bluuebunny/crossref_metadata_2025 huggingface dataset:
Discovery of prognostic and predictive tissue biomarkers in patients with resectable esophageal cancer.
Authors: MacGregor Thomas P. , Gillies Richard S. , Sahgal Natasha , Chetty Runjan , Wang Lai-Mun , Turkington Richard Calvin , Maynard Nicholas D. , McHUGH Peter J. , Kennedy Richard D. , Middleton Mark R. , Sharma Ricky A.
Published: February 2017

45

Background: Patients with operable esophageal adenocarcinoma have a poor prognosis (median survival <2 years). We aimed to discover novel prognostic and predictive biomarkers to be validated as tools for patient selection for optimal neo-adjuvant therapy. Methods: Protein levels of XPF, MUS81, Cyclins A, B1, D1 and E, and Ki67 were assessed by retrospective immunohistochemical analysis of baseline tumor biopsy samples from 3 groups of patients with operable esophageal adenocarcinoma: surgery alone (N=54), 2 cycles of cisplatin-fluorouracil chemotherapy followed by surgery (N=46), and 2 cycles of oxaliplatin-fluorouracil chemotherapy (N=38). Expression of 48,803 genes was studied by Illumina HT-12 chip array followed by functional pathway analysis in oxaliplatin-treated patients before and after chemotherapy (N=38). Results were tested for association with pathological response (Mandard regression grading) (Chi-square test), disease free survival (DFS) and overall survival (OS) (Wilcoxon test). Results: High Ki67 protein levels were associated with worse OS (P=0.034; N=93). None of the markers were predictive of clinical endpoints following cisplatin chemotherapy. In oxaliplatin-treated patients (N=38), functional pathway analysis revealed associations between overexpression of cell cycle/DNA repair genes at baseline and worse clinical outcomes. Expression of 15 DNA repair (DNAR) genes was associated with DFS, and 16 DNAR genes with OS. Expression of 21 DNAR genes significantly increased after chemotherapy. Gene expression associations were validated at the protein level: high MUS81 at baseline predicted poor DFS (P=0.036) and poor OS (P=0.015) following oxaliplatin therapy; high XPF expression was associated with lack of pathological response (P=0.032); high Cyclin B1 predicted poor DFS (P=0.017). XPF protein levels increased following oxaliplatin (P=0.001, paired t-test). Conclusions: By confirmation of mRNA findings at the protein level, XPF, MUS81, and Cyclin B1 have been discovered as predictive biomarkers for response to oxaliplatin chemotherapy that merit prospective validation as tools for patient selection. Funded by Oxford NIHR Biomedical Research Centre and ECMC.
Effect of RAD21 overexpression on prognosis in KRAS-mutant colorectal carcinomas.
Authors: De Campos Silva Thomaz , Deb Siddhartha , Xu Huiling , Yan Yuqian , Fox Stephen B. , Mortensen Neil , McKay Michael
Published: February 2017

453

Background: RAD21 is a component of the cohesion complex and is integral for chromosome segregation and error-free DNA repair. Early studies suggest RAD21 appears to be functionally important in tumour progression and we assessed its clinicopathological and prognostic significance in a cohort of colorectal adenocarcinoma (CRC), and its biological characteristics and cell cell chemosensitivity in vitro. Methods: A multi-institutional study examined RAD21 expression in 652 invasive CRC's. Correlation with clinicopathological factors including gender, differentiation, subtype, nodal status, metastasis and disease specific survival was performed. Correlation with BRAF and KRAS mutation status, p53 accumulation, microsatellite instability, and CpG island methylator phenotype was also undertaken. Cell clones with stable RAD21 knockdown were generated and tested for cellular sensitivity to conventional chemotherapeutic drugs. Results: High RAD21 expression correlated with male gender (56.7% vs. 43.3%), more well differentiated tumours (14.4% vs. 4.0%, p = 0.0001), higher T stage (36.1% vs. 27.0%, p = 0.01), and presence of metastasis (18.8% vs. 12.6%, p =0.03). Both on univariate and multivariate analysis, increased RAD21 expression correlated with worse DSS (hazards ration [HR] 1.4, 95% confidence interval [CI] 1.1 to 1.9, p = 0.01). Subgroup analysis showed RAD21 expression was prognostic in female patients (HR:1.6, 95%CI:1.1-2.3, p = 0.01) and tumours with nodal disease (HR:1.6, 95%CI:1.2-2.2, p = 0.003). RAD21 expression associates worse DSS in patients whose tumors carrying KRAS mutations (HR:2.6, 95%CI:1.4-4.3, p = 0.001), or p53 accumulation (HR:2.2, 95%CI:1.2-4.1, p = 0.02) and in patients receiving adjuvant chemoradiotherapy (HR:1.9, 95%CI:1.2-3.0, p = 0.005). Cells with reduced RAD21 expression exhibited enhanced sensitivity to 5-FU, alone or with Oxaliplatin. Conclusions: RAD21 expression in CRC is associated to advanced disease stage and confers poorer overall prognosis and resistance to chemoradiotherapy. RAD21 expression is of prognostic significance in tumours with somatic KRAS mutations or p53 accumulation. In these subgroups, RAD21 may be a novel important therapeutic target.
Associations between patient (pt) colorectal cancer (CRC) tumor KRAS and BRAF mutation (mut) status and overall survival (OS).
Authors: Hassabo Hesham M. , Sahin Ibrahim Halil , Kazmi Syed Mohammad Ali , Al Mutar Salwan S. , Bugano Diniz Gomes Diogo , Pini Tunghi May , Vilar Sanchez Eduardo , Dasari Arvind , Kopetz Scott , Overman Michael J. , Eng Cathy , Kee Bryan K , Vauthey Jean-Nicolas , Hassan Manal , Garrett Chris R.
Published: February 2017

