KIT D816V Mutation Burden Predicts Prognosis and Survival In Patients With Mastocytosis and Correlates With The WHO Type Of The Disease

Abstract

Mastocytosis is characterized by abnormal expansion and accumulation of neoplastic mast cells in one or more organ systems. Traditionally, mastocytosis is divided into cutaneous mastocytosis (CM) and systemic mastocytosis (SM). In most patients with SM, the transforming somatic mutation KIT D816V is detected. However, only few studies have quantified the KIT D816V allele burden in CM and SM. The aim of the present study was to quantify KIT D816V in various forms of mastocytosis and to correlate the allele burden of KIT D816V with the disease category, serum tryptase levels and clinical outcomes. KIT D816V was quantified in bone marrow (BM) and peripheral blood (PB) cells by a real-time PCR (qPCR) assay based on allele-specific primers. In addition, BM and PB cells were also examined for the presence of KIT D816V by melting curve analysis after PCR clamping in all patients. Overall, 225 DNA samples (BM, n=112; PB, n=113) from 107 patients with mastocytosis (females: n=57; males, n=50; median age 49 years; range 18-73 years) were analyzed. Based on WHO criteria, 14 patients had CM, 3 the provisional diagnosis of mastocytosis in the skin (MIS), 66 indolent SM (ISM), 6 smouldering SM (SSM), 7 aggressive SM (ASM), one mast cell leukemia (MCL) and 10 patients SM with an associated hematologic non-mast cell lineage disorder (SM-AHNMD). KIT D816V was found in in 76/107 patients (71%) by melting curve analysis after PCR clamping, and in 92/107 patients (86%) by qPCR (p<0.005). In paired BM and PB samples of 43 patients an excellent correlation of the KIT D816V burden with almost identical results was found (r=0.98, p<0.001). When examining the KIT D816V allele burden in KIT D816V+ patients (n=92) in various categories of the disease, significant differences were found between CM (median KIT D816V allele fraction: 0.042%), MIS (median: 0.084%), ISM (median: 0.286%), SSM (median: 3.012%), ASM (median: 9.346%) and SM-AHNMD (median: 3.761%) (p<0.001). Moreover, we found that the KIT D816V allele burden correlates significantly with the serum tryptase level in our patients (r=0.50, p<0.005). Consecutive studies revealed that the KIT D816V allele fraction is of prognostic significance concerning survival as determined by Cox regression (p=0.015). As assessed by Kaplan Meier estimates and log rank testing, patients with a KIT D816V allele burden of ≥2% were found to have a significantly shorter survival than those with an allele burden of less than 2% (p=0.001) (Figure 1). Thirty patients were evaluated at diagnosis and during the follow up. In untreated patients with stable disease, the KIT D816V allele burden remained within a constant range. By contrast, in patients with disease progression, an increase in the KIT D816V burden over time was detectable. In patients responding to cytoreductive agents (cladribine n=4; hydroxyurea n=1) a significant decrease in the median KIT D816V allele burden (by 91.6%) after therapy compared to pre-therapeutic samples was observed (p=0.027). In summary, our data show that qPCR is a highly sensitive approach for the detection and quantification of KIT D816V in patients with mastocytosis and that the KIT D816V mutation burden differs significantly among patients in different WHO subtypes. Moreover, the KIT D816V allele burden correlates with serum tryptase levels and is of prognostic significance concerning survival in patients with mastocytosis. Finally, quantification of KIT D816V may serve as follow up parameter useful for determining the natural course and treatment responses in patients with mastocytosis. We recommend that the KIT D816V mutation burden is included as a novel parameter in daily practice and clinical trials in advanced SM.Figure 1Overall survival of patients with KIT D816V+ mastocytosis. Patients were split into those with a KIT D816V allele burden of<2% and those with an allelic burden of ≥2%. Survival was estimated by the method of Kaplan and Meier (p=0.001).Figure 1. Overall survival of patients with KIT D816V+ mastocytosis. Patients were split into those with a KIT D816V allele burden of<2% and those with an allelic burden of ≥2%. Survival was estimated by the method of Kaplan and Meier (p=0.001). Disclosures: Valent: Novartis: Consultancy, Honoraria, Research Funding.

Prospective evaluation of the diagnostic value of sensitive KIT D816V mutation analysis of blood in adults with suspected systemic mastocytosis

“…In the present study, the KIT D816V allele burden was not significantly higher in SM patients who fulfilled the major criterion compared to patients who did not. This result is in line with a previous result by Hoermann et al (2014) who observed no significant correlation between the KIT D816V allele burden and the percentage of MCs in the BM biopsy or BM smear . Together, these results indicate that the association between the allele burden and degree of MC infiltration in the BM is limited, likely because the allele burden also reflects the degree of non‐MC lineage involvement in addition to the neoplastic MC burden .…”

“…We have previously demonstrated that the vast majority (94%) of already diagnosed SM patients test KIT D816V mutation positive in blood when using a sufficiently sensitive method . However, in line with similar studies from other groups also detecting the KIT D816V mutation in blood in high numbers of SM patients, our previous study was primarily based on blood samples obtained at, or later than, the time of the BM investigation. These blood samples were not used in the diagnostic workup to guide the BM biopsy, but were instead sampled after the decision to perform a BM biopsy was made (at the day of the BM biopsy or after the SM diagnosis was established).…”

Characterization of CD34+ hematopoietic cells in systemic mastocytosis: Potential role in disease dissemination

“…In parallel with the above findings, recent studies have demonstrated that most ISM patients, including the majority of ISMs− cases, show KIT ‐mutated cells in PB, even in the absence of multilineage BM involvement of hematopoiesis by the KIT mutation. Nevertheless, no study has investigated so far the specific compartment of circulating PB cells that carries the KIT mutation, particularly at the earliest disease stages.…”

Detection of KIT D816V in peripheral blood of children with manifestations of cutaneous mastocytosis suggests systemic disease

“…There are several published algorithms to assist in the diagnostic approach and determine which adults are more likely to have systemic disease (Alvarez-Twose et al, 2010;Gulen et al, 2014;Carter et al, 2018). Recent studies identifying and quantitating the KIT D816V mutation in peripheral blood (PB) in adults have suggested it to be a valuable tool for diagnosis, (Kristensen et al, 2011(Kristensen et al, , 2014Jara-Acevedo et al, 2015) to determine disease burden and monitoring (Erben et al, 2014), and to assess response to therapy (Hoermann et al, 2014). In adults, the KIT D816V allelic burden in PB correlates with serum tryptase (Kristensen et al, 2013;Erben et al, 2014;Hoermann et al, 2014).…”

“…Recent studies identifying and quantitating the KIT D816V mutation in peripheral blood (PB) in adults have suggested it to be a valuable tool for diagnosis, (Kristensen et al, 2011(Kristensen et al, , 2014Jara-Acevedo et al, 2015) to determine disease burden and monitoring (Erben et al, 2014), and to assess response to therapy (Hoermann et al, 2014). In adults, the KIT D816V allelic burden in PB correlates with serum tryptase (Kristensen et al, 2013;Erben et al, 2014;Hoermann et al, 2014). However, the diagnosis of systemic disease in children remains a challenge because, outside of the clinical finding of organomegaly, which correlates with systemic disease, surrogate markers to help identify those children where a marrow examination might be warranted to establish systemic disease, are lacking (Carter et al, 2015).…”