bluuebunny/crossref_metadata_2025 huggingface dataset:
Expression of antibody-drug conjugates (ADC) biomarkers in colorectal cancer.
Authors: Thyparambil Sheeno P. , Liao Wei-Li , An Eunkyung , Tian Yuan , Heaton Robert , Sylvester Karl G. , Ling Xuefeng B.
Published: February 2020

17

Background: Multiple ADCs are in clinical trials for CRC and the optimal strategy for selecting patients who may benefit from the treatment is evolving. Due to the unique mechanism of ADCs, patient selection should involve screening not only for the presence of the antibody target, but also for markers of resistance or response to the payload. We have built a multiplexed ADC biomarker panel in FFPE tumor tissue that simultaneously quantifies the protein levels of the antibody targets and also the payload markers. Methods: FFPE tumor tissues from 363 CRC patients were microdissected and solubilized for mass spectrometry-based targeted proteomic analysis in our CLIA certified laboratory. We quantified protein levels of EGFR, HER2, HER3, Axl, Mesothelin, FRalpha, Trop2 (antibody targets), tubulin-beta3 and TOPO1 (payload resistance and response markers, respectively) simultaneously. The multiplexed assay also quantified additional 22 clinically relevant proteins. Results: Expression of EGFR(83%), HER2(52%), HER3(21.5%), Axl(3.7%), Mesothelin(26.5%), FRalpha(3.7%), and Trop2(59.8%) may indicate likely response to ADCs. Expression of TUBB3(+) and TOPO1 (>1350amol/µg) in antibody target-positive subset may suggest resistance or response to payloads, such as taxanes and irinotecan, respectively (Table). Previously we identified that HER2 expression >750amol/µg correlated with HER2 positivity. Accordingly, 1.4% (5/355) of CRC patients were HER2 positive, of which 40% (2/5) had TOPO1 expression >1350amol/µg (75th percentile) suggesting that these 2 patients may receive benefit from a HER2/TOPO1 ADC. (+) indicates expression ≥LOQ; (-) indicates expression <LOQ. Conclusions: In patients with CRC, quantitative proteomics identified both antibody targets and markers of resistance or response to the payloads for multiple approved and investigational ADC therapies.

[Table: see text]
Detection of microbial DNA in the blood of colorectal cancer patients for the diagnosis of microbial translocation.
Authors: Sougklakos Ioannis , Messaritakis Ippokratis , Vogiatzoglou Konstantinos , Tsantaki Konstantina , Dretaki Agapi , Koulouridi Asimina , Sfakianaki Maria , Koulouris Andreas Vasileios , Tzardi Maria , Mavroudis Dimitrios , Boukovinas Ioannis
Published: February 2020

171

Background: Dysbiosis has been associated with diseases and is of a major public health importance. Can lead to the passage of viable bacteria, their products or their fragments from the intestinal lumen through the mesenteric lymph nodes and other sites, known as bacterial translocation. The aim of the study was to determine whether microbial translocation occurs in stage II/III-IV colorectal cancer (CRC) patients and evaluate the usefulness of blood PCR for diagnosis of such translocation. Also to correlate the presence of Toll-Like Receptor and Vitamin D Receptor polymorphisms with the detection of microbial DNA fragments in the blood of CRC patients. Methods: Peripheral blood was obtained from 397 CRC patients (adjuvant n = 202 and metastatic n = 195) and 32 healthy individuals. DNA from all subjects was analyzed using PCR for amplification of genomic DNA encoding 16S rRNA, β-galactosidase gene of E. coli, Glutamine synthase gene of B. fragilis and 5.8S rRNA found in C. albicans. Results: Significantly higher rates of 16S rRNA, β-galactosidase, Glutamine synthase and 5.8S rRNA detection was observed in the pool of CRC patients than the controls ( p< 0.001). All microbial DNA fragments detected were also significantly associated with the metastatic disease ( p< 0.001) leading to shorter survival rates ( p< 0.001). Moreover, individuals with the homozygous mutant alleles of either TLR or VDR gene polymorphisms had significantly higher detection rates of microbial DNA fragments. Conclusions: The detection of microbial DNA fragments in patients with CRC highlights the role of these microbes in cancer development, progression and therefore in patients’ survival.
Circulating IL-8 levels as candidate marker of antiangiogenic therapy for metastatic colorectal cancer.
Authors: Suenaga Mitsukuni , Mashima Tetsuo , Kawata Naomi , Wakatsuki Takeru , Seimiya Hiroyuki , Yamaguchi Kensei
Published: February 2020

172

Background: Liver metastasis (LM) confers poor prognosis in metastatic colorectal cancer (mCRC). It has not been well understood whether tumor microenvironment (TM) varies throughout cancer treatment in the presence of LM. We therefore investigated the difference of TM between first-line (FL) and savage-line (SL) anti-angiogenic therapies and tested whether specific angiogenic cytokines could predict clinical outcome in mCRC patients. Methods: A combined cohort consisting of two cohorts (N = 125) was included in this study: 71 patients received first-line treatment with bevacizumab (FL cohort: median age, 60; median follow-up, 28.9 mos) and 54 patients received salvage-line regorafenib (SL cohort: median age, 65; median follow-up, 17.2 mos). Blood samples were obtained from all patients at baseline (BL) and during the treatment, and serum levels of nine angiogenic cytokines were measured using ELISA. Progression-free survival and overall survival (OS) were analyzed using Kaplan-Meier curves, log-rank test, and Cox proportional hazard regression. Results: In univariate analysis for the combined cohort, right-sided CRC, primary unresected, KRAS wild-type, LM, metastatic site ≥2 and SL were associated with shorter OS, and LM and SL remained in multivariable analysis. Ang-2 and IL-8 levels at BL were significantly associated with LM and their cut-off values were 2190.3 pg/ml (COAng2) and 15.1 pg/ml (COIL8), respectively, and no significant interaction was shown between cytokines and cohorts. Patients with Ang-2 levels ≥COAng2 had shorter OS than those with Ang-2 levels < COAng2 (HR2.57, 95%CI: 1.47–4.51, P = 0.001), while IL-8 levels ≥COIL8 was associated with shorter OS than < COIL8 (HR4.31, 95%CI: 2.11–8.79, P < 0.001). IL-8 levels ≥COIL8 remained as significant in multivariable analysis (HR 3.24, 95%CI: 1.47–7.16, P = 0.004). In change analysis, there was no significant correlation of variation of IL-8 and Ang-2 levels with clinical outcome after the first treatment cycle in both cohorts. Conclusions: Circulating IL-8 and Ang-2 levels reflect the TM in the presence of LM, of which IL-8 may serve as solid prognostic marker of anti-angiogenic therapy regardless of the timing of treatment for mCRC.
Defining population response of patient-derived colorectal cancer organoids against prospective clinical outcomes.
Authors: Kratz Jeremy D. , Gillette Amani , Rehman Shujah , Sunil Aishwarya , Johnson Katherine Anne , Pasch Cheri , Lemmon Kayla , Lassen Nicole , Abbott Daniel Erik , Carchman Evie , Foley Eugene F. , Heise Charles , Lawson Elise Hanna , LoConte Noelle K. , Lubner Sam Joseph , Mulkerin Daniel , Matkowskyj Kristina A. , Uboha Nataliya Volodymyrivna , Skala Melissa C. , Deming Dustin A.
Published: February 2020

