Interferon-gamma and perforin-positive T cells in acquired aplastic anemia: implication in therapeutic response.

Acquired aplastic anemia (aAA) is an autoimmune disease, characterized by infiltration of T lymphocytes in the bone marrow with destruction of hematopoietic stem cells by the effector cells. Interferon-gamma (IFN-γ) and perforin are important mediators of cell destruction. In this flow cytometry-based study, we have investigated the percentage of intracellular IFN-γ+ and perforin+ CD5+ T cells in peripheral blood of newly diagnosed aAA patients before and after immunosuppressive therapy (IST). Patients were categorized as per standard disease severity and response to IST. The median percentage of IFN-γ+ and perforin+ CD5+ T cells was higher in untreated patients compared to healthy controls. The percentage of these cells was also increased in untreated severe and very severe aplastic anemia when compared with non-severe aplastic anemia patients. In patients before and after IST the median percentage of T cells producing IFN-γ and perforin was elevated in non-responders as compared to partial plus complete responders. The higher percentage of IFN-γ+ and perforin+ CD5+ T cells may be useful as an early diagnostic marker for aberrant activation of immune system and predict poor response to IST in aAA patients, who will benefit from alternative therapy.

Increased Expression of NOTCH-1 and T Helper Cell Transcription Factors in Patients with Acquired Aplastic Anemia.

Background: Acquired aplastic anemia is an autoimmune disease in which auto-aggressive T cells destroy hematopoietic progenitors. T-cell differentiation is controlled by transcription factors that interact with NOTCH-1, which influences the respective T-cell lineages. Notch signaling also regulates the BM microenvironment. The present study aimed to assess the gene expressions of NOTCH-1 and T helper cell transcription factors in the acquired aplastic anemia patients. Methods: Using quantitative real-time PCR, we studied the mRNA expression level for NOTCH-1, its ligands (DLL-1 and JAG-1), and T helper cell transcription factors (T-BET, GATA-3, and ROR-γt) in both PB and BM of aAA patients and healthy controls. Further, patients of aplastic anemia were stratified by their disease severity as per the standard criteria. Results: The mRNA expression level of NOTCH-1, T-BET, GATA-3, and ROR-γT genes increased in aAA patients compared to healthy controls. There was no significant difference in the mRNA expression of Notch ligands between patients and controls. The mRNA expression level of the above-mentioned genes was found to be higher in SAA and VSAA than NSAA patients. In addition, NOTCH-1 and T helper cell-specific transcription factors enhanced in aAA. We also observed a significant correlation between the genes and hematological parameters in patients. Conclusion: The interaction between NOTCH-1, T-BET, GATA-3, and ROR-γT might lead to the activation, proliferation, and polarization of T helper cells and subsequent BM destruction. The mRNA expression levels of genes varied with disease severity, which may contribute to pathogenesis of aAA.

Clinicopathological Significance of Common Genetic Alterations in Patients With Acute Promyelocytic Leukemia.

OBJECTIVE/BACKGROUND: Acute myeloid leukemia (AML) is one of the common forms of hematological malignancy and acute promyelocytic leukemia (APL) is a unique subtype of AML conferring favorable prognosis. We aimed to determine the prevalence and prognostic impact of Fms-like tyrosine kinase 3 (FLT3), nucleophosmin 1 (NPM1) mutation, epidermal growth factor receptor (EGFR), and flow marker's expression in patients with APL. METHODS: In the present study, 165 de novo APL patients were molecularly characterized for promyelocytic leukemia (PML) breakpoint and additional genetic alterations. Reverse transcriptase polymerase chain reaction (PCR) and real-time PCR assays were used to detect genetic alterations. RESULTS: PML/RARα was detected in 29/165 (17.5%) samples with breakpoint cluster region 1 (bcr1) in 17/29 (58.5%) and bcr3 in 12/29 (41.5%) samples. The prevalence of FLT3-ITD, NPM1, and EGFR were detected in 5/29 (17.5%), 11/29 (38%), and 5/29 (17.5%) patients, respectively. Patients expressing bcr-3 hybrid transcript had lower overall survival compared with bcr1 ( p = .254). White blood cell (WBC) count was significantly higher in bcr3 in comparison with bcr1 patients ( p = .002). Patients with positive EGFR expression ( p = .042) and higher WBC ( p = .002) were significantly associated with poor survival ( p < .05). CONCLUSIONS: We documented the higher prevalence of bcr1 and confirmed that the association of FLT3-ITD significantly reduced the chances of survival in APL. The mortality rate of bcr3 was comparatively higher than that of bcr1. Higher WBC count and EGFR expression were significantly associated with poor survival.

