Shifting gears to differentiation agents in acute promyelocytic leukemia with resource constraints-a cohort study.

BACKGROUND: Treatment of acute promyelocytic leukaemia has emerged as a major success in hemato-oncology. While literature from the developed world boasts of outstanding outcomes, there is a paucity of data from the developing world. This study aimed to assess complications and outcomes of acute promyelocytic leukaemia in a resource-constrained setting. METHODS: We retrospectively collected data from patients diagnosed with APL from January 2016 to December 2020. RESULTS: Sixty-four patients were treated-32 in both the Sanz high and low-risk groups. In the Sanz low-risk group, 12.5% of patients received ATRA with daunorubicin and 81.25% received ATRA with ATO. In the Sanz high-risk group, 18.8% of patients received ATRA with daunorubicin, 34.3% received ATRA with daunorubicin and ATO while 40.6% received ATRA with ATO. 56.25% of patients developed differentiation syndrome. The incidence was higher in Sanz high-risk group as compared to Sanz low-risk group. 57.4% of patients had an infection at the time of presentation. 62.5% of patients developed neutropenic fever during treatment. 17.2% of patients developed pseudotumor cerebri. The 4-year EFS and OS were 71.25 and 73.13%, respectively. Sanz low-risk group had a better 4-year EFS and OS as compared to the Sanz high-risk group. Haemoglobin at presentation and Sanz high-risk group were associated with poorer outcomes with a hazard ratio of 0.8 and 3.1, respectively. Outcomes in high-risk patients were better with the use of ATRA + ATO + daunorubicin. CONCLUSION: In the Indian population, APL patients have a high incidence of differentiation syndrome, pseudotumor cerebri, and infections during induction. CR, EFS, and OS compared to the developed world can be achieved with optimal therapy. Low haemoglobin at presentation and Sanz high-risk group were associated with poorer outcomes. ATRA, ATO, and daunorubicin combination is the preferred protocol for treating high-risk patients.

COVID-19 infection in patients with haematological disease - A tertiary centre experience from north India.

This retrospective study was aimed to understand the clinical, laboratory, radiological parameters and the outcome of COVID-19 patients with underlying haematological disease. All patients with known haematological disease admitted with COVID-19-positive status from April to August 2020 in the COVID-19 facility of a tertiary care centre in north India, were included. Their medical records were analyzed for outcome and mortality risk factors. Fifty four patients, 37 males, were included in the study. Of these, 36 patients had haematological malignancy and 18 had benign disorder. Fever (95.5%), cough (59.2%) and dyspnoea (31.4%) were the most common symptoms. Nine patients had severe disease at diagnosis, mostly malignant disorders. Overall mortality rate was 37.0 per cent, with high mortality seen in patients with aplastic anaemia (50.0%), acute myeloid (46.7%) and lymphoblastic leukaemia (40.0%). On univariate analysis, Eastern Cooperative Oncology Group performance status >2 [odd ratio (OR) 11.6], COVID-19 severity (OR 8.2), dyspnoea (OR 5.7) and blood product transfusion (OR 6.4) were the predictors of mortality. However, the presence of moderate or severe COVID-19 (OR 16.6, confidence interval 3.8-72.8) was found significant on multivariate analysis. The results showed that patients with haematological malignancies and aplastic anaemia might be at increased risk of getting severe COVID-19 infection and mortality as compared to the general population.

Transfusion practices during the COVID-19 pandemic: An experience from a hematology daycare in India.

The Coronavirus disease-19 (COVID-19) pandemic has in multiple ways affected healthcare delivery to non-COVID patients throughout the world. Adequate transfusion services are fundamental in ongoing therapy of patients with hematological ailments. We present the transfusion services in the hematology daycare under the department of Hematology and supported by the Blood Bank at our institution for the period 12th April 2020-30th June 2020, which saw the stringent lockdown and unlocking Phase I in India, declared in lieu of the pandemic. A 56 % reduction in total transfusion sessions was observed in 2020 (588 sessions given to 176 patients) compared to 1336 sessions in 516 patients over the same period in 2019. The reductions were seen across the different blood components (packed red blood cells [PRBC]: 585 vs. 1840, platelet rich plasma: 372 vs. 1313, single donor platelet 18 vs. 16), with a significant reduction in the mean PRBC transfused per PRBC transfusion session (1.11 vs 1.99, p<0.001) in 2020, compared to 2019. There were however no major differences in the transfusion practices across the different phases of the lockdown. Our study highlights the detrimental reduction in transfusion services due to the COVID-19 pandemic and related lockdown and showcases the remedial strategies taken to maximize transfusion support to patients during this period. Our observations might help to provide insights to adequately combat possible similar adverse situations in the future.

