Response and resistance to cladribine in patients with advanced systemic mastocytosis: a registry-based analysis

We sought to evaluate the efficacy of the purine analogue cladribine in 79 patients with advanced systemic mastocytosis (AdvSM) using data from the ‘German Registry on Disorders of Eosinophils and Mast Cells (GREM)’. The overall response rate according to modified Valent criteria (46 evaluable patients) for first- (1L) and second-line (2L) cladribine treatment was 41% (12/29) and 35% (6/17, P = 0.690), respectively, and the median overall survival (OS, all patients evaluable) was 1.9 years (n = 48) and 1.2 years (n = 31; P = 0.311). Univariate and multivariable analyses of baseline and on-treatment parameters identified diagnosis of mast cell leukemia (hazard ratio [HR] 3.5, 95% confidence interval [CI, 1.3–9.1], P = 0.012), eosinophilia ≥ 1.5 × 109/L (HR 2.9 [CI 1.4–6.2], P = 0.006) and < 3 cycles of cladribine (HR 0.4 [CI 0.2–0.8], P = 0.008) as independent adverse prognostic parameters for OS. There was no impact of other laboratory (anemia, thrombocytopenia, serum tryptase) or genetic markers (mutations in SRSF2, ASXL1 or RUNX1) on OS. In consequence, none of the recently established prognostic scoring systems (MARS, IPSM, MAPS or GPSM) was predictive for OS. Modified Valent criteria were superior to a single factor-based response assessment (HR 2.9 [CI 1.3–6.6], P = 0.026). In conclusion, cladribine is effective in 1L and 2L treatment of AdvSM. Mast cell leukemia, eosinophilia, application of < 3 cycles and a lack of response are adverse prognostic markers. Supplementary Information The online version contains supplementary material available at 10.1007/s00277-023-05180-y.


Study population
All cladribine-treated patients (n = 79) from the GREM which were diagnosed between 2003 and 2021 were selected for this project, which is an updated and more detailed analysis of a comparative study between midostaurin and cladribine [23]. The diagnosis of SM was established according to the World Health Organization classification [1,[29][30][31]. All BM biopsies were evaluated by reference pathologists (H.-P.H., K.S.) of the European Competence Network on Mastocytosis (ECNM) [32]. The study design adhered to the tenets of the Declaration of Helsinki and was approved by the institutional review board of the Medical Faculty of Mannheim, Heidelberg University, Germany. Written informed consent was provided by all patients.

Treatment
The number of patients allowed separation of first-(1L) and second-line (2L) treatment. Prior treatment included midostaurin while subsequent treatment approaches included (individually or sequentially) midostaurin, avapritinib, acute myeloid leukemia-like intensive chemotherapy and, rarely, allogeneic stem cell transplantation. Treatment options with a potentially low disease-modifying impact (e.g. interferonalpha) or solely directed towards AHN (e.g. hydroxyurea, azacytidine) were not considered as 1L-or 2L-treatment.

Gene mutation analyses
Quantitative assessment of the KIT D816V expressed allele burden (EAB) was performed by allele-specific quantitative real-time reverse-transcriptase polymerase chain reaction (RT-qPCR) analysis on RNA/complementary DNA as previously described [33]. NGS analyses on DNA were performed through library preparation by the Access Array Technology (Fluidigm, San Francisco, CA) and sequencing on the MiSeq Instrument (Illumina, San Diego, CA). Gene mutations were annotated using the reference sequence of the Ensembl Transcript ID (Ensembl release 85: July 2016).

Prognostic scoring systems
The predictive value and clinical utility of several recently established prognostic scoring systems (MARS, International Prognostic Scoring System for AdvSM [IPSM-AdvSM], MAPS, and GPSM) was conducted according to published criteria [7,11,12,34]. Similarities and differences between the scores are given elsewhere. [11,30]

Response assessment
Response assessment according to modified Valent criteria [21] included regular monitoring of C-findings, serum tryptase and a BM biopsy within 2 months after the last applied course of cladribine. The reasons for not using the more recently established International Working Group-Myeloproliferative Neoplasms Research Treatment-ECNM (IWG-MRT-ECNM) criteria included: (i) the retrospective nature of our analysis did not allow to adequately address the complex IWG-MRT-ECNM criteria, (ii) the modified Valent response criteria were commonly used for response assessment of cladribine in prior studies. Molecular response was defined as KIT D816V expressed allele burden reduction ≥ 25% within 2 months after the last course. [7,23,33,35]