177

Background: No current clinical tool can predict the efficacy of cancer therapeutics for patients with colorectal cancer (CRC). We recently demonstrated the feasibility of using patient-derived cancer organoids (PDCOs) to examine therapeutic response and tumor heterogeneity for individual patients with CRC via optical metabolic imaging (OMI). Here we expand these analyses in a cohort of patients with clinical outcomes. Methods: CRC tissue was collected from patients on IRB-approved protocols. PDCOs were matured and treated with chemotherapy regimens concurrent with patient treatment. Previously established effect size response thresholds were used for diameter ( > 1.5) and OMI ( > 0.5) following 48 hours of treatment. OMI measures the intrinsic autofluorescence of NAD(P)H and FAD using 2-photon microscopy without specific reagents or dyes. Clinical outcomes were prospectively collected by manual chart review. Results: 12 CRC PDCOs were established from patients with CRC. PDCOs were collected from initial diagnosis and advanced setting of both primary and metastatic sites by core needle biopsy and surgical resection. Differential growth rates were observed across lines. PDCOs with RAS/RAF alterations had more uniform growth, while PDCOs without these alterations demonstrated more heterogeneous growth and metabolism. Clinical correlation of PDCOs response with recurrence of disease in the adjuvant setting will be presented. Cases with prior 5-FU-based chemotherapy at the time of PDCO collection had intermediate sensitivity. For PDCOs collected pre-treatment, PDCO response predicted clinical response for 5 of 6 cases using predefined sensitivity thresholds. In the case that overall PDCO response did not predict clinical response, a heterogenous response was observed with distinct sensitive and resistant populations. Across PDCOs, greater post-treatment heterogeneity was observed in resistant lines compared to those with treatment sensitivity. Conclusions: Tumor heterogeneity in treatment response can be assessed using PDCOs growth and metabolism. The utility of PDCOs to predict clinical outcomes for patients with CRC deserves further prospective validation.
A 13-gene colorectal cancer liquid biopsy with greater than 90 percent accuracy in diagnosis and assessment of disease status.
Authors: Modlin Irvin Mark , Kidd Mark S. , Drozdov Ignat A. , Kitz Alexander , Malczewska Anna , Kusnierz Katarzyna , Aslanian Harry
Published: February 2020

18

Background: There are a paucity of blood-based biomarkers with clinical utility for colorectal cancer (CRC) We report the use of a 13-gene colon cancer circulating-free mRNA to diagnose CRC and identify disease burden and status Clinical utility of the assay was assessed in surgical and chemotherapy patients. Methods: Gene identification and validation: Publicly available colon cancer transcriptomes (TTs) (E-MTAB-57); gene expression in 3 CRC cell-lines (LOVO, LS-180 and COLO320DM) and validated in CRC tumors ( n=33). Tumor TT analysis: co-expression network generation and differential expression analysis compared with normal blood-based TT to identify candidate markers. Blood gene expression: CRC set (cancers: n=312, controls n=117) and a CRC artificial intelligence model constructed. Normalized gene expression algorithmically scored (0-100). Matched tumor/ blood samples were available in 33 patients. RECIST criteria Clinical score assessment: Score utility was assessed in surgical and treated cohorts: Surgical: n=37, follow>7 days. Treated chemotherapy: n=75; stable disease (SD): n=20, progressive disease (PD): n=55) The relationship to CEA and CA-19-9 were assessed. Statistics: Non-parametric (Mann-Whitney), Pearson-correlation, Fisher’s and AUROC analyses (Mean±SEM). Results: Transcriptomic analysis identified a13 genes blood signature for CRC. Expression levels were significantly elevated ( p<0.001, 20-100-fold) in cell lines and CRC tumors. The matched tissue/blood correlation r: 0.795 ( p=0.002). In CRC, levels were 54.4±1.5 ( p<0.0001) compared to controls (9.5±1.7); AUROC: 0.91±0.02, accuracy 90.5% (sensitivity 93.1%, specificity 81.2%). Surgical cohort: CRC assay accuracy 100% vs CEA (35%) or CA-19-9 (17%) (both p<0.0001). Resection (R0 – 92%) significantly decreased levels (47±2) at follow-up ( p=0.0055). Treated cohort: Levels were elevated in PD (63±4.1) vs SD (30±3.2, p<0.0001). Conclusions: The diagnostic accuracy of CRC blood-signature was 91%; significantly greater than CEA or CA-19 Surgical resection decreased levels. CRC score was elevated in progressive vs stable disease. A colorectal cancer liquid biopsy has probable clinical utility in management.
Plasma deoxyuridine levels in metastatic colorectal cancer (mCRC) patients and clinical response after 5FU based therapy in combination with arfolitixorin.
Authors: Taflin Helena Anna , Odin Elisabeth , Carlsson Göran Ulf , Tell Roger , Gustavsson Bengt , Wettergren Yvonne
Published: February 2020