Social and Financial Barriers to Optimum TKI Treatment in Patients with Chronic Myeloid Leukemia-A Knowledge-Attitudes-Practices Study from India.

INTRODUCTION: Outcomes in chronic myeloid leukemia (CML) have vastly improved after introducing tyrosine kinase inhibitors. However, patients in low and middle-income countries (LMICs) face many challenges due to social and financial barriers. OBJECTIVE: This study was conducted to understand socio-economic hindrances, knowledge-attitudes-practices, and assessing nonadherence to treatment in chronic phase CML patients taking imatinib. MATERIALS AND METHODS: Patients of chronic phase CML, aged 15 and above, taking imatinib for six months or more were included in the study. A questionnaire (in the Hindi language) was administered, inquiring about the nature of the disease and its treatment, how imatinib was obtained, drug-taking behavior, and the treatment's economic and social burden. Nonadherence was assessed by enquiring patients for missed doses since the last hospital visit and for any treatment interruptions of ≥7 days during the entire course of treatment (TIs). RESULTS: Four hundred patients were enrolled (median age 37 years, median duration on imatinib 63 months). Patients hailed from 16 different Indian states, and 29.75% had to travel more than 500 kilometers for their hospital visit. Scheduled hospital visits were missed by 14.75%. A third of the patients were unaware of the lifelong treatment duration, and 41.75% were unaware of the risks of discontinuing treatment. Treatment was financed by three different means -61.75% received imatinib via the Glivec International Patient Assistance Program (GIPAP), 14.25% through a cost-reimbursement program, and 24% self-paying. 52.75% of patients felt financially burdened due to the cost of drugs (self-paying patients), cost of investigations, the expenditure of the commute and stay for the hospital visit, and loss of working days due to hospital visits. 41.25% of patients reported missed doses in the last three months, and 9% reported missing >10% doses. 16.5% of patients reported TIs. Nonadherence>10% and TIs were significantly higher in self-paying patients (15.6% and 25% respectively). CONCLUSION: We observed that patient awareness about the disease was suboptimal. Patients felt inconvenienced and financially burdened by the treatment. Nonadherence and treatment interruptions were observed in 41.25% and 16.5%, respectively. These issues were prevalent in self-paying patients.

The Prognostic Impact of Epidermal Growth Factor Receptor (EGFR) in Patients with Acute Myeloid Leukaemia.

Expression of Epidermal Growth Factor Receptor (EGFR), an important proto-oncogene, regulates cell differentiation, proliferation, cell migration and survival in most of the cancer types. EGFR expression has been reported in Acute Myeloid Leukaemia (AML), however, many other reports nullified EGFR expression in AML. These contradictory data prompted us to reevaluate the expression of EGFR in AML and carry out a comparative survival analysis between EGFR expressing and non-expressing AML patients (Children and Acute Promyelocytic Leukaemia patients excluded). Bone marrow and/or peripheral blood samples were collected from 60 adult patients with AML with written informed consent. PCR, Real-Time Taqman gene expression assays were used for the detection of genetic alterations. Statistical analysis was conducted using SPSS software (IBM SPSS 20). In our study, EGFR expression was detected in 21 out of 60, in 35% (95% C.I. 23.45-48.48) AML patients. Overall survival was significantly shorter in patients with EGFR (p = < 0.01), with an average survival of 18.57 months (95% C.I. 12.42-24.73 months) compared with 31.27 months (95% C.I. 28.19-34.33 months) in patients without EGFR. EGFR expression was significantly higher in female patients compared to male (p = 0.037).This study confirms the presence of EGFR in AML and indicates that EGFR expression confers poor prognosis in AML. However, the underlying cause of this adverse prognostic effect has not been identified. Further clinical studies are warranted to determine the exact mechanism through which EGFR activity might contribute to AML progression and identify the potential therapeutic target for the reversal of resistance to conventional chemotherapeutics.

