognostic marker in rectal cancers after preoperative radiochemotherapy (RCT) it was recently characterized as a very unspecific feature for colorectal cancer stem cells. We therefore analyzed the association between CD133 expression and mutations in the proto-oncogenes K-Ras and PI3K in rectal cancer patients receiving neoadjuvant RCT. Methods: CD133 expression was evaluated immunhistochemically in pre-treatment biopsies and surgical specimens of 128 patients with locally advanced rectal cancers (cUICC II/III) treated with preoperative RCT within the phase-III German Rectal Cancer Trials. K-Ras mutations were analyzed by sequencing of exons 1, 2, and 3. PI3K mutations were detected by sequencing the p110α subunit (PIK3CA) and correlated with histopathologic parameters, tumor regression and survival. Results: CD133 expression was significantly associated with mutations in the K-Ras gene in both pre-treatment biopsies and post-treatment tumor specimens in uni- and multivariate analyses. However, no significant correlation was observed between CD133 and PI3K mutations. Post-treatment CD133 levels were correlated with neoadjuvant RCT (50.4 Gy/5-FU vs. 50.4 Gy/5-FU+Ox) and tumor regression grading. Anyway, there was no significant association between pre- and post-treatment CD133 expression and disease-free survival. Conclusions: CD133 expression levels are strongly associated with mutations in the K-Ras proto-oncogene in rectal cancers before and after preoperative RCT. Our results strengthen the hypothesis that CD133 is not a specific marker for colorectal stem cells but might be integrated in proliferation pathways like the ras-raf axis.
Micro-RNA-21 (miR21) expression in colorectal cancer (CRC) as a predictor for fluoropyrimidine-based adjuvant chemotherapy.
Authors: Lin Albert Y. , Kouzminova Natalia B. , Pollack Jonathan , Nuovo Gerard
Published: February 2017

408

Background: Fluoropyrimidine-based adjuvant chemotherapy is a standard treatment option for patients with high risk II or stage III CRC. It is, however, unclear if a subset of patients will benefit from chemotherapy more than others. MiR21, a small non-coding RNA, has been associated with promotion of tumor cell growth and metastasis. To assess the effect of miR21 on chemotherapy, we analyzed the association of miR21 expression with clinical outcomes and known prognostic factors. Methods: MicroRNA detection was performed by in situ hybridization on a CRC tissue microarray containing specimens from 130 cases (stage I, 21 patients; II: 44; III: 33; IV: 32). MiR21 expression was graded as negative (no staining in all tissue cores), low (0-10% staining), moderate (20-40%) and strong (50-100%), and was analyzed with stage, grade, expression of VEGF, Ki67, LEF1, OPN and MSH2 by immunohistochemistry. Cox proportional hazards regression analysis was performed to assess the association of miR21 expression with 10-year recurrence-free survival in the subgroup of 77 stage II or III patients. Results: In all, miR21 expression had moderate positive correlation with genes associated with tissue proliferation and invasion, including Ki67 (Spearman's r=0.42, p<0.001), VEGF (r=0.32, p<0.001) and OPN (r=0.32, p<0.001), and weak correlation with LEF1 (r=0.22, p=0.012) and MSH 2 (r=0.18, p=0.039). In the subgroup of 36 patients treated with adjuvant therapy, low or negative miR21 expression (≤10%, n=15) was associated with increased recurrence (HR =3.5; p=0.05). In the multivariate Cox regression model including stage, grade and LEF1 expression, the association of low or negative miR21 with cancer recurrence remained significant along with grade and LEF1. Ki67 was excluded from the multivariate model because of significant association with miR21 (r=0.44, p=0.007) in the analyzed group. Conclusions: Our results suggest that low miRNA21 expression is associated with increased risk for recurrence in CRC patients received adjuvant chemotherapy. Further investigation of miR21 in randomized studies is warranted to establish its role as a predictor for CRC in the setting of adjuvant therapy.
Occurence of initiation and progression of colorectal cancer in the changed intestinal environment.
Authors: Ohigashi Seiji , Sudo Kazuki , Kobayashi Daiki , Takahashi Osamu , Takahashi Takuya , Asahara Takashi , Nomoto Koji , Onodera Hisashi
Published: February 2017

