Abstract A72: Mutation signature associated with colorectal cancer prognosis identified by targeted next-generation sequencing
Authors: Lee Dae-Won , Cha Yongjun , Han Sae-Won , Lee Si-Hyun , Kim Hwang-Phill , Lyu Jaemyun , Han Hyojun , Kim Hyoki , Jang Hoon , Bang Duhee , Huh Iksoo , Park Taesung , Bae Jeong Mo , Won Jae-Kyung , Jeong Seung-Yong , Park Kyu Joo , Kang Gyeong Hoon , Kim Tae-You 
Published: May 2016

Background

Although genomic data of CRC continue to accumulate, only limited information on prognostic role of the alterations have been reported. We have analyzed the prognostic impact of the key mutations in CRC using next generation sequencing technology.

Methods

We selected 40 genes from 5 critical pathways (WNT, TGF-B, PI3K, RTK-RAS and P53) of CRC based on TCGA data. Homogenous population of CRC patients were used to investigate the prognostic implication: 188 stage III or high-risk stage II CRC patients treated with curative surgery followed by adjuvant 5-fluorouracil/ leucovorin/oxaliplatin (FOLFOX) chemotherapy. Archival tissue from Seoul National University Hospital Tumor Bank was used.

The target (size 109kb) enrichment process was proceeded base on in-solution hybridization with biotinylated probes. The captured library was amplified and sequenced using Hiseq 2500 (Illumina, USA). Sequencing data was aligned to GRCh37 and variant call and somatic analysis processes was performed by VarScan2 and were annotated with ANNOVAR.

Using somatic nonsynonymous mutation data, we selected mutant gene signature which is associated with prognosis. We used binary collapsing method in a forward stepwise selection method to combine mutated genes. Test statistics were obtained from Cox-proportional hazard model. Two-sided p-values of less than 0.05 were considered statistically significant.

Results

Among a total of 188 patients, 63 had tumor in proximal location and 125 had tumor in distal location. Tumor stage was high-risk stage II in 21 patients and III in 167 patients. Average coverage of the total samples was 417X (414X tumor and 420X normal mucosa). Mutation frequencies were similar with the TCGA data except for NRAS and DKK2, having lower frequency in the present study. During a median follow-up duration of 58 months, 45 relapse and 23 death events have occurred. Alteration in individual gene or pathway did not have prognostic significance in terms of disease free survival (DFS) and overall survival (OS). By using binary collapsing method, we identified a 4 gene signature comprising ACVR1B, ERBB2, LRP5, and KRAS. 89 patients (47.3%) had 1 or more mutation in these 4 genes. 4 gene mutant group had worse DFS and OS compared with 4 gene wild type group. 3-year DFS and 5-year OS was 72.8% and 82.5% in mutant group compared with 88.7% and 96.0% in wild type group (p-values 0.004 and 0.001, respectively). Multivariate analysis revealed 4 gene signature as an independent negative prognostic factor of DFS and OS (adjusted hazard ratios 2.48 and 4.26, respectively).

Conclusion

Mutations in the 4 genes (ACVR1B, ERBB2, LRP5, and KRAS) were associated with poor prognosis in CRC patients treated with surgery followed by adjuvant FOLFOX chemotherapy. Validation study in an independent cohort is currently underway.

Citation Format: Dae-Won Lee, Yongjun Cha, Sae-Won Han, Si-Hyun Lee, Hwang-Phill Kim, Jaemyun Lyu, Hyojun Han, Hyoki Kim, Hoon Jang, Duhee Bang, Iksoo Huh, Taesung Park, Jeong Mo Bae, Jae-Kyung Won, Seung-Yong Jeong, Kyu Joo Park, Gyeong Hoon Kang, Tae-You Kim. Mutation signature associated with colorectal cancer prognosis identified by targeted next-generation sequencing. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A72.