Clonal Hematological Non-mast Cell Lineage Research Articles

A Rare Case of Systemic Mastocytosis With Associated Clonal Hematological Non-Mast Cell Lineage Disease That Transformed to Acute Leukemia With IDH2 Mutation

An elderly 72-year-old man presented with anemia, thrombocytopenia, monocytosis, splenomegaly and lymphadenopathy. Bone marrow biopsy was consistent with mast cell neoplasm with positive CD117, CD25, CD34 myeloblasts and polymerase chain reaction (PCR) revealed mutation of D816V. He developed bilateral femoral neck fractures and biopsy confirmed that he has systemic mastocytosis (SM). He received cladribine and midostaurin with stable disease for 21 months. His SM with associated clonal hematological non-mast cell lineage disease (SM-AHNMD) transformed to acute myelogenous leukemia with isocitrate dehydrogenase 2 ( IDH2 ) mutation. A trial of enasidenib was given for 5 months but without any response. Patient decided to go with home hospice and died afterwards. J Med Cases. 2020;11(10):317-319 doi: https://doi.org/10.14740/jmc3552

165 A Rare Case of Systemic Mastocytosis With Associated Clonal Hematological Non-Mast Cell Lineage Disease

165 A Rare Case of Systemic Mastocytosis With Associated Clonal Hematological Non-Mast Cell Lineage Disease

Clinical potential of midostaurin in advanced systemic mastocytosis.

Advanced (Ad) systemic mastocytoses (SM) include aggressive SM (ASM) and mast cell leukemia (MCL) with or without an associated clonal hematological non-mast cell lineage disease (AHNMD). They are rare (<15%) but are associated with a poor prognosis due to rapid organ dysfunction. To date, responses to high-dose chemotherapy, cladribine, and imatinib were revealed to be suboptimal with a median survival time of 24 months. Midostaurin is a potent multikinase inhibitor including the most frequent KIT D816V mutation (>80%). We herein present a review of the most recent data of the use of midostaurin in AdSM. First, a multicenter Phase II study (CPKC412D2213) revealed an unprecedented overall response rate (ORR) of 69% regardless of KIT mutational status, with 38% of major response (MR) among 26 AdSM patients treated with midostaurin alone 200 mg daily. Second, a sponsor-initiated, multicenter, single-arm open Phase II study (CPKC412D2201) confirmed a high and durable ORR of 60% including 45% of MR among 89 AdSM patients. Finally, a French compassionate use program managed by the French Reference Centre for Mastocytosis allowed the treatment of almost a hundred AdSM patients to date in France since the CPKC412D2201 study closure. The outcome of the first 28 treated patients under cover of this on-going procedure revealed an ORR of 71% including 57% of MR. Most importantly, survival analysis revealed in comparison to a historical control cohort of AdSM patients who did not receive midostaurin a twofold lower risk of death (p=0.02) in midostaurin-treated patients. Side effects revealed were acceptable and manageable (mostly digestive). Midostaurin appears to be an effective and safe treatment of AdSM. However, its effect on the course of the AHNMD is less clear. For the future, combined therapy (hypomethylating agents, cladribine, mammalian target of rapamycin inhibitors, chemotherapy, and allogeneic bone marrow transplantation) may further improve long-term survival, particularly that of MCL and AdSM patients with AHNMD.

Comparison of gDNA‐based versus mRNA‐based KIT D816V mutation analysis reveals large differences between blood and bone marrow in systemic mastocytosis

Keywords: systemic mastocytosis; KIT D816V mutation; blood; genomic DNA; messenger RNA

Impact of centralized evaluation of bone marrow histology in systemic mastocytosis.

