Higher neutrophil counts are associated with successful tyrosine kinase inhibitor discontinuation in patients with chronic myeloid leukemia

ABSTRACT

Objective

Treatment-free remission (TFR) after tyrosine kinase inhibitor (TKI) discontinuation at the first attempt is a therapeutic goal for patients with chronic phase chronic myeloid leukemia (CML-CP). However, it remains unclear whether discontinuation of TKIs at a second or subsequent attempt can be performed safely.

Patients and method

Here, we report a 72-year-old man diagnosed with CML-CP. He achieved TFR successfully after TKI discontinuation at the third attempt. Before discontinuation, the patient received imatinib, nilotinib, and finally nilotinib. His neutrophil count at the third attempt was higher than after the second attempt. We also performed a retrospective investigation of 53 patients who discontinued TKIs on the first or subsequent attempts.

Results

Overall, 64 TKI discontinuations were documented (first, 53; second, ten; third, one). We found that a higher neutrophil count at the time of TKI discontinuation (>2439/μL; hazard ratio, 0.325; 95% confidence interval, 0.137–0.772; p = 0.011) was associated independently with lower rates of molecular relapse.

Conclusion

We report a case of a patient who successfully achieved third attempt TKI discontinuation and, an increased neutrophil percentage may reflect stronger antitumor immune responses in patients with CML-CP.

KEYWORDS:

• Chronic myeloid leukemia
• ABL1 tyrosine kinase inhibitor
• tyrosine kinase inhibitor discontinuation
• third attempt tyrosine kinase inhibitor discontinuation
• higher neutrophil counts

Introduction

Recently, treatment-free remission (TFR) has become a therapeutic goal for patients with chronic phase chronic myeloid leukemia (CML-CP); indeed, several tyrosine kinase inhibitor (TKI) discontinuation trials show that approximately half of the patients who achieve a durable deep molecular response (DMR) also achieve TFR [1–4]. Unfortunately, approximately half of patients who attempt TKI discontinuation experience molecular relapse. Generally, they resume TKIs, after which the majority re-achieve molecular remission. It is still unclear whether such patients can discontinue TKIs safely at a second or subsequent attempt [5]. Here, we report a rare case of a patient who successfully achieved TFR after TKI discontinuation at the third attempt.

Case presentation

The patient was a 72-year-old man who was diagnosed with CML-CP, classified as low risk according to the Sokal risk score, in August, 2005 (i.e. aged 56). Imatinib was initiated at a standard dose (400 mg/day), and the patient achieved a major molecular response (MMR) after 9 months (May, 2006). He achieved a DMR (BCR-ABL1 transcript <5 copies/0.5 μg RNA, as measured by the transcription-mediated amplification method (TMA)) after 3 years (May, 2009). In January 2013, approximately 8 years after the CML diagnosis and 4 years after the achievement of DMR, the patient decided to discontinue imatinib.

However, 2 months later, he lost the MMR, and so nilotinib (600 mg) was initiated. After 2 months, he achieved DMR again (confirmed by the TMA method). After a durable DMR for 2 years, nilotinib was discontinued in May, 2015. Unfortunately, the patient lost MMR 2 months later, and nilotinib was resumed (600 mg). MR4.0 (on the international scale (IS), measured using the ODK-1201 kit; Otsuka Pharmaceutical Co., Tokyo, Japan) [6] was achieved in October, 2015, and MR4.5 was achieved in January, 2016, with undetectable minimal residual disease (UMRD), measured by polymerase chain reaction, achieved in February, 2017. After a 2 year durable UMRD, nilotinib was discontinued again in February, 2019. No molecular relapse occurred (durable MR4.5) after 18 months (Figure 1(A)). Notably, the average neutrophil count (before 6 months) at the time of second nilotinib discontinuation (5169 ± 376/μL; third TKI discontinuation) was significantly higher than that at first nilotinib discontinuation (2934 ± 445/μL; second TKI discontinuation; p = 0.002) (Mann–Whitney U test, Figure 1(B, C)).

Figure 1. Treatment course of the patient (A). Average white blood cell count (B), neutrophil count (C), and lymphocyte count (D) at the first, second, and third discontinuations of tyrosine kinase inhibitors (before 6 months TKI discontinuation). Abbreviation: MMR, major molecular response.

T cells or natural killer cells within the host immune system may regulate CML cells to prevent progression or relapse after TKI discontinuation [1]. However, it is difficult to confirm the immunological dynamics in patients with CML-CP in a general clinical setting. In the present patient, the lymphocyte count at the third TKI discontinuation was no different from that at the first or second TKI discontinuations (Figure 1(D)); however, the neutrophil count increased. Therefore, we focused on neutrophils.

Previously, we reported a retrospective analysis of patients with CML-CP who made multiple attempts at TKI discontinuation, and we re-analyzed the data about neutrophil count at TKI discontinuation [7]. The study protocol was approved by the institutional review board of Saga University (2021-01-R-04). All procedures involving human participants were conducted in accordance with the ethical standards of institutional and/or national research committees, and the Declaration of Helsinki. Informed consent was obtained by the opt-out method; information on the research, including the aims of the use of specimens and the opportunity to opt-out, was made public, and no patients objected to this study.