473

Background: Evolving information is emerging regarding patient tumor molecular data and associations with patient clinical characteristics and survival outcomes; we sought to evaluate pt clinical data with CRC tumor mut status. Methods: Since January 2008 tumor pts with stage IV CRC (principally primary tumors) were analyzed for KRAS (codon 12, 13, and 61) mut status as standard-of-care. A subset of pts tumors underwent BRAF and KRAS codon 164 mut testing. Results: 1,891 pts with CRC whose tumors underwent molecular mut analysis were identified from the clinical record. There was no statistically significant difference in pt overall survival from time of diagnosis for stages I/II (data not shown) and III; for stage IV pts there was an inferior OS associated with BRAF mut tumors (see Table). Tumor BRAF mut was significantly detected among ever smokers (10.2%) as compared to never smokers (5.4%), p < 0.001; meanwhile, tumor KRAS mut and wild type distribution were similar between ever smokers and never smokers. KRAS mut was significantly observed in CRC patients with distant lymph node, hepatic and lung metastasis, p < 0.001. Type 2 diabetes, intake of metformin or ACE inhibitors, were not significantly correlated with tumor KRAS or BRAF mut status. Conclusions: Pts with stage IV CRC BRAF mut tumors have a higher likelihood of prior or current smoking history in addition to having an adverse survival prognosis; pts with stage IV KRAS mut tumors have an increased association with distant lymph node, hepatic, and lung metastases as compared to KRAS wild type and BRAF mut tumors. Studies are ongoing to determine how tumor molecular characteristics may help to predict clinical outcomes and provide a useful to guide pt care. [Table: see text]
Association of human papillomavirus with unique clinicopathologic features in patients with metastatic squamous cell carcinoma of the anal canal.
Authors: Morris Van Karlyle , Rashid Asif , Rodriguez-Bigas Miguel A. , Das Prajnan , Wolff Robert A. , Chang George J. , Crane Christopher H. , Phillips Jonathan , Ohinata Aki , Rogers Jane , Eng Cathy
Published: February 2017

479

Background: The incidence of anal carcinoma in the United States continues to increase steadily, and infection with the human papillomavirus (HPV) is an established risk factor for the development of anal carcinoma. However, clinicopathologic characteristics of patients with metastatic squamous cell carcinoma of the anal canal according to HPV status have thus far not been defined. Methods: Records of patients treated for metastatic squamous cell carcinoma of the anal canal at the MD Anderson Cancer Center between June 2005 and August 2013 were reviewed. Patients were tested for the presence of HPV DNA by in-situ hybridization and/or the p16 oncoprotein by immunohistochemistry. Associations between the presence of HPV and clinicopathologic attributes were measured by calculation of odds ratios, and survival was estimated according to the Kaplan-Meier method. Results: Patients were followed for a mean of 50.2 months, and the mean age at diagnosis was 54.0 years. Among the 43 patient records reviewed, 39 tumors (90.1%) tested positive for the presence of HPV. Patients with HPV-negative tumors were more likely to have a strong (> 30 pack-year) tobacco history (OR=18.5, p=0.018) and were more likely to develop brain metastases (OR=2.0, p=0.04). A faint association was observed between Caucasian ethnicity and HPV-positive tumors (OR=8.8, p=0.06). Median survival from the time of diagnosis of metastatic disease was 42.4 months, with a trend towards a worse survival seen among patients with HPV-negative tumors (HR for death 4.9, p=0.09). Conclusions: Differences in HPV status and prior tobacco exposure may identify two separate populations of patients with metastatic squamous cell anal carcinoma with different clinicopathologic phenotypes. Our results are similar to prior published data in patients with squamous cell carcinoma of the head and neck in which patients with HPV-positive tumors demonstrate improved survival outcomes. Data from additional patients will be presented.
Potential role of mTOR phosphorylation status as a negative predictor to everolimus plus octreotide in NETs.
Authors: Casanovas Oriol , Capdevila Jaume , Barriuso Jorge , Teule Alex , Castellano Daniel E. , Manzano Jose Luis , Lopez Carlos , Alonso Vicente , Garcia Carbonero Rocio , Dotor Emma , Martinez Alba , Salazar Ramon
Published: February 2017

484

Background: Everolimus treatment is currently approved for a few tumor types and currently evaluated in several other malignancies, including NETs. Nevertheless, there is an urgent need for histological or molecular factors of prediction of response, as these would help discriminate subgroups of patients more likely to respond. Here we report on a biological sub-study on histological markers to unravel the relationship between the activation of the IGFR-mTOR-S6 signal transduction pathway with treatment response to everolimus plus octreotide combined therapy in patients with GI NETs. Methods: IGFR-1 (Abcam #54274), phospho-mTOR (Ser-2448, Cell Signaling #2976) and phospho-S6 (Ser-235/236, Cell Signaling #4857) were evaluated by immunohistochemistry (according to Choe G. et al, Cancer Research. 2003) from paraffin embedded blocks from patients with progressive locally advanced or metastatic non-functioning GI NETs, included in a national phase II clinical trial (EVERLAR, NCT01567488). Study endpoint was PFS, and 50% of the patients had progressed at the time of analysis. Univariant (Chi square) and multi-variant (COX) statistical analysis were performed. Results: 38 patients were evaluated with a median age of 62.7 years, balanced on gender (57 % vs. 42%), of which 76.3% were metastatic, mostly to liver. IGF1R positivity showed a nonsignificant trend to association to treatment response (50% progression vs. 21% progression, p=0.15). Nevertheless, mTOR activation (phospho-mTOR positivity) was associated with progression (median PFS of 100 days vs. 225 days, p=0.019). Multivariant COX regression analysis showed a Hazard Ratio of 2.96 [0.8-10.2] (p=0.087) for phospho-mTOR activation. Conclusions: Contrarily to expected results, mTOR activation status determined by phospho-mTOR immunohistological positivity seems to be associated to worse outcome in terms of progression to everolimus+somatostatin analogs in GI NETs patients. This could be indicative of a refractoriness or primary resistance to treatment based on a threshold of mTOR activation, possibly due to activation feedback-loops.
Influence of genetic variants of genes potentially associated with colorectal brain metastases on overall survival.
Authors: Stremitzer Stefan , Berghoff Anna Sophie , Volz Nico Benjamin , Zhang Wu , Yang Dongyun , Stintzing Sebastian , Ning Yan , Sunakawa Yu , Yamauchi Shinichi , Sebio Ana , El-Khoueiry Rita Elie , Birner Peter , Preusser Matthias , Lenz Heinz-Josef
Published: February 2017