Calreticulin Mutation by Immunohistochemistry: Can It Replace PCR?

Before 2013, the diagnosis of about 30% to 45% cases of primary myelofibrosis (PMF) and essential thrombocythemia (ET) posed a diagnostic difficulty because of the missing reliable clonal marker. Calreticulin (CALR) mutation was identified as a recurrent mutation in about 60% to 88% of JAK2/MPL-negative PMF and ET. Molecular methods like Sanger sequencing and polymerase chain reaction (PCR) are considered gold standard, but they have limited availability, complex techniques, and labor intensive. In contrast to molecular methods, immunohistochemistry (IHC) is a widely available, rapid, simple, and cost-effective option. There are only few studies evaluating the utility of IHC for CALR mutation detection. Hence, we studied the role of IHC in CALR mutation detection and compared it with PCR. Thirty-one JAK2V617F-negative PMF and ET were evaluated for CALR mutation status. PCR was done and interpreted by comparing bands with the expected product size. The bone marrow biopsy was simultaneously put up for IHC using antimutated CALR monoclonal antibody (CAL2). CALR mutation was detected in 64.5% (20/31) cases. Prevalence of CALR mutation in JAK2-negative PMF and ET was 60.9% (14/23) and 75% (6/8), respectively. Sensitivity, specificity, positive predictive value, and negative predictive value of IHC analyzed were 89.4%, 100%, 100%, and 84.6%, respectively. A very good level of agreement (κ=0.86) was observed between PCR and IHC. We suggest that IHC is the best screening test to detect CALR mutation in resource limited countries with limited availability and affordability of molecular methods.

Influence of JAK2V617F allele burden on clinical phenotype of polycythemia vera patients: A study from India.

BACKGROUND: Elevated JAK2V617F allele burden is associated with enhanced expression of downstream target genes in Philadelphia negative chronic myeloproliferative neoplasms (CMPNs) which include PV, ET & PMF. Previous studies have shown the impact of JAK2V617F allele burden on clinical phenotype of CMPNs. However, there is no data from India regarding the association between JAK2V617F allele burden and clinical phenotype in PV. AIMS/SETTINGS AND DESIGN: We aimed to investigate the effect of allele burden on clinical phenotype in 90 JAK2V617F positive PV patients and to see its influence on disease related complications. MATERIAL AND METHODS: Allele burden of 90 JAK2V617F positive PV patients was quantified by Real-time polymerase chain reaction (RQ-PCR). RESULTS: 74/90 (82.22%) were males and 16/90 (17.78%) were females (median 45 years, range 35-78). Patients with age >50 years had significantly higher JAK2V617F allele burden (median 40.15%, range 0.49-91.62 %) than patients with ≤ 50 years age (median 48.59 %, range 0.56-86.74 %; P < 0.032). Patients with splenomegaly had significantly higher JAK2V617F allele burden (mean 50.24%, range 6.91-84.17%) than patients without splenomegaly (mean 33.82 %, range 0.49-71.83 %; P < 0.017). Patients with higher allele burden (median 57.20, range 43.4-72.03%) had significantly raised thrombotic events than the patients with lower allele burden (median 37.38, range 0.49-84.17%; P < 0.043). 49/90 (54%) were homozygous and 41/90 (46%) were heterozygous. CONCLUSIONS: Higher JAK2V617F allele burden showed association with increased age, splenomegaly and thrombotic events. Thus, it may be considered for prognostication and setting up the treatment protocol in PV patients.

