María Jara-Acevedo

Prognosis in adult indolent systemic mastocytosis: A long-term study of the Spanish Network on Mastocytosis in a series of 145 patients

BACKGROUND Indolent systemic mastocytosis is a group of rare diseases for which reliable predictors of progression and outcome are still lacking. OBJECTIVE Here we investigate the prognostic impact of the clinical, biological, phenotypic, histopathological, and molecular disease characteristics in adults with indolent systemic mastocytosis, who were followed using conservative therapy. METHODS A total of 145 consecutive patients were prospectively followed between January 1983 and July 2008; in addition, from 1967 to 1983, 20 patients were retrospectively studied. RESULTS Multivariate analysis showed that serum beta2-microglobulin (P = .003) together with the presence of mast/stem cell growth factor receptor gene (KIT) mutation in mast cells plus myeloid and lymphoid hematopoietic lineages (P = .02) was the best combination of independent parameters for predicting disease progression (cumulative probability of disease progression of 1.7% +/- 1.2% at 5-10 years and of 8.4% +/- 5.0% at 20-25 years). Regarding overall survival, the best predictive model included age >60 years (P = .005) and development of an associated clonal hematological non-mast cell disorder (P = .03) with a cumulative probability of death of 2.2% +/- 1.3% at 5 years and of 11% +/- 5.9% at 25 years. CONCLUSIONS Indolent systemic mastocytosis in adults has a low disease progression rate, and the great majority of patients have a normal life expectancy, with the presence of KIT mutation in all hematopoietic lineages and increased serum beta2-microglobulin the most powerful independent parameters for predicting transformation into a more aggressive form of the disease.

Increased frequency (12%) of circulating chronic lymphocytic leukemia-like B-cell clones in healthy subjects using a highly sensitive multicolor flow cytometry approach.

Monoclonal B-cell lymphocytosis (MBL) indicates the presence of less than 5 x 10(9)/L circulating monoclonal B cells in otherwise healthy subjects. Recently, it has been reported that circulating chronic lymphocytic leukemia (CLL)-like B cells can be detected using 4- or 5-multicolor flow cytometry in 5% to 7% of adults with normal lymphocyte counts. We investigated the frequency of circulating monoclonal B cells in 608 healthy subjects older than 40 years with normal blood counts, using a highly sensitive 8-color flow cytometry approach and systematic screening for total PB leukocyte count higher than 5 x 10(6). We show that the frequency of PB monoclonal B cells is markedly higher than previously reported (12% for CLL-like B cells, found at frequencies of 0.17 +/- 0.13 x 10(9) cells/L), the incidence progressively increasing with age. Most cases (62%) showed clonal B-cell levels below the maximum sensitivity of the techniques described by others (< 0.01%), supporting the notion that detection of MBL may largely depend on the sensitivity of the flow cytometry approach used.

Clinical, biological, and molecular characteristics of clonal mast cell disorders presenting with systemic mast cell activation symptoms

BACKGROUND Systemic mast cell activation disorders (MCADs) are characterized by severe and systemic mast cell (MC) mediators-related symptoms frequently associated with increased serum baseline tryptase (sBt). OBJECTIVE To analyze the clinical, biological, and molecular characteristics of adult patients presenting with systemic MC activation symptoms/anaphylaxis in the absence of skin mastocytosis who showed clonal (c) versus nonclonal (nc) MCs and to provide indication criteria for bone marrow (BM) studies. METHODS Eighty-three patients were studied. Patients showing clonal BM MCs were grouped into indolent systemic mastocytosis without skin lesions (ISMs(-); n = 48) and other c-MCADs (n = 3)-both with CD25(++) BM MCs and either positive mast/stem cell growth factor receptor gene (KIT) mutation or clonal human androgen receptor assay (HUMARA) tests-and nc-MCAD (CD25-negative BM MCs in the absence of KIT mutation; n = 32) and compared for their clinical, biological, and molecular characteristics. RESULTS Most clonal patients (48/51; 94%) met the World Health Organization criteria for systemic mastocytosis and were classified as ISMs(-), whereas the other 3 c-MCAD and all nc-MCAD patients did not. In addition, although both patients with ISMs(-) and patients with nc-MCAD presented with idiopathic and allergen-induced anaphylaxis, the former showed a higher frequency of men, cardiovascular symptoms, and insect bite as a trigger, together with greater sBt. Based on a multivariate analysis, a highly efficient model to predict clonality before BM sampling was built that includes male sex (P = .01), presyncopal and/or syncopal episodes (P = .009) in the absence of urticaria and angioedema (P = .003), and sBt >25 microg/L (P = .006) as independent predictive factors. CONCLUSIONS Patients with c-MCAD and ISMs(-) display unique clinical and laboratory features different from nc-MCAD patients. A significant percentage of c-MCAD patients can be considered as true ISMs(-) diagnosed at early phases of the disease.