487

Background: Brain metastases (BM) in colorectal cancer (CRC) are rare, developing in only 0.3-9% of the patients, and considered a late-stage manifestation of the disease. The aim of this study was to investigate whether genetic variants of genes involved in BM-related pathways, such as integrin, invasion- and adhesion-mediating, angiogenic and tumor suppressing pathways, are associated with outcome. Methods: Genomic DNA was extracted from formalin-fixed paraffin embedded resected BM from 70 patients with histologically proven CRC. Single nucleotide polymorphisms (SNP) in seven genes (CXCR4, MMP9, ST6GALNAC5, ITGAV, ITGB1, ITGB3, KLF4) were analyzed by direct Sanger DNA sequencing and evaluated for association with overall survival (OS) from resection of BM. Only SNPs with an allele frequency of ≥ 10% were analyzed. Results: In univariate analysis, rs17577 (MMP9) and rs4642 (ITGB3) showed a significant difference in OS [(G/G 7.4 months, G/A 5.1 months; HR (95% CI) 1.83 (0.95-3.53), p = 0.0440) and (A/A 9.4 months, A/G 4.8 months, G/G 4.3 months; HR (95% CI) 0.81 (0.44-1.49) and 2.14 (0.98-4.67), p = 0.0354), respectively]. In multivariate analysis adjusted for baseline characteristics [primary tumor site (right colon, left colon, rectal), chemotherapy before BM (yes/no), BM location (supratentorial, infratentorial, both), Karnofsky performance status (<80, 80-100)], rs2236599 (KLF4), and rs10171481 (ITGAV) are significant in OS [(G/G 7.4 months, G/A or A/A 4.8 months; HR (95% CI) 3.19 (1.55-6.53), p = 0.0016) and (A/A 5.7 months, A/G 4.4 months, G/G 15.5 months; HR (95% CI) 0.61 (0.29-1.29) and 0.25 (0.10-0.60), p = 0.0082), respectively]. Conclusions: This study suggests for the first time a prognostic effect of the SNPs involved in the BM pathway. Further analyses are needed to confirm these findings.
Oral nelfinavir before and during radiation therapy for rectal cancer: Changes in tumor perfusion and correlation between tissue and radiological markers of response.
Authors: Hill Esme J. , Enescu Monica , West Nicholas , Franklin Jamie M. , Chu Kwun-Ye , Li Jun , Whittington Deborah , Macgregor Thomas P. , Ashraf Shazad , Blesing Claire , Mukherjee Somnath , Betts Margaret , Slater Andrew , Anderson Ewan M. , Brown Gina , Schnabel Julia A. , Partridge Mike , Quirke Philip , Sharma Ricky A.
Published: February 2017

491

Background: Nelfinavir, an inhibitor of AKT in the PI3-kinase pathway, is a radiosensitizer which increases tumor blood flow and reduces hypoxia in preclinical models. We conducted a pilot study of nelfinavir before and during hypofractionated pelvic radiotherapy (RT) in patients with rectal cancer to evaluate the safety of the combination, changes in tumor perfusion and a tissue biomarker of radiosensitization, tumor cell density (TCD). Methods: Eight patients with T3-4 N1-2 M1 rectal cancer received 7 days of oral nelfinavir (1250 mg bd) and a further 7 days of nelfinavir during pelvic RT (25 Gy/5 fractions/7 days). Safety was evaluated by CTCAE v 4.0. Dynamic contrast enhanced (DCE) CT and MRI scans were performed pre-treatment, after 7 days nelfinavir and prior to last fraction of RT. TCD was performed in biopsies taken pre-treatment and 7 days post-radiotherapy (West N et al. Br J Cancer. 2010, 102; 1519-23). Radiological response and tumor regression grade (TRG) was assessed on MRI 8 weeks from end of RT (Patel UB et al. J Clin Oncol. 2011, 29:3753-60). Results: This was the first clinical trial to combine nelfinavir with hypofractionated RT. There were 3 grade 3 adverse events (diarrhea, rash, lymphopenia). After 7 days of nelfinavir, there was a 40% increase in mean Ktrans on DCE-MRI (p = 0.029, paired t test). On DCE-CT, there was a 30% increase in mean blood flow from before RT to the last day of RT (p = 0.028). The median TCD in biopsies decreased from 41.2% (IQR = 23.5, 53.0%) at baseline to 9.2% (IQR = 2.8, 18.2%) after RT (p = 0.007, paired t test). By RECIST criteria, 3/8 patients treated had complete radiological responses and 4/8 had partial responses. TRG was associated with the magnitude of decrease in TCD measured in paired biopsies for individual patients (p = 0.029, Fisher’s Exact Test). Conclusions: These data suggest that the radiosensitizing drug, nelfinavir, can increase blood flow to rectal cancer when given prior to RT. Nelfinavir added to hypofractionated pelvic radiotherapy has acceptable toxicity and is associated with radiological and tissue biomarker responses. Funded by Oxford NIHR BRC, OCIC, Yorkshire Cancer Research, Oxford and Leeds ECMCs. Clinical trial information: 2010-020621-40.
The impact of low-frequency KRAS mutations on the efficacy of anti-EGFR antibody for patients with metastatic colorectal cancer.
Authors: Hikosaka Tomomi , Yamazaki Kentaro , Hamauchi Satoshi , Tsushima Takahiro , Yokota Tomoya , Todaka Akiko , Machida Nozomu , Fukutomi Akira , Onozawa Yusuke , Yasui Hirofumi
Published: February 2017