Response criteria

Response criteria were defined according to BCR–ABL1 mRNA transcripts adjusted to the international scale (IS), and/or using the transcription-mediated amplification (TMA) method. A major molecular response (MMR) was defined as ≤0.1% (IS) or a BCR-ABL1 transcripts level of <50 copies/0.5 μg RNA (TMA). MR4.0 was defined as ≤0.01% (IS) or undetectable BCR-ABL1 (TMA). MR4.5 was defined as ≤0.0032% (IS). The TMA method for quantification of BCR-ABL1 transcripts was the only method approved by the national insurance body of Japan until June 2015.

Characteristics of the patients who discontinued TKIs

Sixty four TKI discontinuations were documented (53 first attempt, ten second attempt, and one third attempt: this case) in 53 TKI stop patients. The median age at the time of TKI discontinuation was 60 years (interquartile range [IQR]: 55–69 years); 31 patients were male, and 22 were female; and the median TKI treatment duration and follow-up time were 61.8 months (IQR: 43.1–102.7 months) and 48.7 months (IQR: 28.6–69.2 months), respectively. The median white blood cell, neutrophil, and lymphocyte counts were 5150/μL (IQR, 4200–6225), 2439 (IQR, 2025–3771), and 1978 (IQR, 1466–2741), respectively. The variables (continuous variables were dichotomized at the median values) that affected TFR were assessed using a Cox proportional hazards model and the log-rank test. All statistical analyses were performed using the EZR software package (Saitama Medical Center, Jichi Medical University) [8].

Higher neutrophil counts at the time of TKI discontinuation are associated with lower rates of molecular relapse in patients with CML-CP

Univariate analysis of molecular relapse after TKI discontinuation revealed that a higher white blood cell count (≥5150/μL; hazard ratio (HR), 0.446; 95% confidence interval (CI), 0.215–0.925; p = 0.030) and a higher neutrophil count (≥2439/μL; HR, 0.348; 95% CI, 0.166–0.737; p = 0.006) were associated with lower rates of molecular relapse in patients with CML-CP; meanwhile, there was no association with the lymphocyte count (≥1978/μL; HR, 0.809; 95% CI, 0.402–1.627; p = 0.552). Previously, we reported that the male sex might be associated with a lower risk of molecular relapse in patients with CML-CP [7]. In the present study, none of the examined factors (age, sex, Sokal risk score, BCR-ABL1 transcript levels, TKIs before discontinuation, TKI treatment duration, DMR duration before discontinuation, and TKI discontinuation phase (first attempt or subsequent attempt)) were associated with TFR in patients with CML-CP (Table 1). Multivariate analysis identified a higher neutrophil count (≥2439/μL) as being associated with successful achievement of TFR in patients with CML-CP (HR, 0.325; 95% CI, 0.137–0.772; p = 0.011) (Table 2).

Table 1. Univariate analysis to identify clinical characteristics responsible for molecular relapse.

Variable	n	Estimated treatment-free remission at 12 months	P-value	Hazard ratio (95% CI)	P-value
		(%, 95% CI)
Age (years)
<60	26	60.2 (38.7–76.3)		1
≥60	27	44.4 (25.6–61.7)	0.257	1.561 (0.716–3.403)	0.263
Sex
Female	22	40.9 (20.9–60.1)		1
Male	31	60.4 (40.9–75.3)	0.225	0.625 (0.290–1.350)	0.232
Sokal risk score
High	7	71.4 (25.8–92.0)		1
Intermediate	16	43.8 (19.8–65.8)		2.455 (0.530–11.390)	0.251
Low	27	50.4 (30.3–67.5)	0.701	1.948 (0.439–8.653)	0.381
Missing	3
BCR-ABL1 transcript at TKI discontinuation
MR^4.0	29	44.8 (26.5–61.6)		1
MR^4.5	13	53.8 (24.8–76.0)		0.711 (0.280–1.803)	0.472
UMRD	22	56.4 (32.8–74.5)	0.446	0.610 (0.271–1.370)	0.231
TKI before discontinuation
Imatinib	10	46.3 (31.0–60.3)		1
Dasatinib	44	50.0 (18.4–75.3)		0.872 (0.355–2.145)	0.766
Nilotinib	10	70.0 (32.9–89.2)	0.428	0.448 (0.112–1.792)	0.256
TKI treatment duration before discontinuation
<56.0 months	26	50.0 (29.9–67.2)		1
≥56.0 months	38	51.1 (34.1–65.8)	0.936	0.972 (0.479–1.969)	0.936
DMR duration
<38.4 months	32	45.7 (27.8–61.8)		1
≥38.4 months	32	55.8 (37.0–71.0)	0.375	0.732 (0.365–1.467)	0.379
TKI discontinuation phase
First	53	52.2 (37.9–64.6)		1
Subsequent	11	43.6 (14.7–69.9)	0.791	1.128 (0.462–2.751)	0.792
White blood cell count at TKI discontinuation
<5150/μL	32	37.5 (21.3–53.7)		1
≥5150/μL	32	64.8 (45.5–78.8)	0.025	0.446 (0.215–0.925)	0.030
Neutrophil count at TKI discontinuation
<2439/μL	32	34.4 (18.8–50.6)		1
≥2439/μL	32	67.8 (48.4–81.2)	0.004	0.348 (0.166–0.737)	0.006
Lymphocyte count at TKI discontinuation
<1978/μL	32	46.9 (29.1–62.8)		1
≥1978/μL	32	55.2 (36.3–70.5)	0.55	0.809 (0.402–1.627)	0.552

