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Homozygote CRIM1 Variant Is Associated with Thiopurine-Induced

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Homozygote CRIM1 Variant Is Associated with Thiopurine-Induced Park et al. J Transl Med (2020) 18:265 https://doi.org/10.1186/s12967-020-02416-7 Journal of Translational Medicine RESEARCH Open Access Homozygote CRIM1 variant is associated with thiopurine-induced neutropenia in leukemic patients with both wildtype NUDT15 and TPMT Yoomi Park1†, Hyery Kim2†, Heewon Seo1,3, Jung Yoon Choi4,5, Youngeun Ma6, Sunmin Yun1, Byung‑Joo Min1, Myung‑Eui Seo1, Keon Hee Yoo7, Hyoung Jin Kang4,5, Ho Joon Im2* and Ju Han Kim1,8* Abstract Background: NUDT15 and TPMT variants are strong genetic determinants of thiopurine‑induced hematological tox‑ icity that results in therapeutic failure in pediatric acute lymphoblastic leukemia (ALL). However, many patients with both wild‑type (WT) NUDT15 and TPMT still sufer from thiopurine toxicity and therapeutic failure. Methods: Whole‑exome sequencing was done for discovery (N 244) and replication (N 76) cohorts. Age‑ and sex‑adjusted multiple regression analyses of both WT patients were= performed to identify (=p < 0.01, N 188 for dis‑ covery) and validate (p < 0.05, N 52 for replication) candidate variants for the tolerated last‑cycle 6‑mercaptopurine= (6‑MP) dose intensity percentage= (DIP). Both independent and additive efects of the candidate variants on well‑ known NUDT15 and TPMT were evaluated by multigene prediction models. Results: Among the 12 candidate variants from the discovery phase, the rs3821169 variant of the gene encoding Cysteine‑Rich Transmembrane BMP Regulator 1 (CRIM1) was successfully replicated (p < 0.05). It showed high intereth‑ nic variability with an impressively high allele frequency in East Asians (T 0.255) compared to Africans (0.001), Ameri‑ cans (0.02), Europeans (0.009), and South Asians (0.05). Homozygote carriers= of the CRIM1 rs3821169 variant (N 12, 5%) showed signifcantly lower last‑cycle 6‑MP DIPs in the discovery, replication, and combined cohorts (p 0.025,= 0.013, and 0.001, respectively). The traditional two‑gene model (NUDT15 and TPMT) for predicting 6‑MP DIP= < 25% was outperformed by the three‑gene model that included CRIM1, in terms of the area under the receiver operating characteristic curve (0.734 vs. 0.665), prediction accuracy (0.759 vs. 0.756), sensitivity (0.636 vs. 0.523), positive predic‑ tive value (0.315 vs. 0.288), and negative predictive value (0.931 vs. 0.913). Conclusions: The CRIM1 rs3821169 variant is suggested to be an independent and/or additive genetic determinant of thiopurine toxicity beyond NUDT15 and TPMT in pediatric ALL. Keywords: CRIM1, NUDT15, TPMT, 6‑Mercaptopurine, Toxicity, Acute lymphoblastic leukemia *Correspondence: [email protected]; [email protected] †Yoomi Park and Hyery Kim contributed equally to this work 1 Division of Biomedical Informatics, Seoul National University Biomedical Informatics (SNUBI), Seoul National University College of Medicine, 101 Daehak‑ro, Jongno‑gu, Seoul 03080, South Korea 2 Department of Pediatrics, Asan Medical Center Children’s Hospital, University of Ulsan College of Medicine, 88, Olympic‑ro 43‑gil, Songpa‑gu, Seoul 05505, South Korea Full list of author information is available at the end of the article © The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creat iveco mmons .org/licen ses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creat iveco mmons .org/publi cdoma in/ zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Park et al. J Transl Med (2020) 18:265 Page 2 of 13 Background Table 1 Clinical characteristics of pediatric acute Te associations of NUDT15 and TPMT genetic variants lymphoblastic leukemia (ALL) subjects who are normal metabolizers (NMs) for both NUDT15 and TPMT with 6-mercaptopurine (6-MP) intolerance have been very well established in pediatric acute lymphoblastic Characteristic Discovery Replication Combined leukemia (ALL). In European populations, about 50% of Number of 188 52 240 thiopurine-induced severe cytotoxic adverse reactions subjects such as severe neutropenia and leukopenia are explained Age, yearsa 6.9 4.5 7.4 4.5 7.0 4.5 ± ± ± by NUDT15 and TPMT genetic variants [1]. Te Clinical Sex Pharmacogenetics Implementation Consortium (CPIC) Male 108 29 137 [2] publishes practical evidence-based guidelines for the Female 80 23 103 clinical implications of 6-MP based on these two genes, Last‑cycle 6‑MP dose, mg/m2/day supporting the implementation of