496

Background: The benefit of anti-EGFR antibody is limited to metastatic colorectal cancer with KRAS wild-type (WT). However, some KRAS WT patients have low-frequency KRAS mutation (low MT) by pyrosequencing. Recently, it is reported that the efficacy of anti-EGFR antibody on these patients was inferior to no KRAS mutation patients (no MT) (D. Tougeron et al, Ann Oncol. 2013). Methods: Patients were categorized as low MT or no MT according to pyrosequencing cut-off point, and we evaluated anticancer effect of anti-EGFR antibody in each group. Results: A total of 358 patients were analyzed KRAS genotype by pyrosequencing between April 2010 and February 2013 in Shizuoka Cancer Center. 217 (60.6%) patients were KRAS WT of the total, and 93 (42.9%) were low MT, while 124 (57.1%) were no MT among them. 51 of low MT and 53 of no MT patients were treated by anti-EGFR antibody therapy, and response rate (RR) were 41% (CR/PR/SD/PD 1/20/17/13) and 32% (CR/PR/SD/PD 0/17/21/15), median progression-free survival (PFS) were 164 days and 170 days, respectively. 15 of low MT and 11 of no MT patients received anti-EGFR antibody monotherapy, and RR were 33% (CR/PR/SD/PD 0/5/7/3) and 19% (CR/PR/SD/PD 0/1/4/9), PFS were 184 days and 91 days, respectively. 27 patients in both groups were refractory or intolerance to 5-FU/L-OHP/CPT-11, and RR were 37% (CR/PR/SD/PD 0/10/9/8) and 19% (CR/PR/SD/PD 0/5/13/9), PFS were 132 days and 140 days, respectively. In univariate and multivariate analysis, there was no difference of anticancer effect between low MT and no MT. Conclusions: Unlike the previous report, patients with low-frequency KRAS mutations also benefited from anti-EGFR antibody therapy. However, some other biomarkers such as NRAS, BRAF, PIK3CA might affect these results. Now we are evaluating the effect of these candidate biomarkers on these results, and we will report the update data in this meeting.
GPX3 as a predictive biomarker in gastrointestinal malignancies.
Authors: Pelosof Lorraine Cheryl , Yerram Sashidhar R. , Yanagisawa Breann , Azad Nilofer Saba , Herman James Gordon
Published: February 2017

499

Background: Epigenetic heterogeneity within colorectal and pancreatic tumors influences their sensitivity to individual chemotherapy agents. Glutathione peroxidase 3 (GPX3), part of the cellular antioxidant system, is associated with platinum resistance in multiple tumor types. We explored the role of GPX3 methylation in colorectal (CRC) and pancreatic tumor cells’ sensitivity to platinum agents. Methods: Methylation-specific PCR (MSP) and quantitative real-time polymerase chain reaction (qPCR) were used to examine the promoter methylation status and expression level, respectively, of GPX3 in primary tumor xenografts and cell lines. Cell proliferation assays were used to measure cells’ sensitivity to cisplatin and oxaliplatin and short hairpin RNA (shRNA) was used to silence GPX3 in these cells. Results: We examined GPX3 promoter hypermethylation in colorectal and pancreatic primary tumor xenografts. GPX3 was methylated in 25% of colorectal cancer primary tumor xenografts tested and 50% of pancreatic cancer primary tumor xenografts tested; methylation correlated with decreased/absent expression in these samples. Additionally, GPX3 is methylated in 6 out of 9 colorectal cancer cell lines tested and cell lines with methylated GPX3 had significantly decreased GPX3 expression. Importantly, the cisplatin IC50 for the cell lines with methylated GPX3 was 4-fold lower (median 4.5 mM) compared to those lines with unmethylated GPX3 (median 20 mM) and oxaliplatin demonstrates a similar trend with the IC50 of the highly GPX3-expressing line CaCO2 being approximately two-fold that of the fully-methylated line HCT116. Underlying the importance of methylation, azacitidine treatment resulted in GPX3 re-expression in HCT116 cells with sensitivity to oxaliplatin decreasing as would be expected. Currently, we are using shRNA to silence GPX3 in CRC lines to directly test its role in platinum sensitivity; preliminary results demonstrate an approximately two-fold decrease in oxaliplatin IC50 when GPX3 expression is decreased in GPX3-expressing cells. Conclusions: Epigenetically silenced GPX3 may serve as a predictive biomarker for platinum sensitivity in CRC and pancreatic tumor cells.
Use of genetic variants in c-MET to predict clinical outcome in localized gastric cancer (GC) patients (pts) treated with surgery.
Authors: Sunakawa Yu , Wakatsuki Takeru , Zhang Wu , Yang Dongyun , Azuma Mizutomo , Ning Yan , Stintzing Sebastian , Stremitzer Stefan , Yamauchi Shinichi , Sebio Ana , Volz Nico Benjamin , El-khoueiry Rita , Koizumi Wasaburo , Watanabe Masahiko , Lenz Heinz-Josef
Published: February 2017

