Correlation of Expression of CD25 in Hematopoietic Stem/Progenitor Cell Fraction of Bone Marrow Cells with Response to Tyrosine Kinase Inhibitors in Chronic Myelogenous Leukemia Patients
Authors: Karigane Daiki , Kasahara Hidenori , Sakurai Masatoshi , Matsuki Eri , Tozawa Keiichi , Koda Yuya , Toyama Takaaki , Kikuchi Taku , Kato Jun , Shimizu Takayuki , Mori Takehiko , Yatabe Yoko , Arai Tomoko , Mitsuhashi Takayuki , Murata Mitsuru , Kobayashi Hiroshi , Takubo Keiyo , Okamoto Shinichiro 
Published: October 2019

Abstract
Introduction: Tyrosine kinase inhibitors (TKIs) have dramatically improved the prognosis of chronic myelogenous leukemia (CML). The treatment with TKIs continues to improve the depth of response and overall survival of CML patients, but the life-long use of TKI is known to be associated with late complications such as cardiovascular events as well as heavy financial burden, and thus impairs the quality of life. To overcome these issues, many studies evaluating the possibility of TKI discontinuation have been ongoing worldwide. In order to achieve durable treatment-free remission, it is crucial to understand the dynamics of CML-leukemia initiating cells (LICs). We previously reported that CD25 was highly expressed in murine and human CML-LICs (Kobayashi CI et al., Blood, 2014). The aim of this study was to assess whether the proportion of CD25 positive cells in hematopoietic stem/progenitor cell fraction of bone marrow cells in CML patients treated with TKIs is associated with their molecular response and could serve as a novel surrogate marker to stop TKI therapy.
Methods: Bone marrow samples were obtained from patients with CML in chronic phase who were diagnosed and have been treated solely with TKIs at Keio University Hospital (Tokyo, Japan). This study was approved by the institutional ethical committee and informed consent was obtained from each patient. Both quantitative and qualitative PCR of BCR-ABL was performed using bone marrow mononuclear cells (BMMNCs). The proportion of CD25 positive cells in bone marrow hematopoietic stem/progenitor cell (HSPC; CD34+CD38-) fraction was evaluated by flow cytometry using FITC-labeled anti-CD34, PE- labeled anti-CD38 and APC-labeled anti-CD25 antibodies. The response to TKIs at the time of evaluation was determined according to the previous report (Yoshida C et al., Int J Clin Oncol, 2012): complete cytogenetic remission (CCyR) defined as Philadelphia chromosome undetectable and quantitative PCR copy numbers >731 among BMMNCs; major molecular remission (MMR) as quantitative PCR copy numbers ≤731, and complete molecular remission (CMR) as undetectable BCR-ABL by quantitative and qualitative PCR.
Results: Bone marrow samples obtained from 95 patients were evaluated (median age 53 years old; male/female, 67/28). Analysis was performed prior to TKI exposure in nine patients and under TKI therapy including 2nd generation TKI in 64 patients (imatinib, 22; dasatinib, 33; nilotinib, 9). Remaining 22 patients were treatment free because they enrolled in a clinical trial of TKI discontinuation. The proportion of CD25 positive cells in HSPC fraction significantly decreased in patients with prior TKI exposure relative to patients at diagnosis (n=86; Mean 4.2%, SD 7.0% vs n=9; Mean 22.4%, SD 11.3%, P<0.01). In addition, the proportion of CD25 positive cells in HSPC fraction significantly correlated the level of quantitative PCR (Figure, P<0.0001), and MMR was also dividable from CMR by the proportion of CD25 positive cells (MMR; n=29, Mean 4.6%, SD 4.8% vs CMR; n=49, Mean 2.6%, SD 2.5%, P<0.05). The expression of CD25 was still detected in the majority of patients who achieved CMR, including those who sustained CMR after the discontinuation of TKIs.
Conclusion: We confirmed that the expression of CD25 in HSPC fraction of CML patients was significantly correlated with the response to TKI therapy, and may serve as an asset to select patients who are likely to achieve durable treatment-free survival.
Figure Figure.

Disclosures
Karigane: Celgene: Honoraria. Sakurai:Celgene: Honoraria. Matsuki:Bristol-Myers Squibb: Honoraria; Celgene: Honoraria; Nippon Shinyaku: Honoraria. Kikuchi:Celgene: Honoraria; Takeda Pharmaceutical Company: Honoraria; Kyowa Hakko Kirin: Honoraria. Mitsuhashi:LSI Medience: Consultancy. Okamoto:Sumitomo Dainippon Pharma Co., Ltd.: Research Funding; Asahi Kasei Pharma Corp.: Research Funding; Astellas Pharma Inc.: Research Funding; Shionogi & Co., Ltd.: Research Funding; Nippon Shinyaku Co., Ltd.: Research Funding; Alexion Pharmaceuticals, Inc.: Research Funding; Toyama Chemical Co., Ltd.: Research Funding; Otsuka Pharmaceutical Co., Ltd.: Honoraria, Research Funding; Teijin Pharma Limited: Research Funding; Bristol-Myers Squibb K.K.: Honoraria, Research Funding; Kyowa Hakko Kirin Co., Ltd.: Research Funding; Eisai Co., Ltd.: Research Funding; Chugai Pharmaceutical Co., Ltd.: Research Funding; Pfizer Inc.: Honoraria, Research Funding; JCR Pharmaceuticals Co., Ltd.: Research Funding.