pharmacogenetic test- 10 8.68 1.5 (3) 6.29 2.2 (5) 7.19 2.2 (8) ≤ ± ± ± ing in routine clinical practice [3, 4]. > 10 and 15 13.89 NA (1) 13.21 1.8 (3) 13.38 1.5 (4) ≤ ± ± ± Currently, 6-MP dose is clinically titrated based on the > 15 and 25 18.52 3.4 (5) 22.13 1.3 (4) 20.12 3.1 (9) ≤ ± ± ± known risk variants of TPMT or NUDT15. However, a > 25 and 35 29.95 3.3 (16) 30.49 0.8 (4) 30.06 3.0 (20) ≤ ± ± ± substantial proportion of leukemia patients who have no > 35 and 45 39.54 3.6 (8) 40.18 2.6 (5) 39.79 3.1 (13) ≤ ± ± ± genetic variation in NUDT15 or TPMT still sufer from > 45 and 60 52.71 4.0 (41) 54.84 2.8 (5) 52.94 3.9 (46) ≤ ± ± ± life-threatening toxicity, which may result in dose reduc- > 60 and 80 70.79 6.0 (55) 69.35 5.1 (10) 70.57 5.8 (65) ≤ ± ± ± tion and/or discontinuation of 6-MP and resultant thera- > 80 and 100 90.87 5.9 (35) 85.98 5.0 (8) 89.96 6.0 (43)* ≤ ± ± ± peutic failure and relapse. Terefore, further discovery > 100 112.67 16.6 122.66 23.3 (8) 115.17 18.7 (32) of novel genetic variants other than NUDT15 and TPMT (24) ± ± ± variations is urgently needed for preventing 6-MP toxic- Total 68.44 27.6 59.99 38.2 (52) 66.61 30.3 (240) ± ± ± ity and improving pediatric ALL patient care. (188) Te present study aimed to identify novel genetic vari- Data are n, mean SD, or mean SD (N) values ± ± ations associated with the 6-MP intolerance in pediatric 6-MP 6-mercaptopurine, NA not available ALL patients who carry both wild-type (WT) NUDT15 p values are for t-tests or 2 tests as appropriate. *p < 0.05 and TPMT by using whole-exome sequencing (WES) a Data for age were not available for one subject technology. We identifed and systematically evaluated the deterministic efects of novel candidate variants on a clinically important hematological toxicity indicator: the clinical endpoint. Te recorded 6-MP dose per square last-cycle 6-MP dose intensity percentage (DIP) tolerated meter of the body surface over a 12-week cycle was used by pediatric ALL patients. to defne the actual administered dose as a percentage of the planned dose as the last-cycle DIP. Since East Asian Methods ancestry requires signifcantly lower 6-MP dose inten- Subjects sity compared to the other ethnic groups [5], patients A total of 320 Korean pediatric ALL patients receiving who require less than 25% of the protocol planned dose 6-MP treatment during maintenance therapy include were classifed as MP-intolerant groups [6]. We have pre- viously presented a detailed description of the subjects discovery (N = 244) and replication (N = 76) cohorts, recruited from two teaching hospitals [Seoul National and a summary of the measurements [7]. Te present University Hospital (SNUH) and Asan Medical Center study was approved by the SNUH, AMC, and SMC insti- (AMC)] and three teaching hospitals [SNUH, AMC, tutional review boards. Written informed consent was and Samsung Medical Center (SMC)] located in Seoul, obtained from each participant. Korea, respectively (Table 1). Te discovery cohort was retrospectively collected and sequenced before February Whole‑exome sequencing and primary data analysis 2018, while the replication cohort was subsequently col- WES data obtained from the 320 pediatric ALL patients lected and sequenced from October 2018 to November were analyzed in a bioinformatics pipeline as we have 2019. All of the selected individuals conformed with the described previously [7]. Two missense NUDT15 vari- exclusion criteria (i.e., relapse of the disease, stem cell ants with no ofcially designated star alleles were transplantation, Burkitt’s lymphoma, mixed phenotype confrmed using Sanger sequencing, and false posi- acute leukemia, infant ALL, or very high risk of ALL). tive variant calls were removed in the further analysis. Te hematological toxicity was estimated based on the According to the CPIC guideline updated in February measurement of the tolerated last-cycle 6-MP DIP as the 2019, where the activity of NUDT15*9 for 6-MP was changed from ‘uncertain’ to ‘no function’ [1], one patient Park et al. J Transl Med (2020) 18:265 Page 3 of 13 was reclassifed as a poor metabolizer of NUDT15. Te Subsequent genotyping and validation present study analyzed the 240 normal metabolizers (188 To confrm the genotype calls of the fnal candidate vari- and 52 in the discovery and replication cohorts, respec- ant, rs3821169, we performed SNPtype (Fluidigm, San tively) of both NUDT15 and TPMT according to their Francisco, CA) assays for 118 subjects with 2 control star-allele genotypes.
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