Mast cells from different molecular and prognostic subtypes of systemic mastocytosis display distinct immunophenotypes

BACKGROUND Systemic mastocytosis (SM) is a heterogeneous group of disorders with distinct clinical and biological behavior. Despite this, little is known about the immunophenotypic features of the distinct diagnostic categories of SM. OBJECTIVE To analyze the immunophenotypic characteristics of bone marrow (BM) mast cells (MCs) of different subtypes of SM. METHODS Bone marrow samples from 123 patients with different subtypes of SM and 92 controls were analyzed for a broad panel of immunophenotypic markers by flow cytometry. RESULTS Three clearly different maturation-associated immunophenotypic profiles were found for BMMCs in SM. These different profiles were associated with both genetic markers of the disease and its clinical behavior. BMMCs from poor-prognosis categories of SM (aggressive SM and MC leukemia) typically showed an immature phenotype with clonal involvement of all myeloid lineages by the D816V stem cell growth factor receptor gene (KIT) mutation. In turn, a mature activated versus resting BMMC immunophenotype was commonly found among patients with good-prognosis subtypes of SM depending on whether they carried (indolent SM and clonal MC activation disorders) or not (well differentiated SM) the D816V KIT mutation. CONCLUSION Bone marrow MCs from SM show 3 different maturation-related immunophenotypic profiles that are associated with both the genetic markers of the disease and its clinical behavior.

Evaluation of the WHO criteria for the classification of patients with mastocytosis

Diagnosis and classification of mastocytosis is currently based on the World Health Organization (WHO) criteria. Here, we evaluate the utility of the WHO criteria for the diagnosis and classification of a large series of mastocytosis patients (n=133), and propose a new algorithm that could be routinely applied for refined diagnosis and classification of the disease. Our results confirm the utility of the WHO criteria and provide evidence for the need of additional information for (1) a more precise diagnosis of mastocytosis, (2) specific identification of new forms of the disease, (3) the differential diagnosis between cutaneous mastocytosis vs systemic mastocytosis, and (4) improved distinction between indolent systemic mastocytosis and aggressive systemic mastocytosis. Based on our results, a new algorithm is proposed for a better diagnostic definition and prognostic classification of mastocytosis, as confirmed prospectively in an independent validation series of 117 mastocytosis patients.

Nanotechniques in proteomics: Protein microarrays and novel detection platforms

The field of proteomics has undergone rapid advancements over the last decade and protein microarrays have emerged as a promising technological platform for the challenging tasks of studying complex proteomes. Researchers have gone beyond traditional techniques and approached promising disciplines like nanotechnology to satisfy the growing demands of studying proteins in high-throughput format. Applications of nanotechnology in proteomics came from the need to detect low-abundant proteins in complex mixtures for sensitive, real-time and multiplexed detection platform. The scope of this article is to outline the current status and key technological advances of nanotechniques in protein microarrays.

Validation of the REMA Score for Predicting Mast Cell Clonality and Systemic Mastocytosis in Patients with Systemic Mast Cell Activation Symptoms

