Wang et al. Clinical Epigenetics (2020) 12:132 https://doi.org/10.1186/s13148-020-00923-4 RESEARCH Open Access Chidamide, decitabine, cytarabine, aclarubicin, and granulocyte colony- stimulating factor (CDCAG) in patients with relapsed/refractory acute myeloid leukemia: a single-arm, phase 1/2 study Lixin Wang1,2†, Jianmin Luo3†, Guofeng Chen4,5,6†, Meiyun Fang7, Xudong Wei8, Yinghua Li9, Zhuogang Liu10, Yin Zhang11, Sujun Gao12, Jianliang Shen2, Xin Wang13, Xiaoning Gao6, Wei Zhou5, Yigai Ma14, Hui Liu15, Xinquan Li16, Linhua Yang17, Kai Sun11 and Li Yu1,6* Abstract Background: Epigenetic mechanisms play an important role in the chemoresistance of acute myeloid leukemia (AML). The clinical response to epigenetic modifier-based chemotherapy in patients with relapsed/refractory AML (r/ r AML) is unclear. This multicenter clinical trial evaluated the safety and efficacy of epigenetic modifiers (chidamide and decitabine) in combination with aclarubicin, cytarabine, and granulocyte colony-stimulating factor (G-CSF) in patients with r/r AML. Results: Adult patients with r/r AML were treated with chidamide, decitabine, cytarabine, aclarubicin, and G-CSF (CDCAG). The primary measures were overall response (OR), overall survival (OS), and safety. Next-generation sequencing was performed to analyze the correlation between gene mutations and response. A total of 93 patients with r/r AML were enrolled. Overall, 24 patients had a complete remission (CR) and 19 patients achieved CR with incomplete blood count recovery (CRi). The overall response rate (ORR) was 46.2%. The overall survival of these 43 patients who achieved CR/CRi was significantly longer than that of patients who failed to achieve remission (563 vs 152 days, P < 0.0001). Of the patients with mutations in epigenetic and transcription factor-related genes, but without internal tandem duplications in FMS-like tyrosine kinase3 (FLT3-ITDs), 55.6% achieved CR/CRi, whereas the ORR was 28.2% for patients with mutations in other genes. (Continued on next page) * Correspondence: [email protected] †Lixin Wang, Jianmin Luo and Guofeng Chen contributed equally to this work. 1Department of Hematology-Oncology, International Cancer Center, Shenzhen University General Hospital, Shenzhen University Health Science Center, Shenzhen, China 6Department of Hematology, Chinese PLA General Hospital, 28 Fuxing Road, Beijing 100853, China Full list of author information is available at the end of the article © The Author(s). 2020 Open Access 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://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Wang et al. Clinical Epigenetics (2020) 12:132 Page 2 of 11 (Continued from previous page) Conclusions: The CDCAG regimen was well tolerated and effective in r/r AML. Patients with epigenetic and transcription factor-related gene mutations, but without FLT3-ITD mutations, may benefit from this regimen. Trial registration: Clinical Trials, NCT02886559. Registered 01 September 2016 Keywords: Relapsed/refractory acute myeloid leukemia, Next-generation sequencing, Histone deacetylase inhibitor, DNA methyltransferase inhibitor Introduction Results Approximately 30% of acute myeloid leukemia (AML) Patient characteristics cases will be classified as refractory AML due to failure A total of 93 patients, median age 40 years (range, 18– of induction chemotherapy. Additionally, more than 50% 60 years), with primary refractory AML (n = 37, 39.8%), of patients who achieve complete remission (CR) will early relapsed AML (n = 38, 40.9%), or late relapsed eventually relapse [1]. Therefore, the majority of patients AML (n = 18, 19.4%) were enrolled in the study. Among with AML will eventually be classified as refractory or the 56 patients with relapsed AML, 40 were experien- relapsed AML (r/r AML). The prognosis for r/r AML re- cing their first relapse, 15 were in their second relapse, mains dismal despite significant effort devoted to the de- and one was experiencing a third relapse. Patients with velopment of novel single-agent drugs and the design of relapsed AML had a median remission duration before new combination regimens. relapse of 7.6 months (range, 1–51 months). Sixty-nine Resistance to multiple chemotherapeutic agents is a (74.2%) patients had received at least three prior treat- common clinical problem encountered in r/r AML treat- ment regimens, 52 (55.9%) patients had received at least ment [2]. Accumulating research has demonstrated the five prior treatment regimens, and 36 (38.7%) patients importance of epigenetic modification in the pathogen- had received at least seven prior treatment regimens. esis of chemoresistance. DNA methylation and histone Seventeen (18.3%) patients had received at least one acetylation are the most common epigenetic changes hypomethylating agent, and two patients (2.2%) had pre- and can be pharmacologically reversed by DNA methyl- viously undergone allogeneic hematopoietic stem cell transferase (DNMT) inhibitors or histone deacetylase transplantation (allo-HSCT). Patients had received a me- (HDAC) inhibitors. Recent studies have shown that deci- dian of five (1–17) therapeutic cycles prior to enrollment tabine, a DNMT inhibitor, can increase the chemosensi- in this study. Patients were enrolled at a median of 8.8 tivity of several leukemic [3] and solid tumor cells [4, 5]. months (1.3–63.8 months) after initial diagnosis. Patient Furthermore, the addition of HDAC inhibitors, such as baseline characteristics are summarized in Table 1. chidamide or panobinostat, can enhance decitabine’s chemosensitization and cytotoxicity effects on leukemia Clinical responses cells when combined with conventional chemotherapy – Overall, 24/93 (25.8%) patients achieved CR according [6 9]. to the International Working Group (IWG) criteria and The standard dose of CAG regimen, consisting of low- an additional 19/93 (20.4%) patients achieved CR with dose cytarabine, aclarubicin, and granulocyte colony- incomplete blood count recovery (CRi), with an overall stimulating factor (G-CSF), led to a CR rate of 50% with response rate (ORR) of 46.2% (Table 2). Among the 43 well-tolerated toxicity in elderly patients, and the com- patients who achieved CR/CRi, the median duration of bination with decitabine increased the CR rate to 64.7% – leukemia-free survival was 259 days [95% confidence [10 12]. In this regimen, aclarubicin, an anthracycline interval (CI) 215—not available]. Of the 17 patients who topoisomerase II inhibitor, was recently reported to in- received prior hypomethylating agents, nine (52.9%) duce histone eviction in genomic regions and caused im- achieved CR/CRi. portant epigenetic changes [13, 14]. Therefore, exploring The median overall survival (OS) was 266 days (95% CI whether the addition of epigenetic modifiers, such as 235–398 days) with an estimated 6-month OS rate of decitabine and further the HDAC inhibitors, to the CAG 67.2% (95% CI 57.8–78.0%) and a 1-year OS of 36.9% regimen could exert a clinical benefit in r/r AML pa- (95% CI 26.6–51.1%, Fig. 1a). For the 43 patients who tients is of great significance. In this study, we designed achieved CR/CRi, the 6-month relapse-free survival (RFS) a regimen that included chidamide, decitabine, cytara- rate was 66.2% (95% CI 52.7–83.2%), the 1-year RFS rate bine, aclarubicin, and G-CSF (the CDCAG regimen) for was 40.7% (95% CI 26.0–63.9%), and the median RFS was the treatment of patients with r/r AML. We then con- 259 days (95% CI 185–NA; Fig. 1b). Patients achieving ducted a phase I/II study to evaluate the safety and effi- CR/CRi showed a significant improvement in median OS cacy of this regimen. compared with that of non-responders (563 vs 152 days, P Wang et al. Clinical Epigenetics (2020) 12:132 Page 3 of 11 Table 1 Patient demographics and baseline characteristics Table 1 Patient demographics and baseline characteristics (N =93) (N =93)(Continued) $ Characteristic Value$ Characteristic Value Age, years 40.0 ± 12.4 4 12 (13.6) Sex, no. (%) 5 3 (3.4) Male 50 (53.8) Abbreviations: BM bone marrow, HB hemoglobin, WBC white blood cell count, PLT platelets, ECOG PS Eastern Cooperative Oncology Group performance Female 43 (46.2) score, FAB French-American-British, CEBPA-dm CEPBA double mutation $Descriptive statistics were presented as the mean ± standard deviation (mean BM blasts, % 0.4 ± 0.3 ± SD) for continuous data and as numbers and percentages for HB, g/dL 91.0 ± 25.3 dichotomous/categorical data *Measured in 88 patients who underwent gene mutation detection 9 WBC, × 10 /L 9.2 ± 15.9 #Complex karyotype was defined as ≥ 3 clonal chromosomal abnormalities PLT, × 109/L 69.5 ± 66.4 < 0.0001; Fig. 1c). However, the differences of ORR and ECOG PS, no. (%) OS among patients with primary refractory AML, those in 0 34 (36.6) their first relapse, and those who had experienced multiple 1 42 (45.2) relapses were not statistically significant (data not shown). 2 16 (17.2) Of the 43 patients who achieved CR/CRi, eight pa- 3 1 (1.1) tients underwent allo-HSCT treatment.