416

Background: Use of new molecular biology–based methods of bacterial identification is expected to help elucidate the relationship between colorectal cancer (CRC) and intestinal microbiota. However, it remains unclear whether microbial dysbiosis is the cause or the result of CRC onset. We analyzed the intestinal environments to determine whether the changes differed with the stage of CRC or adenoma. Methods: We analyzed fecal microbiota, organic acid concentrations, and pH in CRC patients (n=93), individuals with adenoma (n=23), and individuals with normal intestinal tracts (n=27). After patient hospitalization, the feces of all subjects were collected before any administration of laxatives or antibiotics to prepare the bowel.Thirteen bacterial groups were enumerated in the fecal microbiota by using reverse transcription–quantitative PCR (RT-qPCR). Eight kinds of organic acid were quantified using high-performance liquid chromatography, and fecal pH was measured by pH meter. Results: The counts of total bacteria (10.3 ± 0.7 vs. 10.8 ± 0.3 log10 cells/g of feces; p<0.001), 5 groups of obligate anaerobe (Clostridium coccoides group, C. leptum subgroup, Bacteroides fragilis group, Bifidobacterium, and Atopobium cluster), and 2 groups of facultative anaerobes (Enterobacteriaceae and Staphylococcus) were significantly lower in the CRC group than in the healthy individuals. While the concentrations of organic acids—particularly short chain fatty acids (SCFAs) such as acetic acid, propionic acid, and butyric acid—were significantly decreased in the CRC group, the pH was increased in the CRC group (7.4 ± 0.8 vs. 6.9 ± 0.6; p<0.001). Comparison among the CRC, adenoma, and non-adenoma groups revealed that fecal SCFAs concentrations and pH in the adenoma group were intermediate to the CRC group and the non-adenoma group. Within the CRC group, no differences in microbiota or organic acids were observed among T-stages or Dukes stages. Conclusions: CRC patients showed significant differences in the intestinal environment, including alterations of microbiota, decreased SCFAs, and elevated pH. These changes are not a result of CRC progression but are involved in CRC onset.
Association between genetic variations involved in tumor dormancy related genes and clinical outcomes in Japanese patients with gastric cancer.
Authors: Wakatsuki Takeru , Páez David , Zhang Wu , Labonte Melissa Janae , Yang Dongyun , Azuma Mizutomo , Ning Yan , Volz Nico , Stintzing Sebastian , Bohanes Pierre Oliver , El-Khoueiry Rita , Koizumi Wasaburo , Watanabe Masahiko , Lenz Heinz-Josef
Published: February 2017

43

Background: The major cause of cancer-related death is metastatic growth of disseminated tumor cells (DTCs) from the primary tumor despite approximate resection and adjuvant chemotherapy. Tumor dormancy is a stage in cancer progression in which micrometastases or residual tumor remains occult and asymptomatic until clinical appearance. The regulation of whether DTCs keep to a quiescent state or transit into a proliferative state can be explained through the interaction of the DTCs with the microenvironment, blood supply, and immunosurveillance. This suggests that host genetic background may impact this process. We tested our hypothesis whether polymorphisms involved in tumor dormancy related genes could predict clinical outcomes in gastric cancer (GC). Methods: One-hundred and sixty-nine Japanese patients with histolopathologically confirmed localized GC (stage Ib to IV; AJCC-6th) were enrolled from Kitasato East University Hospital and Fukushima Red Cross Hospital between 2002 and 2010. Twenty-four polymorphisms were selected from VEGF pathway genes, microenvironment related genes, and metastasis suppressor genes. Genomic DNA was extracted from blood or tissue, and all samples were analyzed by PCR-based direct DNA-sequencing. Results: The median age was 68 (31-88) years-old, and 64% of patients were males in this cohort. Pathological stages were 17% in stage Ib, 31% in stage II, 36% in stage III, and 17% in stage IV,. Almost all patients received D2 lymphadectomy based surgery and 65% patients received S-1 based adjuvant chemotherapy. The median follow-up periods were 4.0 years. NME1 rs2302254 was associated with significantly shorter disease-free survival (HR 1.555; 95%CI, 1.026-2.359; p=0.032). NME1 rs34214448 (HR 1.732; 95%CI, 1.082-2.770; p=0.036), PAI1 rs1799889 (HR 1.855; 95%CI, 1.032-3.333; p=0.02), and VEGF-A rs699946 (HR 1.678; 95%CI, 1.014-2.778; P=0.042) were associated with significantly shorter overall survival. Conclusions: NME1 rs2302254 may predict early recurrence of Japanese patients with GC. Additionally, survival after recurrence could be influenced by angiogenic factors in a synergic manner.
The prostaglandine E2 transporting organic anion transporting polypeptide OATP4A1: A potential prognostic marker in colorectal cancer?
Authors: Buxhofer-Ausch Veronika , Ausch Christoph A. , Bauer Heike , Mollik Marina , Larijani Aida , Bajna Erika , Svoboda Martin , Kallay Enikoe , Reiner Angelika , Kriwanek Stephan , Sebesta Christian , Hamilton Gerhard , Zeillinger Robert , Thalhammer Theresia
Published: February 2017