Distribution of common BCR-ABL fusion transcripts and their impact on treatment response in Imatinib treated CML patients: A study from India.

BACKGROUND: Philadelphia chromosome (Ph): Hallmark of CML is caused by reciprocal translocation between chromosomes 9 and 22 resulting in BCR-ABL fusion protein. Most commonly associated breakpoint with CML is M-bcr in exon 13 or exon 14, producing splice variant b2a2 or b3a2 respectively. The distribution of these transcripts and their influence on clinico-hematological parameters is variable. Impact of the fusion transcripts on treatment outcome in Imatinib treated CML patients is still a matter of debate. AIMS/SETTINGS AND DESIGN: We conducted this study on 400 CML-CP patients to look for the distribution of fusion transcripts i.e. b3a2 and b2a2, their clinico-hematological profile and impact on treatment response in patients treated with Imatinib. MATERIAL AND METHODS: CML-CP was diagnosed by reverse transcriptase PCR (RT-PCR) for the BCR-ABL fusion transcript. Real-time quantitative PCR (RQ-PCR) was performed on peripheral blood every 3-6 monthly to look for treatment response. RESULTS: The overall frequency of b3a2 transcript was observed in 288 (72%) followed by b2a2 in 104 (26%) and hybrid fusion transcript (b3a2 + b2a2) was seen in 8 (2%) cases. MMR was attained in 198/288 (68.7%) patients with b3a2 transcript and 90/288 (31.3%) patients failed to achieve MMR after 12 months of Imatinib therapy. Among the patients with b2a2 transcript, 44/104 (42.3%) patients achieved MMR and 60/104 (57.7%) patients failed to achieve MMR after 12 months of Imatinib therapy. CONCLUSIONS: In conclusion, the frequency of b3a2 transcript was more as compared to b2a2 transcript. MMR was significantly higher in patients with b3a2 transcript as compared to patients with b2a2.

Comparative analysis of the Sokal, Euro and European Treatment and Outcome Study score in prognostication of Indian chronic myeloid leukemia-chronic phase patients on imatinib.

INTRODUCTION: The ultimate goal for CML management is risk stratification of the patients to design the appropriate treatment approach. The Sokal, Euro and EUTOS risk scores were established to prognosticate the patients on therapy. AIM: To perform a comparative assessment of the Sokal, Euro and EUTOS prognostic score in Indian CML-CP patients on imatinib. METHODS: This is a retrospective study performed in 260 Ph+ CML-CP patients who were administered oral imatinib (400 mg/day). RESULTS: 166/260 were males and 94/260 were females (M: F::1.6:1) with median age 35 years (range 20-70). 92 (35.38%), 125 (48.07%) and 43 (16.5%) patients were divided into low, intermediate and high risk Sokal score respectively. 102 (39.23%), 106 (40.76%) and 52 (20%) patients were discriminated into low, intermediate and high risk Euro score respectively. 210 (80.7%) and 50 (19.2%) patients were divided into low and high risk EUTOS score. Cumulative incidence of MMR for low, intermediate and high-risk Sokal score was 87%, 76% and 84% respectively (P = 0.016). Incidence of MMR in low, intermediate and high-risk Euro score was 93%, 85% and 68% respectively (P = 0.001). Incidence of MMR was 80 % and 81% for low and high risk EUTOS score (P = 0.764). Both EFS and OS are significantly correlated with Sokal score (P = 0.004, P = 0.007) and Euro score (P = 0.009, P = 0.001) but not with EUTOS score (P = 0.581, P = 0.927). CONCLUSION: The present study highlights the significant prognostic role of Sokal and Euro score in predicting the treatment outcome of the CML- CP patients on imatinib.

Association of JAK2V617F mutation with thrombosis in Indian patients with Philadelphia negative chronic myeloproliferative neoplasms.