Abbreviations: CI, confidence interval; TKI, tyrosine kinase inhibitor; UMRD, undetectable minimal residual disease; DMR, deep molecular response.

Table 2. Multinivariate analysis to identify clinical characteristics responsible for molecular relapse.

Variable	n	Hazard ratio (95% CI)	P-value
Age (years)
<60	26	1
≥60	27	1.749 (0.789–3.879)	0.169
Sex
Female	22	1
Male	31	0.689 (0.318–1.493)	0.345
DMR duration
<38.4 months	32	1
≥38.4 months	32	0.560 (0.245–1.280)	0.169
Neutrophil count
<2439/μL	32	1
≥2439/μL	32	0.325 (0.137–0.772)	0.011

Abbreviation: DMR, deep molecular response; CI, confidence interval.

Neutrophil counts at the time of nilotinib discontinuation were higher than those of imatinib or dasatinib

In the present case, increased neutrophil counts were observed at the second nilotinib discontinuation. We hypothesized nilotinib administration may yield neutrophil elevation in patients with CML. In the 53 patients with CML-CP, neutrophil counts at the time of nilotinib discontinuation (median, 3698.55/μL) were significantly higher than those at the time of imatinib (2192.85/μL, p = 0.0156) or dasatinib (2394.70/μL, p = 0.0015) discontinuation (Figure 2).

Figure 2. Average neutrophil count during treatment with tyrosine kinase inhibitors (TKIs) (imatinib, dasatinib, and nilotinib).

Note: Kruskal-Wallis test with Bonferroni correction.

Discussion

Here, we report the rare case of a patient who achieved TFR after TKI discontinuation at the third attempt. We also show that a higher neutrophil count may be associated with the successful achievement of TFR in patients with CML-CP.

Previous TKI discontinuation trials show that longer TKI treatment duration [2,3], longer duration of DMR [3], and the depth of the molecular response [9,10] are favorable prognostic factors for TFR. In the present patient, successful achievement of TFR3 might be due to a longer DMR time (2–3 years) and a deeper molecular response when compared with TFR2 (MR4.0 to UMRD); this is consistent with the results of previous TKI discontinuation trials [2,3]. Thus, the third attempt at TKI discontinuation may be feasible, with results similar to those achieved after first and second attempts [2,3,5,7].

Host T cells and natural killer cells may regulate CML cells to prevent progression or relapse after TKI discontinuation [1]. In the present patient, the lymphocyte fraction at the time of third TKI discontinuation was no different from that at the first and second attempts; however, the neutrophil fraction increased. Traditionally, neutrophils play an antibacterial role; however, recent studies have identified tumor-associated neutrophils, which play a crucial role in cancer biology [11]. Increased neutrophil fraction possibly contributed successful achievement of TFR in the present case [12], and one possible favorable biomarker for TFR [13]. Hence, we are conducting a multicenter retrospective analysis (2021-12-R-04) to confirm the clinical impact of neutrophil counts on the successful achievement of TFR in patients with CML-CP.

Off-target effects of imatinib or nilotinib include inhibition of the TGF-β signaling pathway [14,15], leading to an increase in the number of mature neutrophils [16]. In our case, neutrophil counts at the time of nilotinib discontinuation were significantly higher than those at the time of imatinib or dasatinib discontinuation. These results indicate that long-term nilotinib treatment may strongly inhibit TGF-β signaling pathway.

In conclusion, we report a rare case of a patient who successfully achieved TKI discontinuation at the third attempt. We also show that long-term nilotinib treatment increases the neutrophil count, which may contribute to the successful achievement of TFR in patients with CML-CP.

Acknowledgements

HU, KK and SK designed study concept, done data collection and data analysis and wrote the manuscript. The other authors contributed to patient’s care. All authors approved the final version.

Disclosure statement

S.K. has received honoraria from Bristol-Myers Squibb, Novartis, Pfizer, and Otsuka Pharmaceuticals, and research funding from Bristol-Myers Squibb, Pfizer, Otsuka Pharmaceuticals, and Ohara Pharmaceuticals. The other authors declare no conflicts of interest.

Additional information

Funding

This work was supported by Grants-in-Aid for Scientific Research from the Shinnihon Foundation of Advanced Medical Treatment Research (to H.U.), the Japan Society for the Promotion of Science (21K16245 to H.U.), the Medical Research Encouragement Prize of the Japan Medical Association (to H.U.), and a Medical Research Grant from Takeda Science Foundation (to H.U.).