430

Background: Organic anions transporters (OATPs) are important for tumor progression and therapeutic response by regulating cellular levels of hormones, second messenger proteins and drugs. OATP4A1 is a transporter of pro-inflammatory prostaglandin E2 and may contribute to cancer progression. Data on the expression of OATP4A1 and its clinical impact in primary colorectal cancer (CRC) is rare. Our study was designed to proof the overexpression of OATP4A1 in primary CRC. Methods: Frozen samples from 20 unselected CRC patients (pat) and five CRC cell lines were analyzed for OATP4A1 mRNA expression by real time PCR (mean level normalized to the calibrator, MNE). Immunohistochemistry was performed on paraffin- embedded tumor sections from 50 CRC pat., UICC 0-II (25/50 with subsequent relapse). An automatic quantitative image analysis program was applied to quantify OATP4A1 expression. Expression and intensity was correlated with clinical parameters and relapses. Results: Significant (p>0.05) higher levels of OATP4A1 mRNA were observed in 20 cancer samples as compared to adjacent non-cancerous tissue (2.44 vs. 0.46 MNE). The highest expression (9.85 MNE) was observed in a well-differentiated tumor sample. Similar high levels were observed in the COGA1A cell line, expression in the other cell lines ranged between 1.83 and 0.28 MNE. Immunoreactive staining for OATP4A1 was located in the membrane and occasionally in the cytosol of tumor cells, it was exclusively membrane located in the adjacent non-cancerous epithelial cells. The staining intensity was significantly higher in cancer cells compared to non-cancerous areas (1528±326 vs.376±218) while staining of stroma cells was only occasionally detectable. Surprisingly, the highest OATP4A1 levels were observed in immune cells (2839±381 vs.298±56). Data on the clinical impact of OATP4A1 in the early stage CRC pat. will be presented at the meeting. Conclusions: The profound expression of OATP4A1 in CRC cells and in the inflammatory infiltrates supports its implication on cancer progression. Suitability of OATP4A1 as a potential prognostic marker has to be established on a larger patient collective.
Effect of hangeshashinto on wound healing in hamster 5-fluorouracil-induced oral mucositis model with increased keratinocyte migration and reduced inducible prostaglandin E2.
Authors: Kono Toru , Kaneko Atsushi , Matsumoto Chinami , Hibino Tomoko , Shigenobu Tatsuro , Miyano Kanako , Fukutake Masato , Uezono Yasuhito
Published: February 2017

432

Background: Chemotherapy-induced oral mucositis (COM) is common, but optimal treatment is not established. COM is characterized by prolonged damage with painful inflammation that involves pathological pain-evoking prostaglandin E2 (PGE2). We previously found that hangeshashinto (HST), a Japanese herbal medicine, was effective as a gargle for the treatment of COM and also reduced ulcer healing time in patients with grade 2 or worse COM by half in a randomized, placebo-controlled, double-blind clinical trial. However, little is known regarding the mechanisms underlying the anti-COM response elicited by HST. Thus, the aim of this study was to clarify the effects of HST on cell migration and PGE2 system. Methods: COM was induced in hamsters by a combination of 5-fluorouracil administration and mild abrasion of the cheek pouch, and healing was examined by measuring lesion size. Hamsters were given a diet containing 2% HST or a control diet throughout the experiments. To estimate the direct effects of HST, human oral keratinocytes (HOK) were used to assay migration and PGE2 production. Migration was evaluated 9–24 h after scratch wounding using sterile pipette tips. PGE2 was induced by addition of 10 ng/mL interleukin-1β, and the amount of PGE2 in 6 h culture fluids were measured by EIA. Results: Cheek pouch lesions induced by abrasion were aggravated by 5-fluorouracil. Lesions were significantly smaller in the HST group than in the control group. Migration of HOK was time-dependently increased by HST (30–100 µg/mL). Inducible PGE2 production was reduced by HST without cytotoxicity, while constitutive PGE2 was unchanged. Gene expressions of cyclooxygenase-2, phospholipase A2, and prostaglandin E synthase were down-regulated by exposure to HST, but cyclooxygenase-1 was not affected. Screening tests for active ingredients showed that berberine promoted scratch wound closure in HOK, and that gingerols, shogaols, and flavonoids significantly decreased PGE2. Conclusions: Our data suggest that the anti-COM mechanisms of HST involve the acceleration of wound healing in hamster COM model and inhibition of inducible PGE2 production in HOK.
Prediction of pathologic response of colorectal liver metastases (CRLM) to transcatheter hepatic therapy with irinotecan-eluting beads by hepatic activation of irinotecan.
Authors: Jones Robert Peter , Sutton Paul , Santoyo-Castelazo Anahi , Malik Hassan Zakria , Fenwick Stephen W , Terlizzo Monica , Poston Graeme John , Kitteringham Neil
Published: February 2017