BACKGROUND: : It is still a matter of debate regarding the association of JAK2V617F mutation with thrombosis in BCR-ABL negative CMPN patients. The role of JAK2V617F mutation in increasing the thrombotic risk in CMPNs is yet unequivocal. AIMS: : To clarify the contribution of JAK2V617F mutation in thrombosis in CMPN patients. SETTINGS AND DESIGN: This retrospective study was done to evaluate role of JAK2V617F mutation in thrombosis in CMPNs. MATERIALS AND METHODS: 65 CMPN patients (PV, ET and PMF) were analyzed for JAK2V617F mutation using ARMS-PCR and detailed history of thrombosis was recorded in these patients. STATISTICAL ANALYSIS: P values were 2 tailed, and statistical significance was set at P < 0.05. RESULTS: : 46/65 were males and 19/65 were females [M: F: 2.4:1] with median age 46 years [range, 14-80 years]. Patients had median Hb 15.6 g/dl [range, 5.1-20.3], median TLC 10.7 × 109/l [range 2.4-216] and platelet count 360 × 109/l [range, 20-1859]. 32 were JAK2V617F positive and 33 were negative for this mutation. On comparing the prevalence of thrombosis in JAK2V617F positive patients with JAK2V617F negative patients, we observed that 20/32 (62.5%) JAK2V617F positive patients had thrombosis as compared to 16/33 (48%) in JAK2V617F negative patients (P = 0.04). We observed significant association of JAK2V617F mutation with thrombosis, however no association of this mutation with thrombosis was observed among the JAK2V617F negative patients. CONCLUSION: Our study suggests that JAK2V617F mutation may increase the risk of thrombosis in CMPNs. This finding could lead to risk stratification, setting up the treatment strategy in CMPNs.

Laboratory Monitoring of Chronic Myeloid Leukemia in Patients on Tyrosine Kinase Inhibitors.

Chronic Myeloid Leukemia (CML) is a myeloproliferative neoplasm characterized by translocation of genetic material from chromosome 9 to chromosome 22 to form a fusion gene (BCR-ABL1) that is responsible for abnormal tyrosine kinase activity and alteration of various downstream signaling pathways. In addition to morphological diagnosis of CML phase, it is essential to detect BCR-ABL1 fusion by either metaphase cytogenetics or reverse transcriptase polymerase chain reaction that also determines type of mRNA transcript. Once treatment begins, monitoring the response to Tyrosine Kinase Inhibitor (TKI) using standardized techniques and guidelines is important to check for failure of response and thus, plan timely intervention by increasing the dose of TKI or opting for second line TKIs. The goal is to stop evolution of CML to accelerated phase or blast crisis that has poor response to treatment. Also, it is desirable to achieve good outcomes and even treatment free remission in patients of CML on TKI. Thus, molecular monitoring by reverse transcriptase quantitative PCR (RT-qPCR) is done at regular intervals. There are international recommendations and quality control measures to standardize the reporting of fusion gene transcript levels by quantitative PCR (RT-qPCR) in CML to achieve and maintain sensitivity in molecular detection of CML disease burden. Various state-of-the-art molecular techniques have emerged to accurately determine the number of fusion-gene transcript levels. This review highlights various methodologies and their practical implications in management of CML patients on TKI.

Prevalence and Clinical Significance of FLT3 and NPM1 Mutations in Acute Myeloid Leukaemia Patients of Assam, India.