Comparison of CRP and procalcitonin for etiological diagnosis of fever during febrile neutropenia in hematology patients- an experience from a tertiary care center in Northern India.

INTRODUCTION: Febrile neutropenia is a common cause in morbidity and mortality during treatment of hematological neoplasms. METHODS: Subjects included all cases admitted under hematology department with febrile neutropenia from February to June 2018. Each febrile episode was investigated by standard investigations (Blood culture, Chest x ray etc.); Procalcitonin (PCT) and c reactive protein (CRP) was sent at fever onset 0, 24, 48 h, day 7 and day 14. RESULTS: Data was analyzed for 52 febrile episodes in 50 patients. PCT cut off value at 24 h of ≤1.2 ng/ml had a sensitivity and specificity of 62.5% and 87.5% for discriminating Invasive fungal infection (IFI) and Microbiologically documented infection (MDI) (p = 0.033). PCT had a negative predictive value of 70% for the diagnosis of IFI as compared to MDI. CRP cut off >160 mg/dl at 48 h was suggestive of fever due to fungal infection with a sensitivity of 100%, specificity of 48%, PPV of 33.3% and NPV of 100%. CRP at 24 and 48 h of fever was useful to distinguish non-infectious causes of fever from infectious causes. CONCLUSION: PCT at 24 h and CRP at 48 h was useful in identifying fungal infection. CRP was a better marker when compared to PCT for identifying disease fever.

Outcomes of paroxysmal nocturnal hemoglobinuria in the pediatric age group in a resource-constrained setting.

Paroxysmal nocturnal hemoglobinuria (PNH) is a rare acquired clonal stem cell disorder. Eculizumab and bone marrow transplantation are disease-modifying treatments for PNH but may not be readily available in resource-constrained settings. Of 52 pediatric patients with PNH, 20 had classical PNH and 32 had PNH/aplastic anemia (PNH/AA). Median time to diagnosis was 30 months in classical PNH patients. Renal failure was present in four patients (20%). Six (30%) achieved complete response, 10 (50%) achieved partial response with androgens in classical PNH. Two underwent allogenic stem cell transplantation. In the PNH/AA group, 16 (50%) were in CR and seven (21%) were in PR with anti-thymocyte globulin ± cyclosporine.

Pediatric myelofibrosis: WHO 2024 update on myeloproliferative neoplasms calling?

OBJECTIVES: Pediatric myelofibrosis is a rare entity with the largest reported series of 19 cases. We describe here the clinicopathological spectrum and outcomes of 15 cases of pediatric myelofibrosis. METHODS: Case files of myelofibrosis of patients less than 18 years were retrieved from January 2016 to January 2019, and patients with idiopathic myelofibrosis after exhaustive work-up were studied. Their clinicopathological profiles were studied and then followed up for resolution and malignant transformation. RESULTS: Of the 15 cases of idiopathic myelofibrosis, transfusion-dependent anemia (14/15) was most common presentation. Only one patient showed leukoerythroblastosis with dacryocytes. Myeloid hyperplasia was seen in 13 of 15 patients and megakaryocytic hyperplasia in 10 patients. Dysmegakaryopoiesis was seen in 8 of 15 patients, and only three had small loose megakaryocytic clustering. None showed hyperchromatic megakaryocytes, intrasinusoidal hematopoiesis, or osteosclerosis. One patient with trisomy 8 tested positive for JAK2V617F. Bone marrow biopsy was hypercellular in 13, and 8 had world health organization (WHO) MF-3 fibrosis. None of the patients developed malignancy, one had spontaneous resolution, and one patient required allogenic stem cell transplant. CONCLUSIONS: Pediatric myelofibrosis is a distinct entity from primary myelofibrosis in adults and merits mention in the WHO manual as a distinct entity.

Concomitant Existence of Paroxysmal Nocturnal Hemoglobinuria in a Patient with Hb E (HBB: c.79G>A) Trait.