438

Background: For patients with CRLM, good pathologic response to neoadjuvant chemotherapy is associated with improved overall survival. However, systemic therapy can increase postoperative morbidity and mortality. DEBIRI gives sustained delivery of drug directly to tumor, maximising response and reducing systemic exposure. Predictive markers of response would allow selection of patients most likely to benefit. This study assessed drug metabolism within normal hepatic parenchyma as a predictor of pathologic response of CRLM to DEBIRI. Methods: Patients with resectable CRLM received single targeted DEBIRI (up to 200mg) 1 month pre-hepatectomy. Hepatic parenchyma was harvested at resection. Parenchymal drug metabolism and protein expression were characterised with a novel LC-MS/MS assay and western blot, and correlated with pathologic tumour response. Results: Ten patients assessed. Median residual tumour 30% (range 10-50%), necrosis 40% (range 40-60%) and fibrosis 20% (range 10-40%). Microsomal formation of SN-38 was 0.52-2.19 nmol/mg/min (mean 1.02), SN-38G 0.30-0.59 nmol/mg/min (mean 0.43), APC 0.1-0.24 nmol/mg/min (mean 0.18). There was a negative correlation between SN-38 production and degree of SN-38 glucuronidation (r2=0.42, p=0.04), with no difference between heterozygous/wild-type UGT1A1*28 patients. SN-38 production demonstrated a strong correlation with CES-2 expression (r2=0.96, p<0.001), with a strong correlation between the glucuronidation of SN-38 and UGT1A1 expression (r2=0.76, p=0.01). A moderate correlation between microsomal SN-38 production and degree of tumour necrosis was obtained (r2=0.30, p=0.07). A highly significant correlation between microsomal SN-38 production and degree of tumour fibrosis was observed (r2=0.54, p=0.01). Conclusions: Metabolism of irinotecan varies widely. Production of the active metabolite SN-38 is directly mediated by CES-2, and hepatic production of SN38 predicts degree of tumour fibrosis after DEBIRI. Metabolic profiling may offer a novel predictor of response to DEBIRI.
Prevalence of DPYD gene mutations among patients receiving 5-FU therapy.
Authors: Saam Jennifer , Bowles Karla , Korst Laurie , Kline Kendel , Roa Benjamin , Wagner Susanne , Eldredge Glenn , Abbott Brian L. , Kaldate Rajesh R.
Published: February 2017

447

Background: Dihydropyrimidine dehydrogenase (DPYD) is the major enzyme that metabolizes 5-Fluoruracil (5-FU), a component of many chemotherapy regimens. Patients with a DPYD gene mutation have higher 5-FU plasma levels, leading to a 50-60% risk of a grade 3-4 toxicity. DPYD mutations have an estimated prevalence of 3-5% in the general population, but full sequencing is rarely performed in published studies. Analyses of the largest set of full DPYD gene sequencing test results from a commercial laboratory database are presented. Methods: A set of 3,083 patients was analyzed. Patient demographics (age, gender, ethnicity) and pre-test grade 3-4 toxicity status (none, 5-FU related, other) were obtained from the test request form. Descriptive analyses were performed to estimate mutation prevalence overall, and by toxicity and ancestry classifications, as well as to characterize mutations of interest. A subset of 24 patients tested for DPYD mutations in response to high 5-FU exposure levels was also analyzed. Results: The overall DPYD mutation prevalence was 7.3%. Mutations were present in 4.7% and 10.3% of patients experiencing none and at least one 5-FU related toxicity pre-test, respectively. Among patients with toxicities pre-test, the prevalence increased from 7.8% to 31.6% for those experiencing a single to all four toxicity types. Among 5-FU related toxicities, mutation prevalence was highest (18.5%) for hematopoietic events. The previously reported founder mutation IVS14+1G>A had a relative prevalence 42.7%, but 30.9% of these mutations were seen in patients not reporting a Western/Northern European ancestry. Among the subset of patients with high 5-FU exposure levels, 29% had a DPYD mutation showing a genetic causality. Conclusions: 5-FU in chemotherapy regimens remains widespread, yet DPYD gene testing utilization remains minimal. Most testing occurs post-treatment in response to a severe toxicity rather than pre-treatment, which would permit physicians to adapt treatment and reduce toxicity risk. Compared to previous studies, this study using full sequencing data from 3083 patients provides robust estimates of DPYD mutation prevalence and helps characterize DPYD mutations of particular interest.
Mutational analysis of locally advanced rectal cancer and response to neoadjuvant chemoradiation.
Authors: Russo Andrea Lyn , Ryan David P. , Borger Darrell R. , Wo Jennifer Yon-Li , Szymonifka Jackie , Kwak Eunice Lee , Blaszkowsky Lawrence Scott , Clark Jeffrey W. , Allen Jill N. , Zhu Andrew X. , Berger David L. , Cusack James C. , Haigis Kevin M. , Hong Theodore S.
Published: February 2017