Bone marrow (BM) histology/immunohistochemistry, KIT D816V mutation analysis and serum tryptase measurements are mandatory tools for diagnosis of systemic mastocytosis (SM). Within the 'German Registry of Disorders on Eosinophils and Mast Cells', we identified 65 patients with SM who had two consecutive BM biopsies. The first biopsy was evaluated by a local pathologist (LP) and the second biopsy by a reference pathologist (RP) of the 'European Competence Network on Mastocytosis (ECNM)'. Final diagnoses by RP were SM (n = 27), SM or aggressive SM (ASM) with associated clonal haematological non-mast cell lineage disease [(A)SM-AHNMD, n = 34)] or mast cell leukaemia ± AHNMD (n = 4). In 15 of 65 patients (23%), initial diagnoses by LP were incorrect (by overlooking SM), for example primary myelofibrosis (n = 3), myelodysplastic/myeloproliferative neoplasm unclassified (n = 3) or B-cell lymphoma (n = 2). Fourteen of 15 patients (93%) with incorrect diagnosis had an advanced SM, mostly (A)SM-AHNMD. In the 50 concordantly diagnosed patients, immunohistochemical markers for quantitative assessment of mast cell infiltration, for example CD117 (KIT) or CD25, were applied by LP in only 34 of 50 patients (68%), and mutational analysis for KIT D816V was performed or recommended in only 13 of 50 patients (26%). Finally, the subclassification of SM was discordant because LP did not diagnose AHNMD in nine of 50 (18%) patients. In summary, adequate diagnosis and subclassification of SM requires an in-depth evaluation of the BM by experienced haematopathologists (preferably in a reference centre) in combination with molecular genetics, serum tryptase level and clinical parameters.

Systemic Mastocytosis with Smoldering Multiple Myeloma: Report of a Case.

Systemic mastocytosis (SM) is a disease characterized by a clonal infiltration of mast cells affecting various tissues of the body. It is grouped into six different subtypes according to the World Health Organization classification. It is called indolent systemic mastocytosis (ISM) when there is no evidence of end organ dysfunction, while the presence of end organ dysfunction defines aggressive systemic mastocytosis (ASM). When SM coexists with a clonal hematological disorder, it is classified as systemic mastocytosis with associated clonal hematological nonmast cell lineage disease (SM-AHNMD). Over 80% of SM-AHNMD cases involve disorders of the myeloid cell lines. To our knowledge, there are only 8 reported cases to date of SM associated with a plasma cell disorder. We report a patient with ISM who was found to have concomitant smoldering multiple myeloma. His disease later progressed to ASM. We discuss this rare association between SM and a plasma cell disorder, and potential common pathophysiologic mechanisms linking the two disorders will be reviewed. We also discuss prognostic factors in SM as well as the management options considered during the evolution of the patient's disease.

A 26-Year-Old Female with Systemic Mastocytosis with Associated Myeloid Neoplasm with Eosinophilia and Abnormalities of PDGFRB, t(4;5)(q21;q33).

Various translocations involving the PDGFRB gene are identified in myeloid neoplasms. However, the PRKG2/PDGFRB fusion gene associated with t(4;5)(q21;q33) has previously been reported in only 3 patients. We present the case of a 26-year-old woman with microcytic anemia, basophilia, thrombocytosis, and massive splenomegaly, who was found to have systemic mastocytosis and associated clonal hematological non-mast cell lineage disease (SM-AHNMD), with myeloid neoplasm with PRKG2/PDGFRB rearrangement. Initial findings included basophilia (37%, 4.1 k/μL), hypercellular marrow with eosinophilia, and increased and atypical megakaryocytes, suggestive of myeloproliferative neoplasm. Additional studies revealed large clusters of CD25 positive mast cells, fulfilling the criteria for the diagnosis of systemic mastocytosis. Consistent with prior reports of this translocation, our patient has responded well to imatinib. This case, in conjunction with others in the literature, suggests a possible connection between t(4;5)(q21;q33) PRKG2/PDGFRB and systemic mastocytosis and highlights their favorable response to imatinib.