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired clonal disorder that manifests with bone marrow failure, thrombosis and hemolysis. We present a 28-year-old male who presented with weakness, jaundice and transfusion dependence. On initial investigation, he was found to have anemia with jaundice with hemoglobin (Hb) capillary zone electrophoresis suggestive of Hb E (HBB: c.79G>A) trait. The same anomaly was also found in his mother. However, transfusion requirement was an unusual feature in the patient. As his corrected reticulocyte count was raised along with lactate dehydrogenase (LDH), which was suggestive of a hemolytic process, he was worked-up for the same. However, the direct Coombs test was negative. A bone marrow aspiration and biopsy was done to rule out hypersplenism but it revealed erythroid hyperplasia with reduced iron stores despite normal ferritin and iron studies. This was unusual as the patient had anemia requiring transfusions. He had no history of hemoglobinuria but a PNH by flowcytomety revealed a large clone of 81.2% in granulocytes and 88.5% in monocytes. The patient was started on Danazol and steroids for anemia which improved. He was counseled for matched sibling stem cell transplant. He had a full match with his brother. At the time of this study he awaits his transplant.

Value of CD16/CD66b/CD45 in comparison to CD55/CD59/CD45 in diagnosis of paroxysmal nocturnal haemoglobinuria: An Indian experience.

BACKGROUND & OBJECTIVES: Diagnosis of paroxysmal nocturnal haemoglobinuria (PNH), a rare haematopoietic stem cell disorder, is challenging in patients with bone marrow failure (BMF) syndrome like aplastic anaemia (AA). This study was conducted with the aim to test the efficacy of the newly recommended markers viz. anti-CD16 and CD66b antibody over the existing anti-CD55 and CD59 antibody for PNH diagnosis in India. METHODS: This study was conducted on 193 suspected cases of PNH by flow cytometry using lyse wash technique to stain the granulocytes with CD16/CD66b and CD55/CD59. RESULTS: Of the 193 suspected cases, 62 patients showed the presence of PNH clone. Forty six patients were detected by CD55/CD59/CD45, whereas 61 were detected by CD16/CD66b/CD45. CD16/CD66b detected 16 (25.8%) additional patients over CD55/CD59 (P<0.05) and was more sensitive in detecting the PNH clone with higher negative predictive value. Most of the patients (11/16) who were picked up by CD16/CD66b were of AA who had small clone sizes. Further, the PNH clones were more discreetly identified in CD16/CD66b plots than by CD55/CD59. Clone size assessed by CD16/CD66b which reflects the clinical severity of classical PNH (thrombosis/haemolysis), was more representative of the underlying clinical condition than CD55/59. INTERPRETATION & CONCLUSIONS: In our experience of 62 patients of PNH, CD16/CD66b proved to be more efficacious in detecting PNH. The new panel was especially useful in monitoring PNH associated with BMF which had small clone sizes.

The Utility of CD43 and CD200 in Differentiating Chronic Lymphocytic Leukemia from Other Mature B-Cell Neoplasms: A Cross Sectional Study.

To evaluate the utility of CD43 and CD200 in differentiating chronic lymphocytic leukemia (CLL) from other mature B-cell neoplasms. This was a cross-sectional study on patients diagnosed with B-cell neoplasms on flowcytometry. The median fluorescence intensity (MFI) of CD43, CD200 expressing neoplastic B-cells were compared between the CLL and non-CLL B-cell neoplasms followed by receiver operating characreristic curve (ROC) analysis. In addition, the sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of CD43 and CD200 in diagnosing CLL were analysed. A total of 137 patients were included. The CLL group consisted 87 patients and non-CLL group consisted 50 patients. The Mann-Whitney U test showed significant CD43 expression (U = 997.5, Z= - 5.265, p < 0.001) and CD200 expression (U = 932.0, Z = - 5.5, p < 0.01) in CLL patients compared to non-CLL patients. The area under the curve were 0.771 and 0.786 for MFI of CD43 and CD200 in differentiating CLL from non-CLL group respectively. The optimal cut-off of MFI for CD43 and CD200 were 1323 and 1775 respectively. The sensitivity, specificity, PPV and NPV of CD43 in diagnosing CLL cases were 97.7%, 66%, 83.3% and 94.2% respectively. The sensitivity, specificity, PPV and NPV of CD200 in diagnosing CLL cases were 100%, 32%, 71.9% and 100% respectively. CD43 and CD200 are useful markers in differentiating CLL from other mature B-cell neoplasms with higher MFI expression of both markers found in CLL.