449

Background: Preoperative chemoradiation (CRT) is the standard of care for locally advanced rectal adenocarcinoma and leads to pathologic complete response (pCR) rates of 8-20%. pCR has been correlated with improved long term outcomes. Known mutations may contribute to lack of pCR. We retrospectively evaluated our single institution experience to determine rates and potential mutational and clinical predictors of pCR. Methods: With IRB approval, we reviewed 79 consecutive patients with stage II/III rectal adenocarcinoma, diagnosed from 7/2005–6/2010. Patients received preoperative CRT to 50.4 Gy and concurrent 5-FU based chemotherapy prior to total mesorectal excision. Primary endpoint was pCR rate by Dworak tumor regression grade. Mutations were analyzed on pre- and/or post-treatment tissue available for 47 patients with a multiplexed clinical assay on nucleic acid (SNaPshot), which tests for 130 mutations across 15 cancer genes. 32 patients did not have enough tissue for analysis. Other variables including T/N stage, grade, circumferentiality, distance to anal verge, tumor size (continuous, per cm), pre and post-CRT CEA levels, and treatment duration were evaluated using logistic regression analysis. Results: Median age was 58 years and 85% had stage III disease. Among 54% of patients with mutations assessed, the following were present: 37% KRAS, 3% APC, 3% BRAF, 5% NRAS, 4% PIK3CA, 8% TP53. TP53 testing was limited to common mutation sites. In the entire cohort, 20% had a pCR. No patients with BRAF, NRAS or KRAS mutations had a pCR. Patients with a pCR were more likely to have a post-RT CEA ≤ 2.5 (93% vs 62%, p=0.002) and no mutations identified (100% vs 38%, p=0.019). pCR patients had a trend to lower rates of KRAS mutations (0% vs 41%, p=0.1). On UVA, predictors of pCR included: post-RT CEA ≤ 2.5 (OR 8.6, 95% CI 1.03 – 71.9, p=0.047), insufficient tissue to analyze (OR 5.1, CI 1.45-18, p=0.011), and smaller tumor size (OR 0.69, CI 0.48 – 1.0, p=0.05). With a median follow-up of 30 months, there were 3 local and 6 distant recurrences. None with a pCR developed recurrence. Conclusions: Patients with the composite characteristics of being BRAF, KRAS, and NRAS wild type rectal cancer are more likely to have a pCR with preoperative CRT.
Comparative analysis of second and subsequent chemotherapy lines on short- and long-term survival of elderly Medicare metastatic colon cancer patients.
Authors: Hanna Nader , Onukwugha Ebere , Bikov Kaloyan A , Zheng Zhiyuan , Seal Brian S. , Mullins C. Daniel
Published: February 2017

455

Background: Metastatic colon cancer (mCC) patients often receive multiple lines of chemotherapy as treatment (TX) to improve survival or quality of life, yet the “real world” benefits and risks of multiple TX lines have not been fully examined. Methods: Elderly (65+) SEER-Medicare patients diagnosed with mCC in 2003-2007 were followed until death or 12/31/09 to examine the survival benefits for different chemotherapy lines. The median time between diagnosis date and the starting date of 2nd line was 352 days. Therefore, we restricted comparative analysis of 2nd and subsequent chemotherapy TX lines to patients who survived at least 1 year after mCC diagnosis date. We used Cox regression framework and adjusted for patients’ TX and censoring histories by using inverse probability weighting method. Separate analyses were conducted for short (2 years) and long-term (5 years) survival to examine different benefits of 2nd and subsequent chemotherapy lines. Results: Of 2,600 elderly Medicare mCC patients diagnosed between 2003-2007 and who survived at least 1 year, 2,530 were dead by the end of 2009. Significant factors associated with long-term survival were 1st line therapy(HR = 0.76; p < 0.01), 2nd line therapy (HR = 0.83; p < 0.01) , and subsequent chemotherapy line therapy(HR = 0.85; p = 0.04), as compared to no therapy, age groups 95+ (HR = 3.07; p < 0.01), 85-94 (HR = 1.33; p < 0.01), and 75-84 (HR = 1.10; p = 0.04) as compared to 65+-74, Asian vs. White (HR = 0.71; p < 0.01), and zip code level household median income (HR = 0.98; p = 0.01). For short-term survival, the benefits of 2nd and subsequent chemotherapy lines were maintained until month 29. Patients with poor performance status were less likely to proceed to 2nd line therapy. No statistically significant variables predicting receipt of subsequent chemotherapy lines were identified. Conclusions: Among elderly Medicare mCC patients who survived at least 1 year after diagnosis, 1st line therapy improved both short and long-term survival. 2nd and subsequent chemotherapy line therapy reduced short-term mortality (2 years); however, they didn’t add any additional long term survival benefit (5 years) as compared to 1st line therapy.
Survival impact of surgical resection of primary tumor (SRPT) in metastatic colorectal cancer (mCRC): A population-based cohort study.
Authors: Ahmed Shahid , Leis Anne , Kanthan Selliah , Karim Safiya , Perron Thomas , Haider Kamal , Reeder Bruce , Pahwa Punam
Published: February 2017