Actualización en mastocitosis. Parte 2: categorías, pronóstico y tratamiento

Mastocytosis is a term used to describe a heterogeneous group of disorders characterized by clonal proliferation of mast cells in different organs. The organ most often affected is the skin. The World Health Organization classifies cutaneous mastocytosis into mastocytoma, maculopapular cutaneous mastocytosis, and diffuse mastocytosis. The systemic variants in this classification are as follows: indolent systemic mastocytosis (SM), aggressive SM, SM with an associated clonal hematological non-mast cell lineage disease, mast cell leukemia, mast cell sarcoma, and extracutaneous mastocytoma. The two latest systemic variants are rare. Although the course of disease is unpredictable in children, lesions generally resolve by early adulthood. In adults, however, the disease tends to persist. The goal of treatment should be to control clinical manifestations caused by the release of mast cell mediators and, in more aggressive forms of the disease, to reduce mast cell burden.

Molecular profiling of myeloid progenitor cells in multi-mutated advanced systemic mastocytosis identifies KIT D816V as a distinct and late event.

To explore the molecular profile and its prognostic implication in systemic mastocytosis (SM), we analyzed the mutation status of granulocyte-macrophage colony-forming progenitor cells (CFU-GM) in patients with KIT D816V(+) indolent SM (ISM, n=4), smoldering SM (SSM, n=2), aggressive SM (ASM, n=1), SM with associated clonal hematologic non-mast cell lineage disorder (SM-AHNMD, n=5) and ASM-AHNMD (n=7). All patients with (A)SM-AHNMD (n=12) carried 1-4 (median 3) additional mutations in 11 genes tested, most frequently TET2, SRSF2, ASXL1, CBL and EZH2. In multi-mutated (A)SM-AHNMD, KIT D816V(+) single-cell-derived CFU-GM colonies were identified in 8/12 patients (median 60%, range 0-95). Additional mutations were identified in CFU-GM colonies in all patients, and logical hierarchy analysis indicated that mutations in TET2, SRSF2 and ASXL1 preceded KIT D816V. In ISM/SSM, no additional mutations were detected and CFU-GM colonies were exclusively KIT D816V(-). These data indicate that (a) (A)SM-AHNMD is a multi-mutated neoplasm, (b) mutations in TET2, SRSF2 or ASXL1 precede KIT D816V in ASM-AHNMD,

Systemic mastocytosis with associated clonal hematological nonmast cell lineage disease

Systemic mastocytosis with associated clonal hematological nonmast cell lineage disease

Modelling of a genetically diverse evolution of Systemic Mastocytosis with Chronic Myelomonocytic Leukemia (SM-CMML) by Next Generation Sequencing.

BackgroundSystemic mastocytosis (SM) is a heterogenous, clonal mast cell (MC) proliferation, rarely associated with clonal hematologic non-mast cell lineage disease (SM-AHNMD). KITD816V is regarded as driver-mutation in SM-AHNMD.MethodsDNA isolated from peripheral blood (PB) of an SM-CMML patient was investigated with targeted next generation sequencing. Variants were verified by Sanger sequencing and further characterized in the SM part of the bone marrow trephine (BMT), normal tissue, and FACS sorted PB cell subpopulations.FindingsLow coverage deep-sequencing (mean 10x) on a GS 454 Junior revealed two as yet unreported SNVs (CBFA2T3 and CLTCL1), both germ-line mutations. High coverage (mean 1674x) targeted re-sequencing on an Ion Proton revealed 177 variants in coding regions. Excluding SNPs, the final list comprised 11 variants. Among these, TET2 (p.Thr1027fs, p.Cys1263Ser) and RUNX1 (p.Asn109Ser) were identified in in the peripheral blood and the SM part of BMT, but not in normal tissue. Furthermore, Sanger sequencing of PB cells revealed similar signal intensities for both TET2 mutations in FACS sorted CD34+ precursor cells and CD16+ granulocytes comparable to signals in the SM part of BMT. In contrast, RUNX1 exhibited a double intensity in CD34+ cells compared to the SM part of BMT and a homozygous variant signal in granulocytes. Both TET2 and RUNX1 mutations were not detectable in B- and T-cells.ConclusionWe present a heterozygous triple-mutation pattern (KIT, TET2, RUNX1) in mast cells (SM disease part) with additional LOH of RUNX1 in granulocytes (CMML disease part). These identified mutations allow a more detailed insight into a multistep pathogenesis which suggests a common tumor progenitor in SM-CMML.