Double heterozygous hemoglobin Q India/hemoglobin D Punjab hemoglobinopathy: Report of two rare cases.

Cation exchange high performance liquid chromatography (CE HPLC) provides an excellent tool for accurate and reliable diagnosis of various hemoglobin (Hb) disorders. HbQ India is a rare alpha chain variant that usually presents in the heterozygous state. Its presence in double heterozygous state with HbD Punjab is extremely rare. The double heterozygosity for α and ß chain variants leads to formation of abnormal heterodimer hybrids, which can lead to diagnostic dilemmas. We report two rare cases of double heterozygous HbQ India/HbD Punjab where the hybrid Hb was seen to elute at retention time similar to HbC on CE HPLC. The first case had unconjugated hyperbilirubinemia at presentation; while, the second case was asymptomatic.

Current state of non-hematologic cancer-associated thrombosis at a tertiary care hospital in India.

INTRODUCTION: Cancer-associated thrombosis is a leading cause of morbidity and mortality in malignancy patients. Prophylactic anticoagulation is under-utilized and the cost of low-molecular-weight heparin (LMWH) and direct oral anticoagulants is a major barrier in developing countries. MATERIAL AND METHODS: A retrospective analysis was performed of all cancer-associated thrombosis patients attending the thrombosis clinic at a tertiary-level referral hospital based in North India between 2011 and 2015. Patient demographics and disease-related parameters were collected and analyzed. RESULTS: A total of 771 patients attended the thrombosis clinic during study period, of which 64 cases were malignancy-associated. Of these, 56% of the patients were female and 20% were bedridden. The median age was 48.5 years, adenocarcinoma (48%) being the most common histological subtype. Gynecological malignancies (30%) were the most common malignancies, followed by genitourinary (11%) malignancies. Most of the cases occurred during first year of diagnosis (51%), and only 14% occurred after 3 years. Most of the patients were on combined treatment. Almost 40% of the patients developed thrombosis within 30 days of surgical treatment. Lower limb thrombosis was the most commonly seen type (56%), while abdominal and pulmonary thrombosis were both seen in 5%. Patients were managed with LMWH and vitamin K antagonists (84.3%) and only 6.25% with LMWH alone. Direct oral anticoagulants were not commonly used during the study period. DISCUSSION: At the hospital studied, most of the cases occurred early in the disease course. Postoperative prophylaxis could have contributed towards reducing thrombosis in the peri-operative period. Early suspicion and prompt treatment can improve quality of life in such patients.

Immunophenotypic Profile of Multiple Myeloma: A Tertiary Care Centre Experience.

Background Immunophenotyping and enumeration of plasma cells (PCs) by flow cytometry are deemed to be prognostically significant. However, PCs enumeration by flow cytometry is challenging owing to discrepancy with morphology and PCs loss during sample processing. Enumeration and differentiation of abnormal plasma cells (APCs) and normal plasma cells (NPCs) is difficult because abnormal antigen expression can be seen in subsets of NPCs. This is particularly true when a limited panel of antibodies are relied upon. Aims and purpose To study the immunophenotypic profile of newly diagnosed multiple myeloma (MM) cases by flow cytometry and evaluate the sensitivities and specificities of individual antigens and combinations. Methods We studied immunophenotype of PCs in newly diagnosed MM cases ( n = 48) and control cases ( n = 10) by a 6-color, 3-tube flow cytometry panel. The sensitivities and specificities of antigens in MM were evaluated and compared with control cases. Results Majority of MM cases ( n = 43) had < 3% NPCs. CD19 was the most sensitive (100%) and CD81 was the most specific marker (100%) for differentiating APCs from NPCs. CD38 MFI came out as a useful marker for APCs identification. In combination, CD19 and CD81 had a higher sensitivity and specificity to detect APCs. Conclusion NPCs may show aberrant antigen expression. A combination of multiple markers including CD81 and CD38 MFI should be used for accurate APC detection.

Prevalence and Impact of HMOX1 Polymorphism (rs2071746: A > T) in Indian Sickle Cell Disease Patients.