50

Background: Dysregulation of the c-MET signaling pathway occurs in a wide range of human cancers including GC. Such derangements result from various molecular mechanisms, including mutations, amplification and overexpression of c-MET. Overexpression and amplification of c-MET have been correlated with poor clinical outcomes in pts with GC. However, the association between c-MET polymorphisms and prognosis in GC is not well defined. We examined the prognostic impact of c-MET polymorphisms on clinical outcomes in pts with localized GC treated with surgery. Methods: One-hundred and sixty-one Japanese pts were included, with localized GC (stage Ib-IV) treated with surgery alone (n=58), or surgery plus adjuvant therapy (n=103) between 2002 and 2010. Median follow up was 4 years. Genomic DNA was extracted from the pts’ blood or tissue. Six functionally significant c-MET SNPs (rs1621, rs40239, rs41736, rs41739, rs184953, rs10234854) were analyzed by PCR-based direct sequencing. All candidate SNPs were analyzed for association with disease free survival (DFS) and overall survival (OS) by uni- and multivariate analyses, adjusting for age, sex, stage and type of adjuvant therapy. Results: The MET rs40239 variant was associated with favorable clinical outcomes. Univariate analysis showed pts with any G (AG/GG) allele had significantly longer DFS and OS compared to those with the AA genotype (HR: 0.43; 95% CI: 0.25-0.74; P=0.001, HR: 0.47; 95% CI: 0.27-0.81; P=0.006, log-rank test, respectively); this remained significant upon multivariate analysis (HR: 0.48; 95% CI: 0.27-0.83; P=0.009, HR: 0.50; 95% CI: 0.28-0.88; P=0.017, respectively). In the subgroup analysis by gender, men, but not women, with the AG/GG genotypes maintained a DFS and OS benefit. Conclusions: Our results show for the first time that the c-MET polymorphism, rs40239, may serve as a prognostic marker in pts with localized GC treated with surgery. There also appears to be a gender-related difference in the impact on clinical outcome by genetic variants of c-MET. Prospective validation of this study is warranted.
Ultra-high-quality sequencing assay for comprehensive genetic panel analysis of tumor-derived circulating cell-free DNA in colorectal cancer patients.
Authors: Lee Jeeyun , Mortimer Stefanie , Mei Gangwu , Sebisanovic Dragan , Siew LaiMun , Eltoukhy Helmy , Talasaz AmirAli
Published: February 2017

504

Background: Current approaches based on invasive biopsy genetic analysis can fail to capture an accurate picture of the real-time cancer profile due to limited spatial window of biopsy into residual disease throughout the body. Moreover, tumor rebiopsy has significant challenges to be widely used in practice for serial monitoring of disease progression or acquired resistance. Analysis of circulating tumor nucleic acids (ctDNA), on the other hand, presents a new tool for the monitoring and treatment of cancer. However, due to high-quality false positives in current NGS assays, the majority of studies on ctDNA have been limited to hotspot analyses, and typically only involve patients where ctDNA fractions are high (>1-5%). Methods: We have developed a differentiated sequencing assay, Digital Sequencing Technology (DST) that enables detection of rare genomic abnormalities with ultra high-specificity and sensitivity. Our assay is able to eliminate the error and distortion created by sample-prep and sequencing processes in standard NGS workflows and produce near-perfect representations of all rare variants. Results: We have shown that in sequencing a comprehensive cancer panel of 80kbp in 0.1% cancer cell line titration samples, standard Illumina SBS generates many high-quality false positive variant calls in the range of 0.05-5%, while our assay resulted in highly sensitive and completely error-free variant calls across the entire panel. We then profiled ctDNA and matched primary tumor samples in more than 50 plasma samples collected from metastatic colorectal cancer patients. We found mutations in ctDNA with allele frequencies in the range of 0.05-40%. We established the concordance of ctDNA mutations with different clones within primary tumors. We further investigated the potential clinical usefulness of discordant mutants based on the treatment history of each patient. Conclusions: This work indicates the remarkable potential of using our assay in deep analysis of ctDNA, thereby allowing researchers and clinicians to comprehensively and non-invasively monitor the genetic dimension of cancer throughout the body.
Identification of hypoxanthine and phosphoenolpyruvic acid as serum markers associated with chemoradiotherapy response in locally advanced rectal cancer.
Authors: Kim Min-Jeong , Yoo Byong Chul , Yeo Seung-Gu
Published: February 2017

507

Background: Patients show variable responses to chemoradiotherapy (CRT), generally performed before surgery for locally advanced rectal cancer (LARC). We aimed to identify molecular markers predictive of CRT responses by analyzing low-mass ions (LMIs) in serum of LARC patients. Methods: LMIs (<1000 m/z) in serum obtained before CRT from 73 LARC (cT3–4) patients were profiled using matrix-assisted laser desorption/ionization mass spectrometry. LMIs with higher weighting factors in discriminating CRT responses were selected using principal components analysis and discriminant analysis. Selected LMIs were identified using the Human Metabolome Database. The concentrations of identified LMIs were determined by colorimetric enzyme assay, and compared according to post-CRT pathological stage (ypStage) or Dworak’s tumor regression grade (TRG). Results: The nine highest-ranking LMIs were selected. Among them, two LMIs with 137.08 and 169.04 m/z were identified as hypoxanthine (HX) and phosphoenolpyruvic acid (PEP), respectively. The HX concentration was higher in patients with ypStage 0-I compared with ypStage II-IV (p = 0.034) or ypStage III-IV (p = 0.030); a similar difference was observed between TRG IV-III and TRG 1 (p = 0.035). In contrast, the concentration of PEP was significantly lower in TRG 4-3 compared to TRG 2-1 (p = 0.012), and a lower PEP level was also found in ypStage 0 compared to ypStage I-IV, with marginal significance (p = 0.061). Conclusions: This study demonstrated that pretreatment serum concentrations of HX and PEP, identified using LMI profiling, may be useful for predicting the CRT response of LARC.
Impact of KRAS on outcomes of patients with unresectable colorectal liver metastases (CRLM).
Authors: Ang Celina , Gewirtz Alexandra N. , Chou Joanne F. , Capanu Marinela , Vakiani Efsevia , Kemeny Nancy E.
Published: February 2017