456

Background: Currently there is very low quality evidence available regarding survival benefit of SRPT in patients with mCRC. In the absence of randomization the reported benefit may reflect selection of younger and healthier patients with good performance status (PS). We have undertaken a retrospective cohort study to determine the survival benefit of SRPT in mCRC. Methods: A cohort of 1,378 patients with mCRC diagnosed between 1992-2005 in the province of Saskatchewan was evaluated. Kaplan-Meier curves were used to determine survival. Log-Rank test was done to compare survival between the two groups. Cox regression model was used to adjust survival for important prognostic variables. Results: Median age was 70 yrs (22-98) and M:F was 57:43. 27% had ECOG PS of >1 and 62% had a comorbid illness. 944 (69%) patients underwent SRPT. Among 1,378 patients, 40% were operated for tumor related symptoms (33% obstruction, 6% perforation, and 4% heavy bleeding, mutually nonexclusive). Median follow up time for whole cohort was 7.1 months (inter-quartile range 2.5-17.5). 42.3% received chemotherapy and 19.1% received 2nd generation therapy. Median survival of patients who received chemotherapy was 15.9 months. Patients who underwent SRPT had median overall survival of 18.3 months vs. 8.4 months if they did not have surgery (p<0.0001).On multivariate analysis 5FU-based chemotherapy (HR 0.53; 95%CI: 0.45-0.61), metastesectomy (HR 0.54; 95%CI: 0.45-0.64), SRPT (HR 0.55; 95%CI: 0.48-0.62), and 2nd generation chemotherapy (HR 0.65; 95%CI: 0.54-0.77) were correlated with a better survival whereas, elevated CEA level (1.56; 95%CI: 1.30-1.90), leukocytosis (HR 1.54; 95%CI: 1.33-1.80), ECOG PS >1 (HR 1.48; 95%CI: 1.30-1.69), low albumin (HR 1.44; 95%CI: 1.26-1.64), age ≥ 65 yrs (HR 1.21; 95%CI: 1.10-1.38), anemia (HR 1.16; 95%CI: 1.03-1.31), and symptomatic disease (HR 1.12; 95%CI: 1.0-1.26) were correlated with poor survival. Comorbid illness, smoking, and gender did not correlate with survival. Conclusions: This is the first large cohort study that reveals that SRPT in patients with mCRC improves survival independent of chemotherapy, age, functional status and comorbid illness.
Phase II trial of combined chemotherapy with irinotecan, S-1, and bevacizumab (IRIS/Bev) in patients with metastatic colorectal cancer (mCRC): Final analysis—Hokkaido Gastrointestinal Cancer Study Group (HGCSG) trial.
Authors: Yuki Satoshi , Komatsu Yoshito , Fukushima Hiraku , Sasaki Takahide , Amano Toraji , Miyagishima Takuto , Hatanaka Kazuteru , Nakamura Michio , Tateyama Miki , Sakata Yuh
Published: February 2017

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Background: The FIRIS study (Muro K et al. Lancet Oncol 2010;11:853–860) previously demonstrated the non-inferiority of Irinotecan plus S-1 (IRIS) to FOLFIRI for mCRC, with progression-free survival (PFS) as the primary endpoint. We previously reported that IRIS plus bevacizumab (IRIS/Bev) is very effective as first-line treatment (Komatsu Y et al. Acta Oncol. 2012 May 4). We now report the final results of this study. Methods: Eligible patients had to have mCRC with a confirmed diagnosis of adenocarcinoma, an age of >20 years, ECOG performance status (PS) of 0-1, and no history of prior chemotherapy. S-1 (40-60 mg depending on patients body surface area) was given orally, twice daily for 2 consecutive weeks, and 100 mg/m2irinotecan and 5 mg/kg bevacizumab was given intravenously on day 1 and 15, followed by 2-week rest period, within a 4-week cycle. The primary endpoint was safety. The secondary endpoints included overall response (OR), PFS, and overall survival (OS). Results: A total of 53 patients were enrolled from October 2007 through March 2009. The results were reported for 52 patients with evaluable lesions. The clinical characteristics of the patients were as follows. The median age was 63.5 years (range, 48 to 82). The male:female ratio was 3:2. The most common grade 3 or 4 adverse events were neutropenia (27%) and diarrhea (17%). Grade 3 or 4 hypertension (21%) was attributed to bevacizumab, but there were no life-threatening adverse events, such as gastrointestinal perforation. On an intention-to-treat basis, OR was 63.5% [95% confidence interval (CI) 50.4-76.5%], and the disease control rate was 94.2%. With a median follow-up time of 51.7 months, median PFS was 17.0 months (95%C.I. 14.2-19.8 months) and median survival time was 39.6 months (95%C.I. 34.2-45.0 months). Conclusions: IRIS/Bev is a remarkably active and generally well-tolerated first-line treatment for patients with mCRC. Randomized control trial comparing this regimen with oxaliplatin containing regimen(XELOX or mFOLFOX6 plus bevacizumab: TRICOLORE study) is already started. Clinical trial information: NCT00569790.
Multicenter phase II study of FOLFOX or biweekly XELOX and cetuximab as first-line treatment in patients with wild-type KRAS/BRAF metastatic colorectal cancer (mCRC) (FLEET study).
Authors: Kim Ho Min , Soda Hitoshi , Hazama Shoichi , Takahashi Takao , Nagata Naoki , Kotaka Masahito , Kono Emiko , Terada Itsurou , Fukunaga Mutsumi , Oba Koji , Hasegawa Junichi , Sakamoto Junichi , Mishima Hideyuki
Published: February 2017