Statistical analyses
All statistical analyses considering clinical, laboratory and molecular parameters were obtained at the time of diagnosis/first referral to our center (initial parameters), treatment initiation with cladribine (baseline parameters) and at multiple time points during treatment (including time point for response assessment). The Mann-Whitney U-test was used to compare continuous variables and medians of distributions. Fisher's exact test was used for categorical variables. We retrospectively analyzed the OS (time of diagnosis/treatment initiation to the date of death/last visit) by using the Kaplan-Meier method with log-rank test for group comparisons/visualizations. Disease progression was defined as a shift to a more aggressive AdvSM subtype (secondary MCL or secondary acute myeloid leukemia [AML]). Duration of treatment was defined as the duration from initiation of cladribine to discontinuation for any reason. For the estimation of hazard ratios (HRs) and multivariable analysis, the Cox proportional hazard regression model was used. All variables that showed prognostic significance in univariate analyses were included in multivariable analyses. The first multivariable analysis was performed in an unmodified cohort of patients irrespective of prior or following treatment approaches (midostaurin, avapritinib, intensive chemotherapy and allogeneic stem cell transplantation); the second multivariable analysis was performed in a modified cohort in which patients with prior or following treatment approaches were either excluded or censored at the time of initiation of the next treatment line. P values of < 0.05 (two-sided) were considered as significant. Data management and statistical analyses were performed with SPSS (SPSS version 20.0; IBM Corporation, Armonk, NY) and GraphPad Prism software (version 8, GraphPad, La Jolla, CA, USA).  An expanded version of this table is given as Appendix Table 2 All First-line Second-line P

Treatment line Median
Years from cladribine initiation

Discussion
In historical cohorts of up to a maximum of 32 AdvSM patients [18,19,21], the ORR on cladribine according to (modified) Valent criteria [21,36] ranged between 50 and 100%. [20] Further interpretation on the impact of treatment with cladribine on progression-free (PFS), relapsefree (RFS), event-free (EFS), leukemia-free (LFS) and overall survival is limited because (i) most reports did not clearly differentiate between ISM and AdvSM, (ii) no report separated between 1L-and 2L-treatment and (iii) the definitions of PFS/RFS/EFS/LFS were not consistent between studies. In a registry-based cross-assessment, we recently reported an ORR (modified Valent criteria) of 35% in midostaurin-treated and 40% in cladribine-treated patients [23]. Notwithstanding, the OS on cladribine was significantly inferior to midostaurin in both 1L-and 2L-treatment cohorts. In the current report, we sought to provide a more detailed analysis on response rates on cladribine in 1L-and 2L-treatment, biomarkers indicating response and resistance and the association between ORR and OS. Multivariable analysis identified hypereosinophilia (> 1.5 × 10 9 /l), as marker of an AHN, diagnosis of MCL, and application < 3 cycles as adverse prognostic markers. This confirms a recent report from the Mayo Clinic registry on 22 cladribine-treated AdvSM patients indicating a diagnosis of an AHN (in addition to older age and absence of KIT D816V) as adverse prognostic markers for survival and is also in line with a previous publication on the poor prognostic impact of eosinophilia in SM [18,37]. Recent data also revealed that midostaurin was superior to cladribine in controlling AHN-associated myeloproliferation [23]. The application of ≥ 3 cycles was further associated with a higher ORR.
In a minority of patients (< 10%), cladribine was used for bridging the interval to the start of the midostaurin trial in 2009 and at later time points, it was used in a few patients for more rapid MC debulking with subsequent preplanned switch to midostaurin. Although myelosuppression became apparent in approximately 20% of patients, infectious complications were not noted as reasons for treatment discontinuation. In contrast to midostaurin, OS on cladribine was not influenced by cytopenias prior to treatment or additional somatic mutations in the S/A/R gene panel. Consequently, none of the prognostic scoring systems (MARS, IPSM, MAPS, GPSM) was predictive for OS. The reasons for this observation are unknown but may be explained at least in part by the fact that the scores more effectively identify low-risk patients on targeted treatment with midostaurin [23,27] or avapritinb [26] than on conventional chemotherapy with cladribine.
In contrast to the recent report from the Mayo Clinic, possibly due to the higher number of patients in our study, any response according to modified Valent criteria in 1L-but also 2L-treatment was associated with improved OS, thus confirming the usefulness of response assessment for guiding further treatment strategies. The data were underscored by the predictive superiority of modified Valent criteria versus a single factor-based response assessment. Although 2L patients presented with a higher disease burden, response and survival were not statistically different from 1L patients.
Recently reported propensity score weighted analyses on LFS/EFS and OS revealed superiority of midostaurin over cladribine and of avapritinib over best available treatment including midostaurin and cladribine [23][24][25]. However, we conclude that (i) cladribine remains a relevant option within the AdvSM treatment algorithm; its application in 1L-, 2Lor 3L-line locally depends on the approval status and availability of midostaurin and avapritinib; (ii) mast cell leukemia, eosinophilia, application of < 3 cycles and lack of response according to modified Valent criteria are adverse prognostic markers, and (iii) commonly used prognostic models for AdvSM are of limited value because of high mortality in low-and intermediate-risk patients.
The genetic and clinical complexity of AdvSM requires further prospective clinical trials to study the effects of KIT inhibitors in combination with simultaneous or intermittent use of other anti-neoplastic drugs, e.g. cladribine or hypomethylating agents. Such an approach may counteract the potential outgrowth of KIT D816V negative or multimutated subclones [38]. For patients with progression into secondary MCL or secondary AML, AML-like chemotherapy with or without subsequent allogeneic stem cell transplantation remains the most reasonable and potentially curative treatment options.