510

Background: In patients with resected CRLM, mutant (MUT) KRAS may confer a worse prognosis than wild-type (WT) KRAS (Karagkounis et al., ASCO 2012, abs 3616; Kemeny et al., ASCO 2013, abs 3609). We analyzed the impact of KRAS on the outcomes of patients with unresectable CRLM treated with hepatic arterial infusion (HAI) + systemic chemotherapy. Methods: Median overall survival (OS) and progression-free survival (PFS) were estimated by Kaplan Meier methods. The log rank test was used to examine the association of patient demographics and disease-related factors including KRAS mutation status with OS and PFS. Cox proportion hazards model was used to examine the association of liver resection with survival outcomes by treating liver resection as a time dependent covariate. Results: Of 93 patients: 59 male, 34 female; 77 Caucasian, 16 non-Caucasian; 66 were age < 60 years, 27 were age > 60 years; 70 were KRAS WT and 23 were KRAS MUT; 59 had chemotherapy pre-HAI pump placement and 53 subsequently underwent liver resection. Median PFS and OS were 13.3 months and 45.7 months, respectively. Age, sex and race were not associated with PFS or OS. Univariate analysis of the impact of KRAS, pre-HAI chemotherapy, and subsequent liver resection on PFS and OS is presented in the Table. Multivariate analysis including important variables did not change outcome. Conclusions: In patients with unresectable CRLM treated with HAI and systemic chemotherapy, overall survival tended to be improved in KRAS WT vs MUT patients. [Table: see text]
Planned interim analysis of phase II study of sorafenib and capecitabine (SorCape) in previously treated metastatic colorectal cancer (mCRC): NCT01471353.
Authors: George Thomas J. , Ivey Alison Marguerite , Daily Karen Colleen , Dang Long H. , Watson Scott , Granicz Renee , McEwan Margaret A. , Scian Joanna , Lu Xiaomin , Iqbal Atif , Tan Sanda , Zendejas Ivan , Allegra Carmen Joseph
Published: February 2017

548

Background: Inhibition of the MEK/ERK pathway through Raf has been shown to resensitize tumor cells resistant to chemotherapy. Sorafenib (Sor) is an oral multi-tyrosine kinase inhibitor (TKI) affecting tumor proliferation and angiogenesis through inhibition of the Ras/Raf/MEK/ERK, VEGF, and PDGF pathways. A similar TKI, regorafenib, has efficacy as monotherapy in advanced mCRC. Demonstrating activity of this compound class with standard cytotoxic therapy represents a clinical unmet need. Methods: Eligible pts had progressive mCRC after prior oxaliplatin and irinotecan, at least 1 measurable target lesion by RECIST, ECOG PS 0-1, and adequate organ function. Pts received Cape (1000mg/m2 PO BID D1-14) and Sor (400mg PO BID max via dose escalation per cycle). Cycles were q21d with tumor assessments q9w. Endpoints included progression-free survival (PFS), objective and biochemical response rates, overall survival (OS), toxicity and quality of life. Sample size was based on a 50% improved median PFS from 3 (historical) to 4.5 months (m). Results: 29 of 43 planned pts have received therapy and the first 26 are included in this planned interim analysis. Baseline pt demographics are in the Table. Most frequently observed adverse events (any AE) included HTN (15%), diarrhea (35%), fatigue (65%) and HFS (91%). 19 pts (73%) had a grade 3 AE (12/19 were HFS). There were no grade 4 or 5 toxicities. 11/25 (44%) evaluable pts have radiographic stable disease. With a median f/u of 6.2 mo, interim PFS is 4.4m. Conclusions: Enrollment continues with manageable AEs, despite the overlapping toxicity of HFS. Planned interim PFS data appears promising in this refractory mCRC population. Molecular correlatives for predictors of response are being analyzed. Clinical trial information: NCT01471353. [Table: see text]
A phase I study of etirinotecan pegol in combination with 5-fluorouracil and leucovorin in patients with advanced cancer.
Authors: Krishnamurthi Smitha S. , Chadha Manpreet , Rodal Mary Beth , Weiss Glen J. , Bokar Joseph A. , Jameson Gayle S. , Mast Catherine Patricia , Zhao Carol , Waluch Bridget , Hoch Ute , Hannah Alison , Dowlati Afshin , Meropol Neal J. , Ramanathan Ramesh K.
Published: February 2017

550

Background: Etirinotecan pegol (EP; NKTR-102) is the first long-acting topoisomerase I inhibitor designed to concentrate and provide continuous tumor exposure. This study was designed to identify the recommended dose of EP in combination with 5-fluorouracil (5FU; 400/2400 mg/m2 by bolus, then 46-hour infusion) and leucovorin (LV; 400 mg/m2). Methods: Patients (pts) were enrolled in a standard 3+3 design. Initially, pts received all 3 agents every 2 weeks (schedule A). Due to higher SN38 exposure with the triplet compared to single-agent EP in the 25 mg/m2dose cohort, the schedule was modified to deliver all 3 agents for the first 4 doses at 2-week intervals; 5FU/LV continued at 2-week and EP at 4-week intervals thereafter (Schedule B). The protocol enrolled adults with an advanced malignancy; pts had ECOG 0 or 1 with adequate organ function. Results: 26 pts were enrolled in 5 cohorts: median age 63 (44-80); male:female 12:14; median prior regimens 3.5 (1-9). Schedule A: 25 mg/m2 (n=8, 6 evaluable for toxicity) and 50 mg/m2 (n=3). Schedule B: 50 mg/m2 (n=3), 75 mg/m2 (n=6) and 100 mg/m2 (n=6). DLTs: Grade (G) 3 abdominal pain following the infusion (1 pt; 25 mg/m2); G3 nausea/dehydration and G4 neutropenia (NTP) >3 days (2 pts; 100 mg/m2). 5 pts at a dose of 75 mg/m2 had a dose delay due to NTP or diarrhea in the first 2 months. Common G3+ toxicity at 75 mg/m2: NTP (50%), diarrhea and lymphopenia (both 33%). At >50 mg/m2EP, SN38 exposure was comparable to that previously observed after single-agent EP. 5-FU exposure: comparable to prior reports. Two pts (ovarian, gall bladder) had a PR; 11 additional pts had stable disease >6months (3 NSCL 6M-9M; 3 CRC 6-8M; esophageal 7M, desmoplastic round cell sarcoma and pheochromocytoma 6M) or tumor marker response (pancreatic pt who had received prior irinotecan 93%, 9M; breast 82%, 5M). ORR or SD>6M was observed in 8 of the 12 patients in the two highest dose levels. Conclusions: 75 mg/m2 EP with 5FU/LV (Schedule B) shows promising clinical activity with an acceptable safety profile. The combination warrants further clinical testing. Single-agent EP is currently being studied in a phase 3 trial in breast cancer compared to physician’s choice (n=852).
The role of adjuvant radiation therapy in T4 nonrectal colon cancers.
Authors: Pail Orrin , Spieler Benjamin , Samueli Benzion , Shacham-Shmueli Einat , Gutman Mordechai , Zmora Oded , Venturero Moris , Golan Talia , Goldstein Jeffrey , Symon Zvi , Lawrence Yaacov Richard
Published: February 2017