Acute Myeloid Leukaemia (AML) is one of the common forms of haematological malignancy in adults. We analysed the prevalence and clinical significance of FMS-like tyrosine kinase 3 (FLT3) and Nucleophosmin 1 (NPM1) mutations in AML patients of North East India. Co-prevalence and clinical significance of three recurrent chromosomal translocations namely t(15; 17), t(8; 21), t(16; 16) and expression of epidermal growth factor receptor (EGFR), flow markers were also documented and co-related with disease progress. We analysed bone marrow aspirates or peripheral blood samples from 165 newly diagnosed AML patients. All clinical samples were analysed by Real Time PCR and DNA sequencing based assays. NPM1 was the most frequently detected mutation in the study population (46/165 = 27.90%, 95% CI 20.75-35.05). FLT3 mutations were detected in 27/165 (16.40%, 95% CI 10.45-22.35) patients with internal tandem duplication (FLT3-ITD) in 24/165 (14.60%, 95% CI 8.91-20.29) and FLT3-D835 in 3/165 (1.80%, 95% CI 0-4.13) patients. NPM1 mutations were associated with a higher complete remission rate and longer overall survival (P < 0.01) compared to FLT3-ITD whereas FLT3-ITD showed adverse impact with poor survival rate (P < 0.01), leukocytosis (P < 0.01) and a packed bone marrow. EGFR expression was more in patients with NPM1 mutation compared to FLT3 mutation (P = 0.09). Patients with FLT3 and NPM1 mutations uniformly expressed CD13 and CD33 whereas CD34 was associated with poor prognosis (P ≤ 0.01) in patients with NPM1 mutation. FLT3-ITD was associated with inferior overall survival. However the clinical significance of FLT3-D835 was not clear due to small number of samples. NPM1 mutation showed better prognosis with increased response to treatment in the absence of FLT3-ITD.

Reversal of Glanzmann thrombasthenia platelet phenotype after imatinib treatment in a pediatric chronic myeloid leukemia patient.

Chronic Myelogenous Leukemia (CML) is a myeloproliferative neoplasm characterized by proliferation of Philadelphia positive clonal pluripotent hematopoietic cells. Bleeding is a rare presentation of CML that can occur due to platelet dysfunction. Both pre-treatment and post-treatment platelet function abnormalities in CML have been described in the literature. We describe a rare case of childhood CML who presented with mucocutateous bleeding manifestations. On laboratory workup, a Glanzmann Thrombasthenia (GT) like platelet phenotype was demonstrated along with confirmation of diagnosis of CML in chronic phase. The acquired nature of platelet function defect was confirmed by demonstrating recovery of platelet antigens glycoprotein IIb/IIIa after achieving complete hematological response with Imatinib. Due to presenting complaint of bleeding diathesis and absence of hepatosplenomegaly, the case was undiagnosed for CML until the patient reported to us. Careful evaluation of complete blood counts, peripheral blood picture and detailed laboratory workup was the window to proper diagnosis and treatment in this case.

NPM1 and FLT3 mutations in acute myeloid leukemia with normal karyotype: Indian perspective.

BACKGROUND: FLT3-ITD and NPM1 mutations are considered to be the major determinants of the patient response to therapy and outcome. The primary aim of this study was to establish the correlation between these molecular mutations and the clinico-hematologic parameters as well as the prognostic outcome of the Indian acute myeloid leukemia (AML) patients. MATERIALS AND METHODS: This prospective study involved newly diagnosed nonpromyelocytic AML patients who had undergone complete diagnostic workup, including immunophenotyping, conventional cytogenetics and molecular analysis for NPM1 and FLT3-ITD mutation by reverse transcriptase polymerase chain reaction at presentation. RESULTS: Overall, the prevalence of NPM1 and FLT3-ITD mutations was found to be 14.4% and 10.8%, respectively. Among patients with normal karyotype, leukocytosis was significantly associated with NPM1+ group than the NPM1- group (P = 0.0019) and more severe degree of anemia was observed in the FLT3-ITD+ patients than the other groups (P = 0.025). No significant correlation was found in terms of age at presentation (P = 0.56), sex ratio (P = 0.467), median platelet count (P = 0.27), and blast percentage between NPM1+ and FLT3-ITD+ groups. Complete remission (CR) rates were better in the NPM1+/FLT3-ITD- group than the other three groups. Unlike most other studies, improved CR rates as well as disease-free survival were observed in the NPM-/FLT3-ITD- group than the FLT3-ITD+ groups although not reaching statistically significant levels. CONCLUSION: Some differences in the clinical behavior of the Indian AML patients in comparison to that of the West in the presence of NPM1 and FLT3-ITD suggests that comprehensive studies are required to confirm the definitive role of these mutations among AML patients, especially with normal karyotype.