185

Background: 5-Fluorouracil (5FU) is one of the most commonly used cancer drugs. FdUMP is a metabolite from 5FU which inhibits thymidylate synthase (TS). TS inhibition causes a rise in the intracellular pool of the natural TS substrate dUMP leading to increased global levels of deoxyuridine (dUR). In order to increase TS inhibition, 5FU is combined with folates, usually leucovorin (LV). However, LV needs enzymatic conversion to become active. The LV conversion capacity differs between patients, with the consequence that many patients will have a reduced effect of 5FU-based treatment. Arfolitixorin is a biologically active folate, 5,10-methylenetetrahydrofolate, which doesn´t need enzymatic conversion; hence it could potentially be more efficacious. Methods: ISO-CC-005 is a phase I/II safety and tolerability study in mCRC patients receiving 5FU/arfolitixorin alone or in combination with irinotecan or oxaliplatin ± bevacizumab. Efficacy was evaluated as Overall Response Rate (ORR) after 4 cycles of chemotherapy. A LC-MS/MS method was developed for quantification of plasma dUr and 5FU. Blood samples were collected before and 24 h after 5FU administration at the first (C1) and fourth (C4) treatment cycle (n = 33). The C4/C1 ratio for dUr and 5FU, respectively, were calculated.The clinical response, which is an ordered categorical data, was modelled using nominal logistic regression using the ratio C4/C1 and the absolute values C1 and C4 as explanatory variables. Results: A positive correlation between dUr and 5FU levels was seen (r = 0.89, p = 0.0011) at C4. The C4/C1 ratio for dUr and 5FU in plasma correlated positively (r = 0.95, p = 0.0001). A positive correlation was also found between arfolitixorin dose and plasma levels of dUr (r = 0.44, p = 0.016) at C1, but not at C4. In the model test, the C4/C1 dUr ratio, as well as the individual dUr levels at C1 and C4, had a highly significant effect on ORR (p = 0.0075). Conclusions: The results demonstrate that the arfolitixorin dose has an impact on plasma levels of dUr and also on clinical response. dUr could potentially be an early surrogate marker for TS inhibition in mCRC patients treated with 5FU/arfolitixorin. Clinical trial information: NCT02244632.
Prognostic and predictive impact of primary tumor sidedness in first-line trials for advanced colorectal cancer: An analysis of 7,828 patients in the ARCAD database.
Authors: Yin Jun , Cohen Romain , Jin Zhaohui , Liu Heshan , Pederson Levi , Adams Richard , Maughan Tim , Venook Alan P. , Van Cutsem Eric , Cremolini Chiara , Tebbut Niall , Seymour Matt , Bokemeyer Carsten , Diaz-Rubio Eduardo , Wasan Harpreet Singh , Heinemann Volker , De Gramont Aimery , Shi Qian , Lenz Heinz-Josef
Published: February 2020

188

Background: Unplanned subgroup analyses from several studies have suggested primary tumor sidedness (PTS) as a potential prognostic and predictive parameter in metastatic colorectal cancer (mCRC). We aimed to investigate the prognostic and predictive impact of PTS on outcomes. Methods: PTS data of 7,828 mCRC patients (pts) from 10 first-line randomized trials in the ARCAD database were pooled. PTS was defined as right-sided (RS) or left-sided (LS) if tumor arose from the cecum to the hepatic flexure or from the splenic flexure to the rectum, respectively; transverse colon cancers were not included. Overall survival (OS) and progression-free survival (PFS) were assessed using Kaplan-Meier and Cox models adjusting for age, sex, performance status (PS), prior radiation/chemo, and stratified by treatment arm. Predictive value was tested by interaction term between PTS and treatment (anti-EGFR plus chemotherapy vs. chemotherapy alone). Results: Compared to RS pts (2407, 31%), LS pts (5421, 69%) had better OS (median: 21.6 v 16.8 mos; HRadj: 0.73, 95% CI 0.69-0.78, P < .001) and PFS (median 8.4 v 7.2 mos; HRadj: 0.81, 95% CI 0.76-0.86, P < .001). Results were consistent among subgroups defined by age, sex, PS, metastatic sites and chemo backbone (irinotecan- and oxaliplatin-based). Interaction between PTS and KRAS mutation was significant (Pinteraction< .001): LS is associated with better prognosis only among KRAS wild-type (wt) (HRadj: OS 0.62, 95% CI, 0.55-0.70; PFS 0.71, 95% CI 0.63-0.80), but not among KRAS mutated pts. Among KRAS wt pts, survival benefit from anti-EGFR was observed for LS, but not for RS (table). Conclusions: The prognostic value of PTS is restricted to the KRAS wt population. PTS is predictive of anti-EGFR efficacy, with a significant improvement of survival for LS mCRC pts. These results suggest treatment stratification in mCRC studies by both PTS and KRAS status.

[Table: see text]
Predicting CD8+ T-cell infiltration in colorectal cancer using versican proteolysis across molecular profiles.
Authors: Johnson Katherine Anne , Emmerich Philip , Matkowskyj Kristina A. , Deming Dustin A.
Published: February 2020