553

Background: The standard treatment for non-metastatic locally advanced (T4M0) non-rectal colon cancer is surgical resection followed by adjuvant chemotherapy. There is strong rational for the use of post-operative radiation therapy (RT) in T4 tumors since negative margins are difficult to obtain. Despite promising single institutional studies, a phase III clinical trial closed early due to poor accrual. The efficacy of adjuvant RT in these patients remains unknown. We hypothesized that patients at lower risk of micro-metastases would benefit most from aggressive local treatment. Methods: Cases were extracted from the Surveillance, Epidemiology, and End Results (SEER) database. Inclusion criteria: T4-anyN-M0 non-rectal colon cancers, diagnosed between 1988 and 2010. Exclusion criteria: rectal tumors and cases lacking staging information. Endpoints: overall survival (OS) and cancer-specific survival. Analyses were based on the Kaplan-Meier method and Cox proportional hazard model. Results: A total of 36,260 patients with T4 colon cancer were identified. Due to changes in the use of systemic adjuvant therapies, an initial analysis was performed to determine effect of adjuvant RT by year of diagnosis. Prior to 2003, adjuvant RT provided no significant improvement in OS. However, between 2003 and 2010, adjuvant RT improved OS by 8 months, from 48 to 56 months (HR: 0.83, p = 0.001). Within this latter period, adjuvant RT improved OS in three subsets of the population: in patients without nodal involvement (N0) (HR: 0.79, p = 0.009); in patients with primary tumor in the sigmoid colon (HR: 0.79, p=0.007); in patients with primary tumor size of 6cm or greater (HR: 0.74, p < 0.001). On multivariate analysis, the following categories remained significant: N stage, tumor grade, year of diagnosis, gender, primary site, and radiation therapy. Conclusions: Some patients with T4 non-rectal colon cancer appear to benefit from adjuvant RT after surgical resection of the primary tumor, especially node-negative tumors located within the sigmoid colon, larger than 6cm in diameter. This benefit has become more evident since the introduction of more effective adjuvant chemotherapy. Our conclusions require verification in a prospective study.
A prognostic nomogram for predicting recurrence and survival in rectal cancers without neoadjuvant treatment.
Authors: Peng Junjie , Ding Ying , Wu Hongbin , Cai Sanjun
Published: February 2017

555

Background: To develop a prognostic nomogram for predicting recurrence and survival in patients with locally advanced rectal cancers that were not able to receive preoperative treatment. Methods: A total of 887 patients with AJCC (7th edition) stage II-III rectal cancers were retrospectively collected from a single institution. All patients did not receive any preoperative treatment before curative resection of the primary tumor. Cox proportional hazards model was performed to develop the predictive model for OS. Results: The 5-year local recurrence rate, distant metastases rate, and overall survival rate were 22.2%, 32.9%, and 63.9%, respectively. A prognostic nomogram was successfully developed to predicting patients’ 5-year overall survival, with a concordance index of 0.7. Factors in this model included patients’ age, gender, CEA value, tumor location, T stage, N stage, metastatic lymph nodes ratio, lymphvascular invasion, perineural invasion, adjuvant chemotherapy, adjuvant chemoradiotherapy. Our nomogram improved the predicting accuracy of current AJCC stage system (7th edition). Conclusions: The prognostic nomogram integrated patients demographic and clinicopathologic factors, which is better able to account for tumor and patient heterogeneity. Our individualized prediction nomogram could help physicians counsel and advise patients about their personalized treatment strategies and follow-up protocols. [Table: see text]
Ziv-aflibercept (Z) in combination with FOLFIRI for second-line treatment of patients (pts) with metastatic colorectal cancer (mCRC): Interim safety data from the global aflibercept safety and quality-of-life program in pts pretreated with bevacizumab (B).
Authors: Ferry David Raymond , Sobrero Alberto F. , Bordonaro Roberto , Siena Salvatore , Pietrantonio Filippo , Bauer Stefan , Taïeb Julien , Garreau-Laporte Pascale , Brette Sandrine , Dochy Emmanuelle , Lledo Gérard
Published: February 2017