Epidemiology of systemic mastocytosis in Denmark.

Mastocytosis is a heterogeneous group of diseases characterized by abnormal proliferation of mast cells. Systemic mastocytosis (SM), in which abnormal mast cells are present in tissues beyond the skin, is divided into seven subcategories with varying degrees of severity and prognosis. Very little is known about the epidemiology of SM and its subcategories. This retrospective cohort study of 548 adults with SM diagnosed 1997-2010 was constructed using linked Danish national health registries. The most common subtype of mastocytosis was indolent SM (including urticaria pigmentosa) (n=450; 82%), followed by SM with subtype unknown (n=61; 11%), SM with associated clonal haematological non-mast cell lineage disease (n=24; 4%), aggressive SM (n=8; 2%), and mast cell leukaemia (n=5; 1%). The incidence rate for SM (all subtypes including urticaria pigmentosa) was 0·89 per 100000 per year. Cumulative incidence was 12·46 per 100000, and the 14-year limited-duration prevalence as of 1 January, 2011 was 9·59 per 100000. This nationwide cohort from Denmark is the first population-based epidemiological study of mastocytosis. In this cohort of patients aged 15years and older, SM was found to be overall relatively rare with notable variation by subtype for patient characteristics, survival and epidemiological measures.

Allogeneic Hematopoietic Cell Transplantation Is Effective In Patients With Advanced Systemic Mastocytosis: A Multicenter Retrospective Analysis