Chronic myeloid leukemia with a rare fusion transcript, e19a2 BCR-ABL1: A report of three cases from India.

The µ-bcr breakpoint connects exon 19 of BCR with ABL giving rise to the e19a2 transcript corresponding to the p230 fusion protein (micro-BCR breakpoint) which is rarely seen in chronic myeloid leukemia (CML) patients. Here we report three patients with p230 fusion protein presenting with different clinical presentations and diagnosed as CML-CP. These patients received Imatinib (tyrosine kinase inhibitor-TKI) and are still in remission.

Does Indian Myelodysplastic Syndrome Have a Biology Different from That in the West ?

abstract.

Calreticulin mutation profile in Indian patients with primary myelofibrosis.

INTRODUCTION: Somatic mutations in Calreticulin (CALR) have been recently discovered in JAK2/MPL unmutated patients with primary myelofibrosis (PMF) or essential thrombocythemia. METHODS: Clinical and hematologic features were obtained for 80 patients with PMF. JAK2V617F mutation was analyzed by DNA tetra-primer amplification refractory mutation system (ARMS-PCR). CALR and MPL mutations were identified by bi-directional Sanger sequencing. RESULTS: CALR mutations were detected in 11.2% (9/80) of all PMF patients and 25.7% (9/35) of all JAK2V617F and MPL unmutated patients all of which were Type I mutation or deletions. A novel CALR mutation pattern (c.1241_1288del) was identified in one (1/9) patient. No case of Type II mutations or scattered point mutations was found in any of these patients. Uni-variate analysis at presentation showed that CALR mutations were significantly associated with younger age (P = 0.003) and larger spleen size (P = 0.001). No significant correlation was found between CALR mutation and clinico-hematologic characteristics or international prognostic scoring system (IPSS) scoring of the PMF patients. CONCLUSION: CALR mutations have a distinct molecular profile in Indian patients, different from that of other studies worldwide. Larger prospective studies need to be designed to establish the impact of paucity of Type II mutations in contributing to disease phenotype and prognostic outcome of patients.

Correlation of JAK2V617F mutational status in primary myelofibrosis with clinico-hematologic characteristics and international prognostic scoring system scoring: a single center experience.

INTRODUCTION: Somatic mutation in the exon 14 of Janus Kinase 2 gene is an established diagnostic marker in bcr-abl negative myeloproliferative neoplasms, especially primary idiopathic myelofibrosis (PIMF). AIM: Our primary aim was to find out the correlation between the JAK2V617F mutational status and the clinico-hematologic characteristics, as well as the international prognostic scoring system (IPSS) scoring of patients with PIMF. MATERIALS AND METHODS: Clinical and hematologic features were reviewed for 68 patients with primary idiopathic myelofibrosis (PIMF). JAK2V617F mutation status was analyzed by amplification refractory mutation screening-polymerase chain reaction. The patients were further stratified into low, intermediate-1, intermediate-2 and high-risk groups on the basis of IPSS scoring. RESULTS: The JAK2V617F mutation was detected in 58.8% patients. Univariate analysis of variables at presentation identified that JAK2V617F negative patients were significantly associated with more severe anemia (P = 0.045), younger age (P = 0.008), higher transfusion requirement (P = 0.017), and thrombocytopenia (P = 0.015). Patients who were homozygous for JAK2V617F mutation were associated with thrombocytosis (P = 0.014) and also had higher median total leucocyte count (P = 0.20) than the other groups. No significant correlation was detected between JAK2V617F mutational status and the presence of constitutional symptoms, spleen size, grade of bone marrow fibrosis or prognostic risk stratification of the PIMF patients. CONCLUSION: The variations in the prognostic implication of PIMF patients with mutation status as stated by various publications worldwide, reinstates the need for larger prospective studies using standardized JAK2V617F quantification methods as well as estimation of other newer molecular markers to develop deeper insight into various molecular alterations involving PIMF patients in India as well as worldwide.