556

Background: In the VELOUR study, adding Z (known as aflibercept outside the United States) to FOLFIRI resulted in improved OS, PFS, and RR in mCRC pts who had received prior oxaliplatin. Prior treatment with B (~30% of the ITT population) did not appear to impact the safety profile of Z. Results from VELOUR supported initiation of the global Aflibercept Safety and Quality-of-Life (QoL) Program composed of two clinical studies (ASQoP [NCT01571284]; AFEQT [NCT01670721]) to capture QoL and safety data from a population similar to that of VELOUR in a real-life setting. We report early safety data from this interim analysis in pts pretreated with B. Methods: ASQoP and AFEQT are single-arm, open-label trials evaluating safety and QoL of Z in mCRC pts previously treated with an oxaliplatin-containing regimen. Eligible pts receive Z (4 mg/kg) q2wks on day 1 of each cycle followed by FOLFIRI until disease progression, unacceptable toxicity, death, or investigator/pt decision. Initial starting doses and subsequent modifications are at treating physicians’ discretion. The percentage of pts with grade (G) 3/4 adverse events (AEs) in the combined safety population of ASQoP and AFEQT in B-pretreated pts is compared with that of B-pretreated pts in VELOUR. Results: At data cut-off, the safety population comprised 116 pts with ≥1 completed treatment cycle; 67 (57.8%) were pretreated with B. At least 1 G3/4 AE was experienced by 49.3% of pts vs 82.5% in VELOUR. Most G3/4 AEs were G3. There were no reports of G4 hypertension or proteinuria. Conclusions: Thisinterim safety analysis from ASQoP/AFEQT in pts pretreated with B has identified no new safety signals for Z and a trend toward decreased incidence of G3/4 events. The early analysis provides additional safety data and suggests an acceptable toxicity profile in this real-life setting. Clinical trial information: NCT01571284. [Table: see text]
Inhibition of migration and invasion of gastric cancer cells by Snail-regulated MiR-375 through JAK2 targeting.
Authors: Yanjun Xu , Tianhua Zhou , Jianmin Si , Wei Zhuo
Published: February 2017

56

Background: Migration and invasion of cancer cells are essential process during cancer metastatic procession. In gastric cancer, cells invasion into the surrounding tissue is a crucial early step. However, the mechanisms have not been fully understood. MicroRNAs, which are a class of small single-stranded non-coding RNA, participate in the malignant progressions of cancer, including metastasis. We study the association between specific dysregulated miRNA and specific metastasis step of gastric cancer, which will provide insights into the potential mechanisms of gastric cancer cells migration, invasion and metastasis. Methods: The expression of miR-375 was assayed using the quantitative real-time PCR analysis. Scratch-wound healing assay, Transwell migration and invasion assay were conducted to study the migration and invasion abilities of cells. Animal experiment was also conducted to examine the effect on liver and lung metastases by overexpression of miR-375. Luciferase assay was conducted to study the association between Snail and miR-375. Results: MiR-375 is downregulated in gastric cancer cells with greater migration and invasion abilities. The expression level of miR-375 is decreased in gastric cancer tissues from metastasis-positive patients compared with that from metastasis-free patients. Overexpression of miR-375 inhibits the migration and invasion abilities of gastric cancer cells. JAK2, which may be a target gene of miR-375, could reverses miR-375 induced inhibition of gastric cancer cells migration and invasion. Liver metastasis was not detected in mice injected with miR-375 overexpressed cells but was apparent in mice injected with cells which were transfected with control vector. The transcription factor Snail, which binds directly to the putative promoter of miR-375, could reduce the expression level of miR-375 significantly. A distinct inverse correlation was found between miR-375 expression and Snail mRNA level. Conclusions: These findings demonstrate that tumor suppressor miR-375, whose expression is directly regulated by the transcription factor Snail, inhibits gastric cancer cells migration, invasion and metastasis by targeting an important protein JAK2.
Use of preoperative tattooing to increase lymph nodes (LNs) retrieved in patients with locally advanced rectal cancer who receive preoperative chemoradiotherapy (CRT): A randomized controlled trial.
Authors: Okada Kazutake , Sadahiro Sotaro , Suzuki Toshiyuki , Tanaka Akira , Saito Gota , Kamijo Akemi
Published: February 2017

561

Background: Retrieval of 12 or more LNs has been recommended for accurate staging of colorectal cancer. Patients with 12 or more LNs revealed better survival than those with less than 12 LNs. Preoperative tattooing has been reported to improve LNs retrieval in colorectal cancer. Preoperative CRT had been reported to decrease the number of LNs in rectal cancer. We conducted a randomized controlled study to validate whether preoperative tattooing increases the number of LNs in patients who receive preoperative CRT. Methods: Fifty patients with cStage II or III rectal cancer were randomly assigned to a group in which tattooing was performed at the time of colonoscopy one day before surgery (T group) or a group without tattooing (C group). Submucosal injection of India ink was performed into two sites at the lower tumor margin. All patients had received 40/45 Gy of radiotherapy in 20/25 fractions with concurrent oral UFT or S-1 chemotherapy. Mesorectal excision was performed 6 to 8 weeks after completion of CRT. Results: There were no significant differences in characteristics between the groups. The numbers of retrieved LNs (mean±SD) in T group and C group were 14±7 and 8±6, respectively (p < 0.01). The number of patients with 12 or more LNs was greater in T group (56%) than in C group (20%) (p < 0.01). The numbers of metastatic LNs were similar in both groups (0.5±1.2 and 0.5±1.4, respectively p = 0.92). In patients with poor histologic tumor regression (TRG grade 3 to 5), the number of LNs was significantly greater in T group (17±9) than in C group (6±5; p < 0.01). However, in patients with good histologic response (TRG grade 1 or 2), the numbers of LNs in T group and C group were 11±5 and 9±6, respectively (p = 0.31). In T group, the numbers of LNs and metastatic LNs were significantly greater only in patients with poor histologic regression. Conclusions: Preoperative tattooing increases the number of retrieved LNs. In patients with poor histologic response following CRT, tattooing significantly increases the numbers of LNs. However, tattooing does not increase the numbers of retrieved LNs and metastatic LNs in patients with good histologic response. Clinical trial information: 11R-004.