Systemic mastocytosis (SM) is a rare hematologic neoplasm characterized by abnormal growth and accumulation of tissue mast cells (MC) in various organ systems, including bone marrow (BM). Indolent and advanced forms of SM have been described. Whereas patients with ISM have a normal or near normal life-expectancy, patients with advanced SM, including those suffering from mast cell leukemia (MCL) have a poor prognosis. In these patients, neoplastic MC are usually resistant against conventional drugs and various targeted drugs. In rapidly progressive aggressive SM (ASM) and MCL, polychemotherapy followed by allogeneic hematopoietic stem cell transplantation (alloHCT) has been proposed. However, outcome of alloHCT in advanced SM is unknown, and it also remains uncertain whether clinically relevant graft-versus-SM (GVSM) effects may occur in these patients, as only sporadic case reports have been published. We performed a retrospective multi-center analysis to evaluate the outcome of alloHCT in patients with advanced SM. Fifty-four advanced SM patients receiving SCT in 32 transplantation centers in Europe and America were identified between 1990 and 2013. The median patient age was 45 years. Donors were: HLA identical siblings (31), unrelated donors (URD) (15), umbilical cord blood donors (UCB) (2), and haploidentical donors (1). In 5 patients, stem cell source was not defined (5). Thirty-four patients received myeloablative conditioning (MAC) and 18 received reduced intensity conditioning regimens (RIC). In 2 patients, conditioning regimen was not specified. Indications for alloHCT were SM with an associated clonal hematologic non-mast cell lineage disease (SM-AHNMD) (n=32), MCL (n=13, including one with MCL-AHNMD), 8 with ASM and 1 with myelomastocytic leukemia (MML). The most prevalent AHNMD was acute myeloid leukemia (AML, n=16). With follow-up of 35-6180 (median 365) days, SM responses (defined as ≥50% decrease in BM mast cells ± decrease in serum tryptase ± regression of other organ manifestations) were observed in 39 patients (72%), including complete responses (CR) documented in 12 patients (22%). Eleven patients had stable disease, whereas 4 patients (7%) progressed immediately after alloHCT (primary resistance). In addition, 10 patients progressed (5 of them within 100 days) after an initial response. Progression was most frequently seen in MCL patients (n=6, 50%). In the AHNMD group, only 8 patients relapsed/progressed (25%). The overall survival (OS) and SM progression-free survival (PFS) at 1 year were 63% and 50% for all patients, 77% and 68% for SM-AHNMD, 63% and 50% for ASM, and 25% and 17% for MCL, respectively. The strongest predictive variable associated with inferior survival was a diagnosis of MCL. Other factors associated with poor outcome were: Karnofsky performance status ≤70%, ≥2 SM regimens given before alloHCT (e.g., steroids, cladribine, chemotherapy, tyrosine kinase inhibitor), donor source (alternative donors-UCB and haploidentical compared to sibling or URD), SM progression within the first 100 days, normal cytogenetics (compared to t(8;21) (q22;q22), and RIC (compared to MAC). The following variables were not associated with poor outcome: patient and donor age, recipient-donor sex match status, graft source (BM vs. peripheral stem cells), BM mast cell percentage at time of alloHCT, and CR status of AML or SM response at time of alloHCT. This largest multi-center analysis of results in advanced SM provides evidence for clinical efficacy of alloHCT, presumably because of a GVSM effect of alloHCT (achieving CR, and response to donor-lymphocyte infusions and RIC alloHCT). However, responses varied among different SM categories: while patients with SM-AHNMD enjoyed excellent outcomes, the OS for MCL patients in general, was poor. Nevertheless it is remarkable that 3 of 13 patients with MCL – an otherwise fatal disease with a median survival of <12 months – became long-term survivors after alloHCT. In summary, our results support development of a prospective alloHCT study, with the aim to optimize therapy and to improve overall outcome in advanced SM. Based on our pilot study, alloHCT should also be considered in practice for eligible and fit patients with SM-AHNMD (if treatment of AHNMD component needs alloHCT or SM component is aggressive), rapidly progressing ASM, MCL and MML when a suitable HLA-matched sibling donor or URD is available. Disclosures:Vercellotti:Sangart Inc.: Research Funding; Seattle Genetics: Research Funding. Akin:Novartis: Consultancy. Valent:Novartis: Consultancy, Honoraria, Research Funding.

Comprehensive mutational profiling in advanced systemic mastocytosis

AbstractTo explore mechanisms contributing to the clinical heterogeneity of systemic mastocytosis (SM) and to suboptimal responses to diverse therapies, we analyzed 39 KIT D816V mutated patients with indolent SM (n = 10), smoldering SM (n = 2), SM with associated clonal hematologic nonmast cell lineage disorder (SM-AHNMD, n = 5), and aggressive SM (n = 15) or mast cell leukemia (n = 7) with (n = 18) or without (n = 4) AHNMD for additional molecular aberrations. We applied next-generation sequencing to investigate ASXL1, CBL, IDH1/2, JAK2, KRAS, MLL-PTD, NPM1, NRAS, TP53, SRSF2, SF3B1, SETBP1, U2AF1 at mutational hotspot regions, and analyzed complete coding regions of EZH2, ETV6, RUNX1, and TET2. We identified additional molecular aberrations in 24/27 (89%) patients with advanced SM (SM-AHNMD, 5/5; aggressive SM/mast cell leukemia, 19/22) whereas only 3/12 (25%) indolent SM/smoldering SM patients carried one additional mutation each (U2AF1, SETBP1, CBL) (P < .001). Most frequently affected genes were TET2, SRSF2, ASXL1, CBL, and RUNX1. In advanced SM, 21/27 patients (78%) carried ≥3 mutations, and 11/27 patients (41%) exhibited ≥5 mutations. Overall survival was significantly shorter in patients with additional aberrations as compared to those with KIT D816V only (P = .019). We conclude that biology and prognosis in SM are related to the pattern of mutated genes that are acquired during disease evolution.