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Background: Cetuximab and chemotherapy as first-line therapy for patients with KRAS wild type prolong survival. However, COIN trial has not demonstrated the survival benefit of FOLFOX or XELOX and cetuximab therapy. Few data are available on its benefit for patients with KRAS and BRAF wild-type. Methods: The aim of this study was to assess the efficacy of first-line FOLFOX or bi-weekly XELOX and bi-weekly cetuximab in KRAS/BRAF wt mCRC. Chemonaive patients received FOLFOX or biweekly XELOX (oxaliplatin 85 mg/ m2/day 1 plus capecitabine 2000/m2/days 1-7) and biweekly cetuximab 500mg m2/ day 1 every 2 weeks. Primary endpoint was response rate(RR), other secondary endpoints were PFS, OS, DCR, safety, DI and resection rate. KRAS test (codon12,13) and BRAF test (V600E) by direct sequence were performed in Yamaguchi University. Patients with KRAS/BRAF wt were enrolled in this study. The regimen of FOLFOX or XELOX were selected by investigator’s preference, not randomized. Results: From April 2010 to May 2011, 139 pts were preregistered. KRAS and BRAF were examined from paraffin-embedded sample. 70 (50.3%) pts were KRAS/BRAF wt, and 62 pts were enrolled: The main characteristics of the entered pts were: sex (M/F) 34/28, median age 66 yrs (range 34-83 yrs). Grade 3/4 adverse events were leucopenia 4.8%, neutropenia 25.8%, skin toxity (paronychia/fissure) 9.7%, and acne 9.7%. Two CR (3.2%) and 40 PR (64.5%), 15 SD (24.2%) and 3 PD (4.8%) 2NE were observed, with an overall response rate of 67.7% and a disease control rate (CR+PR+SD) of 91.9%. The RR of FOLFOX or XELOX were 64.9% (24/37) and 72.0% (18/25), DCR were 89.2% and 96% respectively. Conclusions: FLEET was the first multicenter phase II study with prospective KRAS/BRAF analysis as a predictive biomarker for cetuximab in first-line mCRC in Japan. Results of this study indicate that both biweekly combination regimens are feasible, tolerable, and clinically active. Biweekly XELOX+cetuximab study (FLEET2) is ongoing. Clinical trial information: UMIN000003253.
A novel molecular tool for lymph node upstaging in colon cancer patients: Results of a prospective European, multicenter study.
Authors: Zuber Markus , Geppert Carol-Immanuel , Bader Franz , Nitsche Ulrich , Rosenberg Robert , Zettl Andreas , Tarragona Jordi , Matias-Guiu Xavier , Croner Roland
Published: February 2017

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Background: A new diagnostic semi-automated system, One Step Nucleic Acid Amplification (OSNA), was designed for the detection of cytokeratin 19 (CK 19) mRNA in lymph node (LN) metastases of colon cancer (CC) patients (Güller U et al. CANCER 2012: DOI 10.1002/cncr.27667). Recently it has been shown that occult metastases detected in pN0 patients have an adverse effect on survival based on stage migration. In this study we investigated whether pN0 patients diagnosed by standard histopathology can be upstaged by OSNA analysis to stage III. Methods: In this international, multicenter (4 institutions: CH, G [2], E) study 103 patients-originally diagnosed as pN0 by the standard use of 1 single section haematoxylin & eosin (H&E)-were subjected to OSNA analysis. LN were isolated from fresh tissue, homogenised, and directly subjected to OSNA without prior isolation of mRNA. OSNA results were provided in qualitative categories (- / + / ++), along with the exact CK19 mRNA copy number (+: 250-5000 copies, ++: >5000 copies). Results: In total, 1,594 LN from 103 CC patients (median 14 LN / patient, range 1-46) were analyzed with OSNA. From the 103 histology pN0 patients 26 were positive with OSNA, resulting in an upstaging rate of 25.2%. Sixteen patients had 1 OSNA positive LN, three had 2, three had 3, and one had 4 positive LNs, resulting in a total of 35 positive LN. Patients with pT1, pT2, and pT4 tumors only had 1 positive LN, while all patients with multiple positive LN were from the pT3 group which also contained 9/10 patients with a (++) OSNA result. Concerning stage categories, 6/37 (16.2%) CC patients with stage I were upstaged and 20/66 (30.3%) patients with stage II. Conclusions: OSNA is a novel molecular tool for LN upstaging in CC patients with similar performance as very intensive histological investigation (Güller U et al. CANCER: 2012 DOI 10.1002/cncr.27667). The present study confirms that OSNA improves staging in CC patients and provides a standardized, observer-independent technique for a more accurate staging than the standard histopathology.
A phase II trial of 5-fluorouracil, leucovorin, and oxaliplatin (mFOLFOX6) chemotherapy plus bevacizumab (bev) for patients (pts) with unresectable stage IV colon cancer and a synchronous asymptomatic primary tumor: Updated results of NSABP C-10 with definitive survival analysis.
Authors: McCahill Laurence E. , Yothers Greg , Sharif Saima , Petrelli Nicholas J. , Lopa Samia H. , O'Connell Michael J. , Wolmark Norman
Published: February 2017