189

Background: The clinical indications for immunotherapies continue to increase across cancer types. In colorectal cancer (CRC), there has been little progress in the use of these therapies outside of mismatch repair deficient cancers (dMMR). However, even in dMMR cancers only a minority actually respond to the FDA-approved anti-PD1 agents. The tumor microenvironment is increasingly implicated in the resistance of cancers to immune-based therapies. Our group has previously described that accumulation of a matrix proteoglycan, versican, correlates with a reduction in CD8+ T-cell infiltration in CRCs, while proteolysis of versican, releasing the bioactive fragment versikine, correlates with increased infiltration. Here we examine the impact of pathogenic mutations on the utility of MMR status and versican proteolysis to predict CD8+ T-cell infiltration. Methods: Matched normal colon and CRC tissues from 122 patients were stained for versican, versikine, MLH1, MSH2, MSH6, PMS2, CNNB1, and CD8. Each was reviewed by a blinded GI surgical pathologist and CD8 quantified as tumor infiltrating lymphocytes (TILs) per high power field (hpf). 107 of the CRC samples were available for sequencing using the Qiagen Comprehensive Cancer Panel examining 160 genes across cancer relevant hotspots. The molecular profile was correlated with the IHC staining. Results: As previously reported, dMMR tumors had higher CD8+ T-cell infiltration. This trend persisted across dMMR genotypes (dMMR vs proficient (p)MMR p = 0.0016). Versican proteolysis correlated with increased CD8+ T cell infiltration in dMMR and pMMR cancers and was present in cancers with/without APC, TP53, and KRAS mutations. Across common mutations, cancers with the versican proteolysis predominant phenotype had more CD8+ T-cell infiltration than those without (APC mutant (mt): 11.82 vs 1.97 CD8+ TILs/hpf, p < 0.001; KRAS mt: 9.39 vs 3.08, p = 0.15; BRAF mt: 25.00 vs 7.50, p = 0.13; TP53 mt: 8.61 vs 1.63, p < 0.001). Conclusions: Across common mutations, versican proteolysis predicts CD8+ T-cell infiltration in both dMMR and pMMR CRC. Further investigation into whether this increase in infiltration will lead to greater immunotherapy response is warranted.
Effect of previous chemotherapy treatments on circulating tumor-associated cells in colorectal cancer.
Authors: Limaye Sewanti , Crook Timothy , Page Raymond , Patil Darshana , Akolkar Dadasaheb , Datta Vineet , Schuster Stefan , Sims Cynthe , Patil Revati , Patil Sanket , Mhase Vishakha , Apurwa Sachin , Khan Shabista , Srinivasan Ajay , Datar Rajan
Published: February 2020

194

Background: Resistance to combination regimens of fluorouracil, oxaliplatin and irinotecan are commonly observed in Colorectal cancers (CRC). There are presently no viable approaches for ‘real-time’ monitoring of innate and acquired chemoresistance. We used a novel method for chemo-interrogation (CI) by harvesting from peripheral blood sufficient numbers of Circulating-Tumor Associated Cells (CTACs) which are defined as apoptosis-resistant cells of tumorigenic origin which are positive for Epithelial Cell Adhesion Molecule (EpCAM) and pan-cytokeratins (pan-CK) irrespective of CD45 status. Methods: Peripheral blood was collected from 110 patients with confirmed diagnosis of CRC, among whom 56 were recently diagnosed and therapy naïve while 54 were pre-treated. Peripheral blood mononuclear cells (PBMCs) were harvested by centrifugation and treated with commercially available stabilizing agents by a proprietary protocol to stabilize apoptosis resistant C-TACs. Surviving C-TACs were confirmed by immunostaining for EpCAM and pan-CK. Harvested C-TACs were treated in vitro with 5-fluorouracil, irinotecan and oxaliplatin and the fraction of surviving cells estimated to determine resistance profiles. Results: Among the 56 cases of recently diagnosed therapy naïve CRC, innate chemoresistance was observed in 37.5%, 47.3% and 41.8% of samples (unique patient-drug combinations) towards 5-fluorouracil, irinotecan and oxaliplatin respectively. Among the 54 cases of previously treated CRC, acquired chemoresistance was observed in 92.6%, 92.6% and 95.5% of samples towards 5-fluorouracil, irinotecan and oxaliplatin respectively. Conclusions: We show for the first time that sufficient C-TACs can be harvested for meaningful CI in newly diagnosed treatment naïve CRC as well as refractory CRCs. Post-treatment chemoresistance being an order of magnitude higher than the untreated cohort indicates that C-TACs in CRC are resistance-educated by previous treatments and can guide treatment strategy.
5-hydroxymethylation signatures in plasma circulating cell-free DNA as markers for appendiceal and colorectal peritoneal metastasis.
Authors: Berger Yaniv , Cui Xiao-Long , West Diana , Rivas Marco , Hsu Phillip , Dougherty Urszula , Deng Zifeng , Hindi Enal , Eng Oliver , Bissonnette Marc , Polite Blase N. , Turaga Kiran
Published: February 2020

195

Background: Noninvasive tests for peritoneal metastasis (PM) detection lack sensitivity. Genome-wide mapping of 5-hydroxymethylcytosine (5hmC) on nanogram quantities of peripheral plasma circulating cell-free DNA (PcfDNA) was previously shown to differentiate non-metastatic colorectal cancer from healthy subjects. We aimed to examine if patients with colorectal cancer (CRC), high grade appendiceal cancer (HGA) or low grade appendiceal cancer (LGA) with PM have distinct signatures of 5hmC in PcfDNA compared to each other and to patients matched for tumors without PM. Methods: We analyzed plasma samples from a prospectively collected tissue bank. To correlate 5hmC signatures with intraoperative findings, only patients who underwent abdominal surgery in proximity to plasma collection were selected. Key steps of PcfDNA processing included extraction from plasma, nano-hmC-Seal chemical labeling and enrichment of 5hmC-modified fragments, next-generation sequencing, and mapping to the reference human genome. DESeq2 R package was finally used to compare relative 5hmC enrichment and detect distinct 5hmC signatures according to disease histology and PM presence. Results: Plasma samples were collected between 11/2016 – 3/2019 from 46 patients with CRC (n = 21), HGA (n = 17) and LGA (n = 8). Of those, 32 (70%) had PM based on intraoperative findings (median peritoneal cancer index score = 15, range 2-39) and 14 did not have PM. Most samples (91%) were collected on the same day as surgery. An average of 24 million paired-end reads were sequenced for each sample. Four samples (8.7%) were excluded from the analysis due to low sequencing coverage or high duplication level. Unique 5hmC enrichment patterns were found to discriminate with p < 0.05 between CRC PM and HGA PM (n = 616 differentially hydroxymethylated genes (DHMGs)), CRC/HGA PM and LGA PM (n = 1074 DHMGs), and CRC/HGA PM and CRC/HGA patients without PM (n = 1576 DHMGs). Conclusions: Distinct signatures of 5hmC in PcfDNA could differentiate patients with CRC/HGA/LGA PM from each other and from patients with similar tumor histologies without PM. Thus 5hmC signatures in PcfDNA might potentially serve as a sensitive marker of occult PM.
Association of KRAS and BRAF mutations with progression-free survival (PFS) with second-line FOLFIRI +/- regorafenib in metastatic colorectal cancer (mCRC).
Authors: Innocenti Federico , Selitsky Sara R. , Parker Joel S. , Auman James Todd , Hammond Kelli , Patil Sushant A. , Sanoff Hanna Kelly , Lee Michael Sangmin
Published: February 2020

