Abstract 2872: Acquisition of drug resistance mutations during chemotherapy treatment in pediatric acute lymphoblastic leukemia
Authors: Li Benshang , Brady Samuel W. , Ma Xiaotu , Shen Shuhong , Zhang Yingchi , Li Yongjin , Liu Yu , Wang Ningling , Flasch Diane , Myers Matthew , Mulder Heather , Ding Lixia , Lu Yanling , Tian Liqing , Hagiwara Kohei , Xu Ke , Sioson Edgar , Wang Tianyi , Yang Liu , Zhao Jie , Zhang Hui , Shao Ying , Sun Hongye , Sun Lele , Cai Jiaoyang , Lin Ting-Nien , Du Lijuan , Yang Fan , Rusch Michael , Edmonson Michael , Easton John , Zhu Xiaofan , Zhang Jingliao , Cheng Cheng , Raphael Benjamin , Tang Jingyan , Downing James , Zhou Bin-Bing , Pui Ching-Hon , Yang Jun , Zhang Jinghui 
Published: August 2019

Abstract
Acute lymphoblastic leukemia (ALL) is a leading cause of cancer-associated death in children. To study the mechanisms of drug resistance in ALL, we performed whole-genome sequencing of diagnosis-relapse-germline trios from 103 Chinese patients and ultra-deep sequencing of 208 serial bone marrow samples from 17 of them. Relapse-specific somatic alterations were enriched in 12 genes (NR3C1, NR3C2, TP53, NT5C2, FPGS, CREBBP, MSH2, MSH6, PMS2, WHSC1, PRPS1, and PRPS2), which were predominantly involved in response to thiopurines, glucocorticoids, methotrexate, and other drugs. Four lines of evidence indicate that these resistance mutations frequently developed during treatment, rather than pre-existing at diagnosis. First, two novel, relapse-specific mutational signatures (novel signatures 1 and 2), most likely caused by chemotherapeutic regimens, were detected in 15% and 14% of relapsed cases, respectively. Drug resistance mutations frequently appeared at novel signature-associated trinucleotide contexts, indicating that chemotherapy may directly cause drug resistance mutations in ALL. The signatures were validated in NCI TARGET relapsed ALL samples, 2% and 23% of which harbored novel signatures 1 and 2, respectively. The varying signature prevalence between cohorts may reflect treatment differences. The novel signatures were not detected in &gt;2,000 adult cancers from the PCAWG study. Novel signature 1 induced C&gt;G transversions, particularly at GCC and TCT trinucleotides, and showed transcription-strand bias indicating guanine adducts. Novel signature 2 favored C&gt;T and C&gt;G mutations at CCG, and correlated with relapse-specific dinucleotide variants and structural variants, indicating an agent causing multiple mutation types. The drugs inducing these novel signatures are being explored in vitro. Second, mathematical modeling using growth curves of drug-resistant ALL indicated that drug resistance mutations occur, in some cases, long after diagnosis, during active treatment. Third, some patients acquired multiple drug resistance mutations sequentially through successive relapses, a finding inconsistent with their pre-existence at diagnosis. Indeed, 20% of relapses had multiple drug resistance mutations targeting different drug classes. Fourth, most relapsed ALLs derived from a subclone detected at diagnosis, which then evolved additional mutations, including drug resistance mutations, not detectable at diagnosis using 2000X targeted sequencing. Drug resistance mutations were often subclonal at relapse, suggesting later appearance. Together these data indicate that fully drug-resistant clones may not necessarily pre-exist at diagnosis in ALL, but may be acquired later during treatment. Thus, early intensive or targeted treatment strategies in slow responders may forestall the subsequent development of drug resistance mutations.
Citation Format: Benshang Li, Samuel W. Brady, Xiaotu Ma, Shuhong Shen, Yingchi Zhang, Yongjin Li, Yu Liu, Ningling Wang, Diane Flasch, Matthew Myers, Heather Mulder, Lixia Ding, Yanling Lu, Liqing Tian, Kohei Hagiwara, Ke Xu, Edgar Sioson, Tianyi Wang, Liu Yang, Jie Zhao, Hui Zhang, Ying Shao, Hongye Sun, Lele Sun, Jiaoyang Cai, Ting-Nien Lin, Lijuan Du, Fan Yang, Michael Rusch, Michael Edmonson, John Easton, Xiaofan Zhu, Jingliao Zhang, Cheng Cheng, Benjamin Raphael, Jingyan Tang, James Downing, Bin-Bing Zhou, Ching-Hon Pui, Jun Yang, Jinghui Zhang. Acquisition of drug resistance mutations during chemotherapy treatment in pediatric acute lymphoblastic leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2872.