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Background: Surgical resection of asymptomatic primary colon tumor for pts presenting with synchronous yet unresectable metastatic disease is controversial. We published results for the primary endpoint in JCO in September 2012. Here we update the primary endpoint and present definitive survival results. Methods: Eligible pts had ECOG Performance 0 or 1, an asymptomatic colon tumor and unresectable distant metastases. Primary endpoint (PE) was major morbidity, defined as surgical resection or death related to the intact primary tumor. Major morbidity rate of 25% was considered acceptable and the trial had 85% power to rule out a rate of 40%. Secondary endpoints were overall survival and morbidity related to the intact primary requiring hospitalization, transfusion, or interventional procedure. Results: Between March 2006-June 2009, 90 pts were registered and 86 eligible pts with follow-up comprise this analysis. 52% were female and 47% were age 60+. Median follow-up was 61.5 mos. There were 15 (17.4%) major morbidity events, 12 (14.0%) required surgery: obstruction (9), perforation (2), and pain (1). Three (3.5%) resulted in pt death: perforation (2), obstruction (1). Cumulative incidence of major morbidity at 42 mos was 17.7% (95% CI 9.5-26.0%). Fourteen other primary tumor resections were performed: attempted cure (11), other reasons (3). There were 6 secondary endpoint events: 4 obstructions (2 required stents, 2 resolved with conservative management), 2 pts with suspected perforation required percutaneous drainage (1) and hospitalization and antibiotics (1). Median survival was 18.3 mos (95% CI 15.2-24.2). Two year OS rate was 38.7% and 3 year OS rate was 15.1%. Conclusions: The primary endpoint holds with the updated analysis justifying observation for asymptomatic primary colon cancers even in patients with a good clinical response of distant metastases. Clinical trial information: NCT 00321828.
On-treatment progression-free survival analysis of aflibercept-FOLFIRI treatment within 28 days of progression in metastatic colorectal cancer: Updated efficacy results from the VELOUR study.
Authors: Ferry David Raymond , Kim Tae Won , Guren Tormod Kyrre , Desai Jayesh , Villanueva Luis Marcelo , Andria Michael L. , Van Cutsem Eric , Tabernero Josep
Published: February 2017

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Background: The phase III VELOUR study demonstrated that adding the novel antiangiogenic agent aflibercept (known as ziv-aflibercept in the United States) to FOLFIRI in patients with metastatic colorectal cancer previously treated with oxaliplatin significantly improved overall survival, progression-free survival (PFS), and overall response rate versus placebo-FOLFIRI. We performed an additional analysis of PFS “on-treatment,” censoring events that occurred more than 28 days after last treatment dose. Methods: Patients were randomized to receive aflibercept 4 mg/kg or placebo every 2 weeks in combination with FOLFIRI. An independent review committee determined progression based on radiologic review. PFS was estimated using Kaplan-Meier analysis, with censoring of events after the last dose plus 28 days. Treatment groups were compared using a log-rank test stratified by ECOG performance status and prior bevacizumab therapy. Hazard ratio (HR) and confidence interval (CI) were estimated using a Cox proportional hazard model. Results: On-treatment PFS results are shown in the Table. Patients on aflibercept-FOLFIRI showed significantly increased on-treatment PFS compared with patients on placebo-FOLFIRI. More patients were censored in the aflibercept arm due to adverse events, thus decreasing the number of events. Conclusions: On-treatment PFS with aflibercept-FOLFIRI was significantly increased compared with placebo-FOLFIRI, which is consistent with the PFS benefit observed in the primary analysis. Clinical trial information: NCT00561470. [Table: see text]