196

Background: LCCC1029 was a 2:1 randomized phase II trial of 2nd-line FOLFIRI plus either regorafenib or placebo in mCRC. The addition of regorafenib improved PFS (median PFS 6.1 vs 5.3 mo, HR 0.73, 95% CI 0.53-1.01). However, the effect of somatic mutations on regorafenib activity has not been tested. Methods: We performed whole exome sequencing on archival primary tumor tissue and paired normal tissue in 85 patients of LCCC1029. We compared PFS and OS using Kaplan-Meier method and log-rank tests, and hazard ratios (HR) were estimated using Cox proportional hazards method. Results: Among the 85 subjects, 54 (64%) had tumors wild-type (WT) for KRAS and BRAF, 26 (31%) had tumors with KRAS mutations in exons 2-4, and 5 (6%) had tumors with BRAF V600E. The addition of regorafenib to FOLFIRI improved PFS in the KRAS/ BRAF WT subgroup (median PFS 8.0 vs 4.9 mo, HR 0.68, 95% CI 0.48-0.97, log-rank p=0.028), but not in the KRAS mutant subgroup (median PFS 6.8 vs 5.5 mo, HR 0.90, 95% CI 0.61-1.35, log-rank p=0.617) or the BRAF mutant subgroup (log-rank p=0.156). In all of these subgroups, the addition of regorafenib was not associated with significant difference in OS. BRAF V600E was prognostic and associated with significantly worse OS (median OS 8.4 vs 18.0 mo, HR 2.59, 95% CI 1.01-6.66, log-rank p=0.04). Conclusions: The addition of regorafenib to FOLFIRI improves PFS among the subgroup of patients with KRAS and BRAF dual WT CRC, but not among the KRAS mutant subgroup. These results indicate that the addition of anti-angiogenic therapy to second-line chemotherapy backbones may be more effective in KRAS/ BRAF WT tumors in particular. More confirmatory studies are needed to corroborate this finding.
Real-world costs of cetuximab + chemotherapy administered every two weeks versus weekly for treatment of metastatic colorectal cancer.
Authors: Pescott Chris , Batech Michael , Boutmy Emmanuelle , Ronga Philippe , Lamy Francois-Xavier
Published: February 2020

22

Background: Cetuximab (CET) 250 mg/m2 weekly (q1w) after an initial dose of 400 mg/m2 added to chemotherapy (CT) is licensed for treatment of (K)RAS wild-type metastatic colorectal cancer (mCRC). In practice, administration of CET 500 mg/m2 plus CT every 2 weeks (q2w) is common. We compared healthcare costs between q2w and q1w regimens in a US claims database study. Methods: A cohort of 2,943 mCRC patients CET-treated between 2010 and 2016, identified in IBM MarketScan, was analyzed for costs associated with CET+CT q2w vs q1w. All-category costs (ACC), stratified by overall outpatient (OO), inpatient (OI), and pharmacy (OP) claimed costs during the exposure period, were compared between groups. Additionally, subcategories of CRC- and skin toxicity (ST)–related claims were explored, and imputation of capitated claim costs was performed. Patients were weighted by the stabilized inverse probability of treatment (IPTW) based on a high-dimensional propensity score to control for confounding. Generalized linear models (GLMs) with gamma distributions were used to compare regimens. Inflation-adjusted costs (2016 US dollars) are presented per patient per month ($PPPM) with 95% CIs. Results: 1,779 and 951 patients were classified as q1w and q2w, respectively. Median ACC were 14,089 (q2w) vs 14,034 (q1w) $PPPM. Mean ACC and OO, OI, and OP costs are summarized in the table. CIs overlapped in each category, with GLMs showing no statistically significant differences. This finding was similar for CRC and ST subcategories. Conclusions: No cost differences were found between q2w and q1w regimens. In line with published noninferiority of overall survival with the q2w regimen, it represents an effective, cost-neutral option for treating mCRC patients.

[Table: see text]
Clinical utility of the systemic inflammatory response (SIR) in identifying high-risk stage II colon cancer.
Authors: Golder Allan Matthew , McMillan Donald C. , Mansouri David , Horgan Paul G. , Roxburgh Campbell SD
Published: February 2020