Use of octreotide for relief of gastro-intestinal (GI) symptoms in systemic mastocytosis

e present the case of an 82-year-old Caucasian female who was evaluated by hematology oncology for further work up and management of her chronic but progressive anemia. She reported weight loss, poor appetite, fatigue, nausea, several episodes of non-bloody diarrhea and increased abdominal girth over the past 3 months. On admission, her hemoglobin was 8.9 gm/dL, platelets were 43 k/mcl, WBC was 5.49 K/mcl with absolute neutrophil count of 3.73 k/mcl and 0% peripheral blasts. Her metabolic panel was abnormal for elevated LDH (245 U/L) and alkaline phosphatase (207 U/L). Her liver function tests were normal except for albumin of 2.5 g/dl. Computed tomography (CT) of the abdomen/pelvis showed splenomegaly, ascites, and retroperitoneal lymphadenopathy, a concern for a lymphoproliferative or metastatic disease. A bone marrow biopsy and aspirate performed to determine the cause of her progressive anemia and thrombocytopenia revealed hypercellularity (90%) and increased blasts (35% on the touch preparation). The blasts had scant cytoplasm and fine chromatin (blue arrows in Figure 1). In addition, there were paratrabecular infiltrates of round to spindled mast cells (red arrow in Figure 1), which by immunohistochemistry were strongly positive for tryptase (shown in Figure 2), CD117, CD68 and CD25. The blasts were weakly positive for CD117 and strongly positive for myeloperoxidase, CD56, Bcl-2, and CD43 (not shown). The aspirate was severely hemodiluted, likely due to the reticulin fibrosis observed in the biopsy. Flow cytometry specimen was also hemodiluted (9% blasts) with the following myeloid immunophenotype: CD33 positive CD13 positive CD56 positive CD117 partially positive CD34 negative HLA-DR negative. Based on these findings the patient was diagnosed with AML and SM, a subset of systemic mastocytosis with associated clonal hematologic non-mast cell lineage disease (SM-AHNMD). The patient’s diarrhea responded briefly to antimotility agent loperamide, but while awaiting definitive treatment plan, her symptoms worsened rapidly and required admission for severe, profuse non-bloody diarrhea, dehydration and abdominal distension. Splenomegaly and ascites were noted on clinical exam. Labs continued to show anemia and thrombocytopenia. Initial test was positive for clostridium difficile (C. diff). This was successfully treated with metronidazole and then with oral vancomycin. Subsequent tests for C. diff, other infectious agents and transglutaminase IgA antibody were negative. However, the patient continued to have multiple episodes of diarrhea with little relief from antimotility agents. Diarrhea was deemed secondary to SM. Extensive work up for portal hypertension was non-diagnostic. She could not undergo colonoscopy and liver biopsy due to thrombocytopenia. As a consideration for treatment with imatinib she was tested for BCR/ABL and PDGFRA fusion genes which were negative. KIT mutation analysis could not be performed due to lack of tissue sample. After a lengthy discussion with the patient regarding various options of treatment, she chose palliative care and hospice. However, severe diarrhea limited her transition to hospice; she was then started on a trial of octreotide every 8 h for her diarrhea. She had a rapid response with dra-

Systemic mastocytosis with associated clonal hematological non‐mast cell lineage disease: Clinical significance and comparison of chomosomal abnormalities in SM and AHNMD components