Introduction Fetal hemoglobin (HbF) levels play significant role in lowering down the morbidity and mortality in sickle cell disease (SCD) patients. Coinheritance of heme oxygenase-1 (HMOX1) rs2071746:A > T polymorphism may contribute to variable HbF levels in Indian SCD patients. Objective This study was aimed to evaluate the role of HMOX1 polymorphism and its impact on HbF level in Indian SCD patients. Materials and Methods One-hundred twenty confirmed cases of SCD and 50 healthy controls were recruited. Their mean age was 11.5 ± 8.6 years (range: 3-23 years). Quantification of Hb, HbA2, HbF, and HbS was done by capillary zone electrophoresis. Allele-specific polymerase chain reaction was used to genotype HMOX1 (rs2071746:A > T) gene polymorphism. Results Out of the 120 cases of SCD, 65 were hemoglobin sickle-shaped (HbSS) and 55 were sickle-beta thalassemia (Sß). Out of 65 HbSS patients, 29 (44.6%) were heterozygous (AT), 20 (30.76%) were homozygous (TT), and 16 (24.61%) were found wild-type (AA) genotype. Out of 55 Sß, 22 (40%) were heterozygous, 18 (32%) were homozygous and 15 (28%) were wild-type. Patients carrying HMOX1 (rs2071746:A > T), AT, and TT genotypes had less anemia, painful crisis, splenomegaly, hepatomegaly, jaundice, and blood transfusion. HbF level was found higher in TT genotype (in HbSS the HbF levels was 25.1 ± 4.4; in sickle-beta thalassemia the HbF levels was 36.1 ± 4.7) than wild-type(AA) and was statistically significant ( p -value <0.001). Conclusion The TT genotype of the rs2071746:A > T polymorphism was associated with increased levels of Hb F ( p < 0.001). It can serve as a HbF modifier in Indian sickle cell diseases patients.

Evaluation of Diagnostic Usefulness of CD200 Expression in B-cell Chronic Lymphoproliferative Disorders.

Immunophenotyping by flow cytometry (FCM) is a useful diagnostic tool for the evaluation of mature B-cell neoplasms (MBN). Here, CD200 expression may play a significant role and improve the distinction between various MBNs, but any potential as a prognostic marker is yet to be established. The present prospective study was conducted on all the suspected cases of MBNs. Immunophenotyping was done using a BD FACS Canto FCM using a panel of 4 to 6 color combinations of monoclonal antibodies; CD45, CD34, CD5, CD19, CD20, CD22, CD23, CD79b, FMC7, CD10, CD38, ZAP70, CD200, IgG, IgM, CD25, CD103, CD2, CD3, CD11c as well as κ and λ light chains. CD200 expression was compared in different subgroups. Of the total of 130 cases included in the study, CD200 was positive in 118 cases (90%). CD200 was expressed in 100% of the cases of CLL(86 cases), atypical CLL(06 cases), HCL(14 cases), FL(02 cases), SMZL(04 cases), LPL (01 case), and low-grade NHL (05 cases), with the highest intensity of fluorescence in HCL followed by CLL. All the cases of MCL and PLL were exclusively negative for CD200. In conclusion, the results of the present study support inclusion of this marker in the flow cytometric panels for the differential diagnosis of MBNs.

Comparison of HbA2 Using High Performance Liquid Chromatography Versus Haemoglobin Capillary Zone Electrophoresis.

Thalassemia is among the most common hereditary disorders in the world. Approximately 5% of the world's population are carriers of hemoglobinopathies, and 2.9% are carriers of beta thalassemia. Haemoglobin A2 (HbA2) constitutes less than 3% of the total hemoglobin (Hb) in adults, and the determination of Hb A2 levels is important to diagnose the beta thalassemia trait (BTT). In some cases, the level of HbA2 is not typically elevated, and some difficulties may arise in making the diagnosis. Cation exchange high-performance liquid chromatography (HPLC) and HbCZE (haemoglobin capillary zone electrophoresis) are considered acceptable methods to diagnose BTT, but these vary in their accuracy and cut-offs. In this study, we attempted to compare HbA2 values using two methods, HPLC and HbCZE, in 536 whole blood samples sent by physician-ordered hemoglobinopathy screening over two years. This included antenatal women, patients with anemia not responding to iron, and cases of familial screening where either a child or a sibling had been diagnosed with hemoglobinopathy or thalassemia. The performance characteristics of both machines were compared for the detection of the 5 most common hemoglobin variants: Hb A, HbF, HbS, Hb C, and HbE. On comparing the HbA2 values, the HPLC showed higher values for HbA2 as compared to HbCZE, while the HbF and HbS measurement agreement was good between both methods. Normal ranges and mean normal values of HbA2 differ between different methods and different manufacturers; hence, each institute using these machines should validate its cutoffs.