220

Background: Surgery for TNM Stage II colon cancer is considered curative however approximately 20% of patients will have recurrence of their disease. A number of high risk pathological features guide the use of adjuvant chemotherapy. More recently the preoperative SIR has been consistently shown to have prognostic value but to date has not been utilised clinically as a high risk feature. The present study compared the influence of the SIR versus established high-risk clinical features on overall/cancer specific survival (OS/CSS). Methods: Patients in the West of Scotland undergoing curative resection for Stage II colon cancer from 2011-2015 were identified with survival updated until December 2018. Additional data was obtained from online records. Through uni/multivariate analysis (UVA/MVA) we compared the effect on survival of the SIR measured using the modified Glasgow Prognostic Score (mGPS), neutrophil-lymphocyte ratio (NLR) and lymphocyte-monocyte ratio (LMR) when entered individually into a multivariate model alongside established high-risk features. Results: 982 patients were identified having had a curative resection of Stage II colon cancer. Median follow up was 61 months and there were 307 deaths during follow up. For OS: emergency presentation, T stage, adjuvant chemotherapy, nodal harvest, margin involvement, mGPS, LMR, NLR (all p≤0.001) and EMVI (p < 0.05) were significant on UVA. On MVA: age (HR 1.51), T stage (HR 1.59), nodal harvest (HR 1.67), margin involvement (HR 1.94), adjuvant chemotherapy (HR 0.47), mGPS (HR 1.38), NLR (HR 1.35) and LMR (HR 1.50) remained significant (all p < 0.05). For CSS: age, emergency presentation, T stage, margin involvement, mGPS, NLR, LMR (all p < 0.001), nodal harvest and adjuvant chemotherapy (both p < 0.05) remained significant on UVA. On MVA emergency presentation (HR 1.88), T stage (HR 2.02), margin involvement (HR 2.98), adjuvant chemotherapy (HR 0.51) and mGPS (HR 1.34) remained significant (all p < 0.05). Conclusions: The present study suggests that the SIR is an independent predictor of worse OS/CSS in Stage II colon cancer and should be considered a high risk feature in future prospective studies.
Systemic inflammation, adjuvant chemotherapy, and survival in stage III colorectal cancer: Results from the ScotScan Colorectal Cancer Collaborative.
Authors: Park James Hugh , Fuglestad Anniken Jorlo , Kostner Anne Helene , Oliwa Agata , Roxburgh Campbell SD , Horgan Paul G. , Graham Janet Shirley , Kersten Christian , McMillan Donald C.
Published: February 2020

228

Background: The systemic inflammatory response (SIR) is a poor prognostic marker in patients with colorectal cancer (CRC), and predicts poor outcome following adjuvant chemotherapy. Whether this may be influenced by chemotherapy regime is not known. The present study examined the relationship between the pre-operative SIR, adjuvant therapy regime, and survival of patients with stage III CRC in the ScotScan cohort. Methods: Patients with stage III CRC in Scotland (1997-2015, n= 317) and Norway (2000-17, n= 312) were included. The pre-operative SIR was measured using C-reactive protein (CRP≤10mg/L or > 10mg/L). Adjuvant status was categorised as none, 5-fluorouracil-only (5FU or capecitabine), or oxaliplatin-combination (Ox). Relationship with 3 year overall (OS) and cancer-specific survival (CSS) was examined. Results: Rates of Ox were comparable between cohorts (Scotland – 26% vs. Norway 28%), although more patients from Norway received single 5FU (4% vs. 19%, P= 0.005). 36% of each cohort were systemically inflamed. Ox was associated with superior OS (90%) and CSS (92%) when compared to 5FU (77% and 84%) and no therapy (61% and 72%, both P< 0.001). Stratified by SIR, patients with CRP≤10mg/L receiving Ox or 5FU had comparable 3yr OS greater than those receiving none (90% vs. 88% vs. 67%), whereas those with CRP > 10mg/L receiving Ox had superior survival than those receiving 5FU or no therapy (89% vs. 64% vs. 53%, P-for interaction = 0.101). Results were similar for CSS (CRP≤10mg/L: 91% vs. 94% vs. 79%; CRP > 10mg/L: 94% vs. 72% vs. 62%, P-for interaction= 0.01). Although patients receiving Ox were younger and less comorbid, both use of Ox and SIR remained independently associated with OS and CSS. Conclusions: Although selection bias in the choice of adjuvant therapy may confound analysis, this study suggests the SIR may aid in determining response to adjuvant therapy. Whereas non-inflamed patients with stage III CRC may benefit from single 5FU, those with an elevated SIR may benefit greater from more intensive, Ox-based regimes. These results remain to be validated, however support the use of the SIR as a prognostic and predictive biomarker in patients with stage III CRC.
The potential for combined measures of the systemic inflammatory response (SIR) in colon cancer: An analysis of 2,300 patients.
Authors: Golder Allan Matthew , McMillan Donald C. , Mansouri David , Park James Hugh , Roxburgh Campbell SD , Horgan Paul G.
Published: February 2020

233

Background: The preoperative SIR clearly demonstrates independent prognostic significance following curative resection for colon cancer, independent of TNM stage. SIR can be measured either using acute phase proteins, predominantly the modified Glasgow Prognostic Score (mGPS) or using the differential white cell count, predominantly neutrophil-lymphocyte ratio (NLR) or lymphocyte-monocyte ratio (LMR). The present study investigates the potential to combine these scores to better predict overall/cancer specific survival (OS/CSS). Methods: Patients in the West of Scotland undergoing curative resection for Stage I-III colon cancer from 2011-2015 were identified with survival updated until December 2018. Through uni/multivariate analysis (UVA/MVA) we compared the effect on OS/CSS of the SIR measured using a combination of mGPS and either NLR or LMR with other clinicopathological features. Results: 2312 patients were identified having underwent curative surgery. Median follow up time was 60 months and there were 756 deaths during follow up. On UVA: mGPS (0/1/2), NLR ( < 3/3-5/ > 5) and LMR ( < 2.5/2.5+) were significant for OS and CSS (all p < 0.001). In the multivariate model when a combination of mGPS and NLR were entered, both remained significant for OS (mGPS: HR 1.20, p = 0.002 and NLR: HR 1.23, p = 0.001) and CSS (mGPS: HR 1.25, p = 0.003 and NLR: HR 1.21, p = 0.017) adjusted for age, sex, emergency presentation, site, ASA, TNM stage and margin involvement. With mGPS and LMR in the model similar results were seen for OS (mGPS: HR 1.24, p = 0.005 and LMR: HR 1.49, p = 0.003) and CSS (mGPS: HR 1.20 p = 0.001 and LMR: HR 1.45, p < 0.001). When mGPS and NLR were scored 0/1/2 (mGPS 0/1/2 and NLR < 3/3-5/ > 5) and combined to form a cumulative grade (0/1/2/3/4) this offered better stratification of %OS (86/84/72/61/55) than mGPS (83/75/60) or NLR (86/79/66) alone (all p < 0.001). Similar results were seen for %CSS – mGPS (90/79/71), NLR (91/85/75) and combined grade (92/90/77/71/67). Conclusions: The results of the present study show that the combined use of mGPS and a differential white cell-based score (NLR/LMR) stratify OS/CSS better than a single measure of preoperative SIR alone.
The Reg4-CD44ICD pathway and tumor aggression in early-stage colon adenocarcinoma.
Authors: Sninsky Jared , Bishnupuri Kumar , Gonzalez Ivan , Trikalinos Nikolaos , Chen Ling , Dieckgraefe Brian
Published: February 2020