Some patients with systemic mastocytosis have concurrent hematological neoplasms, designated in the World Health Organization (WHO) classification as systemic mastocytosis with associated clonal hematological non-mast cell lineage disease (SM-AHNMD). In this study, we analyzed 29 patients with SM-AHNMD and compared them to 40 patients with pure SM. The AHNMDs were classified as chronic myelomonocytic leukemia (CMML) (n = 10), myelodysplastic syndrome (MDS) (n = 7), myeloproliferative neoplasms (n = 4), B-cell lymphoma/leukemia/plasma cell neoplasms (n = 7), and acute myeloid leukemia (n = 1). Patients with SM-AHNMD were older, more frequently had constitutional symptoms and hematological abnormalities, less often had skin lesions, and had an inferior overall survival compared with pure SM patients (48 months vs. not-reached, P < 0.001). Karyotypic abnormalities were detected in 9/28 (32%) patients with SM-AHNMD but not in pure SM patients (P < 0.001). Combined imaging/ fluorescence-in-situ hybridization performed in four SM-AHNMD cases revealed shared abnormal signals in mast cells and myeloid cells in two patients with SM-CMML and one patient with SM-MDS, but not in the mast cells of a case SM-associated with chronic lymphocytic leukemia with ATM-deletion. Quantitative mutation analysis showed higher levels of mutant KIT D816V in SM-CMML and SM-MDS than in pure SM (P < 0.001). Our data indicate that the SM-AHNMD category in the WHO classification is heterogeneous, including clonally related and unrelated forms of AHNMD. The presentation, treatment, and outcome of patients with SM-AHNMD is often dictated by the type of AHNMD.

A Case of Systemic Mastocytosis Associated with Acute Myeloid Leukemia Terminating as Aleukemic Mast Cell Leukemia after Allogeneic Hematopoietic Stem Cell Transplantation

In up to 40% of systemic mastocytosis (SM) cases, an associated clonal hematological non-mast cell lineage disease such as AML is diagnosed before, simultaneously with, or after the diagnosis of SM. A 40-yr-old man was diagnosed with AML with t(8;21)(q22;q22). Mast cells were not noted at diagnosis, but appeared as immature forms at relapse. After allogeneic hematopoietic stem cell transplantation (HSCT), leukemic myeloblasts were not observed; however, neoplastic metachromatic blasts strikingly proliferated during the state of bone marrow aplasia, and finally, aleukemic mast cell leukemia developed. As the disease progressed, we observed serial morphologic changes from immature mast cells with myeloblasts to only metachromatic blasts and atypical mast cells as mast cell leukemia; FISH analysis showed that the neoplastic mast cells originated from the same clone as the leukemic myeloblasts of AML.

Advanced systemic mastocytosis: the impact of KIT mutations in diagnosis, treatment, and progression

Apart from indolent systemic mastocytosis (SM), which is associated with a favorable prognosis, other subtypes of SM (SM with associated clonal hematologic non–mast cell lineage disease, aggressive SM, and mast cell leukemia – collectively referred to in this review as advanced SM) can be debilitating. The complexity of SM makes both the diagnosis and design of response criteria challenging for clinical studies. The tyrosine kinase KIT has been shown to play a crucial role in the pathogenesis of SM and has been a focal point in the development of targeted therapy. Mutations within various domains of the KIT receptor that lead to constitutive activation have been identified in patients, and those involving the activation loop of the KIT receptor are the mutations most frequently detected in patients with mastocytosis. Aberrant activation of the KIT receptor results in increased production of mast cells in extracutaneous organs that may lead to organ failure or early death. This review discusses the diagnosis and management of patients with advanced SM, including the relevance of KIT in this disease, potential therapies targeting this kinase, and criteria for measuring responses to these therapies.

Systemic mastocytosis with associated clonal hematological non-mast cell lineage disease: Clinical significance and comparison of chomosomal abnormalities in SM and AHNMD components

Systemic mastocytosis with associated clonal hematological non-mast cell lineage disease: Clinical significance and comparison of chomosomal abnormalities in SM and AHNMD components

Development and Validation of a Sensitive Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) Assay for the Simultaneous Quantification of Midostaurin (PKC412) and Its Two Major Metabolites: A Suitable Tool for the Monitoring of the Drug in Patients Suffering From Advanced Systemic Mastocytosis