Study of Bone Marrow Lymphocyte Subset in Acute Myeloid Leukemia.

Introduction Acute myeloid leukemia (AML) is a heterogenous disorder consisting of clonal expansion of myeloblasts. Tumor immunity plays an important part in the pathobiology of AML. Understanding the components of tumor immunity is important for understanding tumor pathogenesis and the principles of immunotherapy. Methods We studied 41 patients with AML, for total lymphocyte, CD4 positive helper T cells, CD8 positive cytotoxic T cells, and CD16/56 positive natural killer (NK) cells proportion. Quantification was done on bone marrow aspirate sample by flowcytometry. Whenever available, post induction bone marrow was also analyzed for the lymphocyte subset. Results No significant difference was noted in the percentage of blasts among the three risk categories: favorable, intermediate, and adverse. However, there was significant difference in the total lymphocyte among the risk stratification groups, being highest in the favorable group and lowest in the adverse group. CD8 positive cytotoxic T cells were significantly less in Acute Promyelocytic Leukemia (APML) cases ( p = 0.001). Total lymphocytes were, however, more numerous in APML ( p = 0.005). NK cell proportion was not significantly different between APML and non-APML patients. On completion of induction chemotherapy, bone marrow samples for 12 patients could be processed for lymphocyte subset. On comparing the baseline against the post induction bone marrow, it was observed that there was significant increment in the proportion of CD4 positive T lymphocytes ( p = 0.046). Conclusion There is a difference in lymphocyte subset amongst patients with AML. Larger studies including functional aspects are needed to better define the role of lymphocytes in disease pathogenesis.

Thyroid Dysfunction in Patients With Immune Thrombocytopenia: Prevalence and it's Impact On Outcome.

Immune thrombocytopenia (ITP) is an autoimmune disorder characterized by isolated thrombocytopenia with or without bleeding. Mild to moderate thrombocytopenia can be seen with thyrotoxicosis and hypothyroidism. This study was conducted to assess prevalence of thyroid dysfunction in ITP and impact on treatment and outcomes. This prospective study included patients more than 12 years old, diagnosed as ITP. Serum Thyroxine (T4) and Thyroid Stimulating Hormone levels and anti-thyroid peroxidase (TPO) antibodies were done by electro- chemiluminescence immunoassay. A total of 168 patients were enrolled, with thyroid function tests available in 146 patients. Mean age was 30.6 years, with 67.8% females (n = 114). Sixty percent patients had chronic ITP and 25% had persistent ITP. Overall prevalence of thyroid disease was 25.7% in the study population, with overt hypothyroidism seen in 21 (16.4%) patients, subclinical hypothyroidism in 9 (7.0%) patients and subclinical thyrotoxicosis in 3 (2.3%) patients, while no patient had overt thyrotoxicosis. Thyroid status of the patients did not correlate with duration of ITP or response to treatment. Presence of anti TPO antibodies was associated with abnormal thyroid function and chronic ITP. The prevalence of thyroid dysfunction is increased in ITP patients. Many such cases have anti TPO antibodies, suggesting autoimmune pathology. Role of glucocorticoids, estrogens and systemic illness in these findings must be further studied before firm conclusion can be drawn about their routine inclusion in diagnostic work up.

CD157 Can Replace CD24 and CD14 in a Single-Tube Flow-Cytometric Assay to Detect Paroxysmal Nocturnal Hemoglobinuria (PNH) Clones on Both Neutrophils and Monocytes: A Prospective Study From North India.