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Background: Reg4, or regenerating islet-derived protein 4, is highly expressed in gastrointestinal malignancies and acts as a mitogenic factor. Our prior work suggests a direct invitro interaction between Reg4 and CD44, a non-kinase transmembrane glycoprotein over-expressed in cancer stem cells. Reg4 is hypothesized to activate CD44 proteolytic cleavage to release intra-cytoplasmic domain of CD44 (CD44ICD). The goal of this study was to demonstrate clinical validity of the Reg4-CD44/CD44ICD pathway in stage II/III colon cancer and its association with disease recurrence and clinical parameters of aggression. Methods: We constructed a tissue microarray (TMA) consisting of 94 stage II/III matched colon adenocarcinoma patients, 23 of which had recurrent disease after potentially curative treatment. The TMA was immunohistochemically stained for Reg4, CD44, and CD44ICD proteins with specific antibodies. Spearman’s correlation, Chi-square tests, and Kaplan-Meier analyses were used to identify protein associations with tumor characteristics, recurrence and overall survival. Results: Expression of Reg4, CD44, and CD44ICD was associated with tumor location, size, tumor grade, mismatch repair (MMR) status, and disease recurrence. TMA data analysis supported our proposed mechanism of Reg4-mediated release of CD44ICD into the cytoplasm. A significant correlation was observed between Reg4 and CD44 (r2= 0.23, P = 0.028), CD44 and CD44ICD (r2= 0.36, p = 0.0004), and Reg4 and CD44ICD (r2= 0.45, p = < 0.0001). Reg4 expression was associated with larger tumor size (r2= 0.23, p = 0.026) . No association was observed between Reg4, CD44, or CD44ICD expression with disease recurrence. In patients who recurred, Reg4-positive patients had a median survival of 4 years vs. 7 years for Reg4-negative patients (p = 0.04). Our lab has performed in vitroexperiments with CD44ICD interfering peptides blocking the pro-proliferative effect of Reg4. Conclusions: These results suggest that Reg4-positive tumors, while not at higher risk of recurrence, are associated with decreased survival in established recurrent colon adenocarcinoma. Inhibition of the Reg4-CD44/CD44ICD pathway may be a future therapeutic target.
BRCA mutations in metastatic colorectal cancer (mCRC) and response to chemotherapy.
Authors: Feng Sophie , Al-Showbaki Laith , Krzyzanowska Monika K. , Prince Rebecca M. , Stockley Tracy , Yu Celeste , Bedard Philippe L. , Siu Lillian L. , Chen Eric Xueyu
Published: February 2020

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Background: Mutations in BRCA 1/2 are typically associated with breast, ovarian, pancreatic and prostate cancers. BRCA mutations have been reported in colorectal cancer in sporadic case series. Unlike other cancers, the significance of BRCA mutations in mCRC is not known. We report the prevalence and molecular characteristics associated with BRCA 1/2 mutations in mCRC, and investigate the impact of these mutations on chemotherapy response since both oxaliplatin (OX) and irinotecan (IRI) interfere with DNA repair pathways. Methods: The Ontario-wide Cancer Targeted Nucleic Acid Evaluation (OCTANE) database was queried to identify mCRC patients (pts) harbouring BRCA 1/2 mutations. BRCA 1/2 mutations were detected using panel-based next generation sequencing (NGS) on archival tumour tissue. Clinical and molecular variables were collected, together with treatment outcomes. Results: Of 279 mCRC pts within the OCTANE database as of March 2019, 9 pts with BRCA 1/2 mutations were identified (3.2%): 4 BRCA 1 and 5 BRCA 2 mutations. Each patient had a unique variant with 8/9 missense mutations and 1/9 splicing error. Allele frequency ranged from 0.11 to 0.57. RAS or BRAF mutations were present in 67%. Common co-mutations included TP53 (56%), APC (56%), TSC1 (44%), ROS1 (33%) and ATM(33%). 2 pts were mismatch repair deficient. Median age was 48.5 years (range: 31-69 years), 56% males. 5 pts presented with de novo metastatic disease. First line OX-containing chemotherapy was administered to 4 pts, and IRI to 3 pts. 2 pts did not receive chemotherapy (1 had surgery only post-adjuvant OX, and 1 immunotherapy). Overall response rate (ORR) was 71%, with all pts achieving a partial response or stable disease. The median progression-free survival was 7.5 months (range: 1.8- 31.7 months) and median overall survival 68.5 months (range: 10.5- 68.5 months) respectively. Conclusions: BRCA 1/2 mutations are present in a small subset of mCRC pts. Pts with these mutations tend to be younger at diagnosis. BRCA 1/2 mutations are associated with favourable response to first line chemotherapy. Targeting BRCA 1/2 mutations may broaden treatment options for these patients.
KRAS status and survival in multicenter study of RAS mutations (MURAS) in patients with colorectal liver metastases receiving Y90 radioembolization treatment.
Authors: Dendy Case Meaghan , Ghodadra Anish , Novelli Paula M. , Wu Vincent , Ganguli Suvranu , Wildgruber Moritz , Kohler Michael , Robinson Charles , Kim Charles , Wang David S. , Sze Daniel Y. , Kolbeck Kenneth , Russell Lori , Uhlig Johannes , Ludwig Johannes M. , Kim Hyun S.
Published: February 2020