Abstract 4913 Background:Among tyrosine kinase inhibitors, midostaurin (PKC412) takes place in the treatment of patients presenting with advanced systemic mastocytosis and, specifically aggressive systemic mastocytosis (ASM) or mast cell leukemia (MCL) with or without associated clonal hematological non-mast cell lineage disease (AHNMD). Our knowledge of the pharmacokinetic (PK) behavior of this major drug remains unfortunately sporadic. We developed a powerful liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay, which is able to sensitively detect PKC412 as its two major metabolites CGP52421 and CGP62221 in human plasma. The method was validated in clinical situation and, especially in the aim to monitor plasma concentrations of PKC412 in 11 patients, already enrolled in a Phase II clinical trial (Novartis Lab.). In addition, we also determined the PK profile of PKC412 at steady-state for one specific patient. Methods:A sensitive LC-MS/MS assay was developed for the simultaneous quantification of the product PKC412 and its two major metabolites in human plasma. Midostaurin-D5 was used as internal standard. The method was then validated in accordance with international guidelines and within the bioanalytical ±15% acceptance limits.In any case, PKC412 regimen was 100 mg bid. A total of 61 blood samples were collected from 11 patients and, between 15 min to 12 hrs following oral administration. Sampling took place over a period of 1 to 924 days following the beginning of treatment. PKC412 residual concentration at steady-state, range and mean concentration and, intra- and inter-individual variability coefficients were also calculated. On the other hand, a model independent method was used to determine the PK profile of PKC412 for a 12th patient (using 22 samples collected within 24 hrs): maximal concentration post-dosing (Cmax – ng/mL), time to Cmax (Tmax - hrs) were experimental values; terminal half-life (t1/2lz - hrs) and AUC0–12 were calculated during each 12 hrs interval post-dosing. Data are expressed as means ± SD. Results:This LC-MS/MS method is satisfactory for the key validation criteria i. e. trueness, bias, precision (repeatability and intermediate precision), accuracy and, linearity, over a range of concentrations of 75 to 2500 ng/mL for both PKC412, CGP52421 and, CGP62221. Patients monitored (8/11 males) were 57±14 yrs-old. All except one were treated for ASM±AHNMD (the last one had MCL), 10/11 patients carrying the ckit D816V mutation. The main characteristics at inclusion were as followed: body weight = 65±11 kg, BMI = 22±3 kg/m2, albumin = 31±6 g/L, GFR = 64±12 mL/min, bilirubin = 14±6 mg/L, GGT = 103±78 UI/L, ALP = 264±158 UI/L, tryptase = 412±411 μg/L. At the last follow-up visit i. e. between 54 to 924 day after initiation therapy, 8/11 patients were either in major or partial response with the following clinical picture: body weight 74±16 kg, BMI 24±4 kg/m2, albumin 40±2 g/L, DFG 59±11 mL/min, bilirubin 12±8 mg/L, GGT 56±57 UI/L, ALP 128±59 UI/L, tryptase 256±264 μg/L. It is important to point out that the tolerability profile was very good. The PKC412 residual concentrations value determined at steady-state were between 487 to 3254 ng/mL (1333±867 ng/mL). The intra- and inter-individual coefficients of variations were 18 and 65% respectively. About our 12th subject, major PK parameters were: Cmax = 3692±447 ng/mL, Tmax = 2, 5±2 hrs, t1/2lz = 95±88 hrs, AUC0–12= 34326±2197 ng/mL. hrs. Conclusions:The LC-MS/MS method is efficient and enables a sensitive and discriminative analysis of PKC412 and its major metabolites in human plasma. These first results highlight the high intra- and inter-individual variability of the PK behavior of the drug and, suggest the interest of its routine therapeutic monitoring vs. both adherence to the treatment and, its tolerability profile. Further, it has been shown that the 2 major metabolites of PK412 are active; under these conditions, we are working to the simultaneous therapeutic drug monitoring of the 3 compounds. Disclosures:No relevant conflicts of interest to declare.