Introduction As per current guidelines, detection of paroxysmal nocturnal hematuria (PNH) clones on leucocytes requires the demonstration of the loss of at least two glycosyl-phosphatidyl-inositol (GPI)-linked molecules on both neutrophils and monocytes by flow cytometry. CD24 and CD14 are GPI-linked molecules expressed on neutrophils and monocytes respectively, whereas another GPI-linked molecule, CD157, is expressed on both neutrophils and monocytes. This prospective study evaluated the ability of CD157 to replace both CD24 and CD14 in a single-tube flow-cytometric assay to detect PNH clones on both neutrophils and monocytes. Materials and methods PNH clones were newly detected in 52 patients by an existing "standard" single-tube six-color flow-cytometric method, which was routinely performed in our laboratory at the time of undertaking this study. Six antibodies (CD45/CD15/CD64/CD24/CD14/FLAER) were used in this "standard" technique. Subjects were divided into two groups: (i) PNH disease (n=10), and (ii) aplastic anemia/myelodysplastic syndrome (AA/MDS) (n=42). Diagnosis of PNH disease and AA/MDS were made as per standard literature and guidelines. Results were compared with a single-tube five-color "test" assay using the antibodies CD45/CD15/CD64/CD157/FLAER by flow cytometry. Samples from 20 healthy control subjects were used to calculate cut-off values for the "test" assay. Results By the "test" method, cut-off values for detecting PNH clones obtained from receiver operating-characteristic curve analysis were >0.4% for neutrophils (sensitivity=96.15%, specificity=95%), and >0.9% for monocytes (sensitivity=98.08%, specificity=95%). There was significant correlation between PNH clone sizes measured by both the "standard" and "test" assays in neutrophils (PNH disease: r=0.976, p<0.001; AA/MDS: r=0.980, p<0.001) as well as monocytes (PNH disease: r=0.806, p=0.005; AA/MDS: r=0.915, p<0.001). Bland-Altman analysis showed agreement between both assays in all the 52 patients and in individuals with AA/MDS. The cost of the test to the patients was about 15% less in the "test" method than the "standard" technique, with improved technical efficiency. Conclusion CD157 can replace both CD24 and CD14 in a single-tube flow-cytometric assay to detect PNH clones on both neutrophils and monocytes, with reduced cost to the patients and improved technical efficiency.

Evaluation of utility of immunohistochemistry markers as a tool for objective diagnosis of low-grade myelodysplastic syndrome in routine reporting: Prospective observational study.

Purpose: Diagnosis of myelodysplastic syndrome (MDS) primarily relies on the detection of morphological dysplasia in bone marrow. It is subjective and many studies have reported lack of interobserver agreement in reporting. Biopsy is preferred specimen for megakaryocyte assessment. We studied 43 bone marrow biopsies from 40 suspected MDS patient having persistent undiagnosed cytopenia. Utility of immunohistochemistry (IHC) with CD61 and p53 in detecting low-grade MDS was analyzed over routine morphology. Method and Results: Total number of megakaryocytes and number of dysplastic megakaryocytes seen on CD61 IHC was significantly higher than that on H and E stain (P value < 0.05) Out of total 43 biopsies, 13 [30.2%] cases showed dysplastic megakaryocytes that were confirmed by interobserver agreement after IHC. From 30 cases with no significant dysplasia on morphology, 21/43 [48.8%] cases showed >10% dysplastic megakaryocytes on CD61 (P value 0.0001). Nine cases showed no significant dysmegakaryopoiesis with either H and E or CD61 IHC. Fourteen cases could meet higher cut off (30%) of dysmegakaryopoiesis with CD 61 IHC. Out of total 34 cases showing significant dysplasia 7 cases (20.6%) showed positivity for p53 on IHC, which is little less than that reported in low-grade MDS. Conclusion: CD61 IHC is helpful in making correct diagnosis of MDS in cases with minimal dysplasia and should be performed before excluding possibility of MDS on morphology in a patient with undiagnosed cytopenia. IHC is cost effective tool for MDS diagnosis in developing world where access to extensive flow cytometery and molecular testing is limited.

Isolated third cranial nerve palsy as the first presentation of multiple myeloma.

A 50-year-old woman presented with a right-sided isolated third cranial nerve palsy. MRI brain showed a mass lesion arising from the right clivus with extension into the cavernous sinus. Blood investigations and bone marrow biopsy were suggestive of multiple myeloma with hypercalcaemia and renal dysfunction. It was unclear at first if the intracranial lesion was due to myelomatous involvement or a separate disease entirely. The patient declined consent for a biopsy and cerebrospinal fluid analysis was inconclusive. She was treated with bortezomib based chemotherapy and the palsy resolved by day 6, which helped clinch the rare diagnosis of central nervous system (CNS) involvement by multiple myeloma. Most patients with CNS myeloma have a dismal survival of under 6 months but she is on therapy for relapse 26 months after diagnosis. While placed under the umbrella of CNS myeloma, patients with osteodural myeloma have better outcomes, perhaps due to their distinct aetiopathogenesis.