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Background: Assessment of the clinical outcome and prognostic factors of patients receiving Y90 radioembolization to treat colorectal liver metastases. Methods: Multi-institution retrospective study of patients with CRLM who received Y-90 radioembolization after undergoing molecular testing was performed. Patients treated between 2008 and 2019 were included from 7 institutions within the United States and Europe. Patient demographics, systemic therapy regimens, tumor characteristics, laboratory values and overall survival were analyzed between patients with differing histopathologic and genomic status. Genomic profiling was obtained for KRAS, BRAF, PIK3CA, AKT, MEK, NRAS and MMR genes. Kaplan-Meier survival estimation and multivariate Cox regression were analyzed. Results: 445 patients treated with Y90 radioembolization fulfilled the inclusion criteria. Of the total cohort, 430 patients were available who had documented KRAS mutation status. Average age at diagnosis was found to be 58.4 years for all patients (57.6% male, 42.4% female). Decreased survival post Y90 treatment was shown in those patients with mutant KRAS. The median survival in patients with mutated KRAS was 5.7 months, as compared to 8.0 months in those patients with wild-type KRAS (p< 0.0001). Patient demographics including age, gender and race were not shown to be significant in overall survival post-Y90 treatment (p-values >0.05). The average number of systemic chemotherapy treatments prior to Y90 was 2.15 and was similar between KRAS groups (mutant vs wild type.) KRAS status persisted as a significant predictor of survival on multivariate analysis (p< 0.0001; HR 2.1 CI: 1.54-2.85). Conclusions: KRAS status is shown as a significant prognostic factor in patients with colorectal liver metastases receiving Y90 radioembolization.
A comparison study of the intratumoral microbiome in younger verses older-onset colorectal cancer (COSMO CRC).
Authors: Weinberg Benjamin Adam , Wang Hongkun , Geng Xue , Shokralla Shadi , Bakhshi Emily , Chaldekas Krysta , Harris Brent T. , Clagett Dionyssia , Marshall John
Published: February 2020

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Background: Although colorectal cancer (CRC) incidence has declined overall, CRC in individuals under age 45 has risen dramatically, particularly in the distal colon and rectum (left-sided CRC). The intratumoral microbiome (MB) of young individuals may be responsible. Certain bacteria disrupt colonic luminal integrity and promote inflammation, leading to oncogenic mutations in colonic epithelial cells. Fusobacterium nucleatum ( F. nuc) promotes CRC by suppressing immune response within the tumor microenvironment, activating the β-catenin pathway, and causing chemoresistance due to autophagy. Methods: We compared the intratumoral MB in CRC patients (pts) diagnosed before age 45 and after age 65. Primary and metastatic tumors were included. DNA was extracted from tumors and analyzed using 16S ribosomal gene sequencing. We compared the frequency of F. nuc and other bacterial and fungal DNA in tumors from younger- vs. older-onset CRC pts. Results: Tumors from 18 younger pts (median age 39.2 years) and 13 older pts (median age 72.8 years) underwent analysis. In total, 478 unique bacterial and fungal species were detected. F. nuc was found in tumors of 5 younger pts (28%, 4 left-sided and 1 right-sided primary) and 3 older pts (23%, 1 left-sided and 2 right-sided; P = NS, Fisher’s Exact test). A significant difference was seen in the rate of Moraxella osloensis (11% vs. 46%, P = 0.043, Fisher’s Exact test) in younger vs. older pts. There was no significant difference in MB diversity in younger vs. older pts. Conclusions: F. nuc is present in a greater number of tumors in pts with CRC diagnosed before age 45 than previously thought. Intratumoral bacterial profiling may discover patterns that explain the rising incidence of CRC in younger individuals and might eventually inform the development of novel therapeutics and adaptive cancer screening methods.
Tumor area and microscopic extent of invasion to determine circulating tumor DNA fraction in plasma and detectability of colorectal cancer (CRC).
Authors: Bredno Joerg , Lipson Jafi , Venn Oliver , Gross Samuel , Fields Alexander P. , Beausang John F. , Liu Qinwen , Brooks Jacqueline D. , Chen Xiaoji , Lopatin Rita , Liu Hai , Zhang Nan , Zhang Lori , Wu Tony , Allen Brian , Hartman Anne-Renee , Hubbell Earl , Jamshidi Arash , Aravanis Alex
Published: February 2020

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Background: Circulating Cell-free Genome Atlas (CCGA; NCT02889978) is a multi-center, case-control, observational study with longitudinal follow-up to develop a cfDNA assay in which classifiers were trained on whole-genome bisulfite sequencing (WGBS) and targeted methylation (TM) sequencing data for detection of multiple cancer types. Previously, we showed that the fraction of ctDNA fragments (TF) was a stronger predictor of cancer detection than clinical stage and an equivalent predictor for survival. Given that CRC tumors can be described via surface area (TSA) and microscopic tumor extent (microinvasion), CRC was used as a model to examine the biophysical determinants of TF. Methods: Detection of multiple cancers with WGBS at 98% and TM at > 99% specificity, and methods for determining TF, were previously reported. A model to predict the presence of detectable cfDNA fragments for CRC adenocarcinomas of stages I, II, and III included TSA and microinvasion beyond the subserosa. Predictors were combined assuming a linear increase of cfDNA shedding with tumor size, with scaling factors depending on microinvasion. Model parameters were determined for 27 participants (7, 11, 9 for stages I, II, III, resp.) with WGBS and applied to 40 participants (12, 15, 13 for I, II, III, resp.) with TM assay and information on tumor size and microinvasion. Results: CRC detection at stages I/II/III was 33/46, 61/73, 57/74% for WGBS/TM. TF predicted detection with AUC = 97.6. The model predicted TF as TSA multiplied by 3.81*10−6 / mm2 for tumors that invaded beyond the subserosa (p < 0.001). This was 4.4x higher than estimates for tumors below the subserosa. The model trained on the WGBS assay predicted CRC detection in the TM assay with an AUC of 0.844. Conclusions: This model used TSA (number of tumor cells) and microinvasion (bloodstream access) to predict the fraction of CRC ctDNA fragments in blood without needing to account for stage. Tumors not penetrating the subserosa had low ctDNA shedding that likely limited detection. These findings may generalize to other cancer types, providing principles to predict ctDNA shedding and thus cancer detectability based on microinvasion and surface area. Clinical trial information: NCT02889978.