Background: A variable percentage of patients with systemic mast cell (MC) activation symptoms meet criteria for systemic mastocytosis (SM). We prospectively evaluated the clinical utility of the REMA score versus serum baseline tryptase (sBt) levels for predicting MC clonality and SM in 158 patients with systemic MC activation symptoms in the absence of mastocytosis in the skin (MIS). Methods: World Health Organization criteria for SM were applied in all cases. MC clonality was defined as the presence of KIT-mutated MC or by a clonal HUMARA test. The REMA score consisted of the assignment of positive or negative points as follows: male (+1), female (–1), sBt <15 µg/l (–1) or >25 µg/l (+2), presence (–2) or absence (+1) of pruritus, hives or angioedema and presence (+3) of presyncope or syncope. Efficiency of the REMA score for predicting MC clonality and SM was assessed by receiver operating characteristic (ROC) curve analyses and compared to those obtained by means of sBt levels alone. Results: Molecular studies revealed the presence of clonal MC in 68/80 SM cases and in 11/78 patients who did not meet the criteria for SM. ROC curve analyses confirmed the greater sensitivity and a similar specificity of the REMA score versus sBt levels (84 vs. 59% and 74 vs. 70% for MC clonality and 87 vs. 62% and 73 vs. 71% for SM, respectively). Conclusions: Our results confirm the clinical utility of the REMA score to predict MC clonality and SM in patients suffering from systemic MC activation symptoms without MIS.

An immature immunophenotype of bone marrow mast cells predicts for multilineage D816V KIT mutation in systemic mastocytosis

D816V KIT mutation of bone marrow (BM) mast cells (MC) is a common feature to systemic mastocytosis (SM) patients. Nevertheless, occurrence of the KIT mutation in BM cell compartments other than MC is associated with progression to more aggressive forms of the disease and poor outcome in indolent SM (ISM). Here, we assessed the potential association between the immunophenotype of MC and multilineage KIT mutation in the BM of SM patients through the investigation of the flow cytometric protein expression profile (PEP) of bone marrow mast cells (BMMC) from 70 control individuals and 206 SM patients, classified according to the WHO (World Health Organization), and the degree of involvement of BM hematopoiesis by the D816V KIT mutation; additionally, we developed a score-based class prediction algorithm for the detection of SM cases with multilineage mutation. Our results show that aberrant expression of CD25 with a FcɛRIlo, FSClo, SSClo and CD45lo immature phenotype of BMMC, in the absence of coexisting normal MC in the BM, was associated with multilineage involvement by the D816V KIT mutation, regardless of the diagnostic subtype of the disease (for example, indolent vs aggressive SM), which supports the utility of the immunophenotype of BMMC as a surrogate marker to screen for multilineage KIT mutation in ISM.

Immunophenotyping in systemic mastocytosis diagnosis: 'CD25 positive' alone is more informative than the 'CD25 and/or CD2' WHO criterion.

Aberrant expression of CD2 and/or CD25 by bone marrow, peripheral blood or other extracutaneous tissue mast cells is currently used as a minor World Health Organization diagnostic criterion for systemic mastocytosis. However, the diagnostic utility of CD2 versus CD25 expression by mast cells has not been prospectively evaluated in a large series of systemic mastocytosis. Here we evaluate the sensitivity and specificity of CD2 versus CD25 expression in the diagnosis of systemic mastocytosis. Mast cells from a total of 886 bone marrow and 153 other non-bone marrow extracutaneous tissue samples were analysed by multiparameter flow cytometry following the guidelines of the Spanish Network on Mastocytosis at two different laboratories. The ‘CD25+ and/or CD2+ bone marrow mast cells’ World Health Organization criterion showed an overall sensitivity of 100% with 99.0% specificity for the diagnosis of systemic mastocytosis whereas CD25 expression alone presented a similar sensitivity (100%) with a slightly higher specificity (99.2%). Inclusion of CD2 did not improve the sensitivity of the test and it decreased its specificity. In tissues other than bone marrow, the mast cell phenotypic criterion revealed to be less sensitive. In summary, CD2 expression does not contribute to improve the diagnosis of systemic mastocytosis when compared with aberrant CD25 expression alone, which supports the need to update and replace the minor World Health Organization ‘CD25+ and/or CD2+’ mast cell phenotypic diagnostic criterion by a major criterion based exclusively on CD25 expression.

CD30 expression by bone marrow mast cells from different diagnostic variants of systemic mastocytosis

CD30 expression by bone marrow (BM) mast cells (MC) has been reported recently in systemic mastocytosis (SM) patients. The aim of this study was to investigate the potential diagnostic and prognostic value of CD30 expression in SM as assessed by multiparameter flow cytometry.