Cytogenetic Profiling In Paediatric Acute Leukaemia; A Report On 746 Newly Diagnosed Paediatric Cases Analyzing The Spectrum Of Recurring Chromosomal Rearrangements In B Cell Lymphoblastic And Acute Myeloid Leukaemia.

Background: Cytogenetics is evolving and different molecular mechanisms we know now have proved to be of diagnostic and prognostic significance in both acute lymphoid (ALL) and myeloid leukaemia (AML). This study aims to find out and compare the occurrence of different cytogenetics in paediatric acute leukaemia. Methods: This is a cross-sectional study of diagnosed B-ALL and AML patients presenting at The Indus Hospital. We studied FISH and karyotype in BALL and FISH in AML patients. FISH analysis shows a total of 69 (12.8%) of B ALL patients had cytogenetic abnormalities. BCR-ABL1 was positive in 5.1%, ETV6/RUNX1T1 in 8.6% and KMT2A in 2.3% individuals. Karyotype reveals hyper diploidy in 24.3%, Monosomy in 1.94%, and t (1:19) and t (17:19) were observed in 5.8% and 0.24% cases respectively. FISH analysis in AML cases reveal positivity of t (8:21) in 26.4%, INV (16) in 6.1% while PML-RARA t(15:17) was done on morphological suspicion in 17 cases; all of which showed positivity; making 7.9% of the total AMLs. The study demonstrated a wide spectrum of heterogeneity in paediatric acute leukaemia. Conclusion: Hyperdiploidy was the most common cytogenetic abnormality. We report a lower incidence of t (12:21), compared to the world. We showed a higher prevalence of RUNX1/RUNX1T1 in young children. The prevalence of core binding factor AML was 32.5%.

Infantile B-lymphoblastic leukemia: a case series and review of the literature.

Infantile leukemia is a rare hematological malignancy that occurs in the first year of life. It is an aggressive disease with peculiar immunophenotypic, cytogenetic, and molecular characteristics. It can be myeloid or lymphoid in origin. More than 80% of cases involve KMT2A gene rearrangement in the lymphoblastic subset, versus 50% in the myeloid subset. In this study, we present three cases of this rare entity to add knowledge about its clinical presentation and diagnostic profiles. These cases of infantile B-lymphoblastic leukemia (B-ALL) were retrospectively reviewed at the Department of Hematology, Section Cytogenetics at Indus Hospital and Health Network. The clinical characteristics, complete diagnostic profile, immunophenotypic profile, fluorescence in situ hybridization (FISH) results, treatments, and outcomes of the patients were assessed. All three infants were girls who presented with hyperleukocytosis, and they were diagnosed by eight-color flow cytometry. FISH studies revealed KMT2A gene rearrangement in two of the three patients. Infantile B-ALL is a biologically distinct disease carrying a poor prognosis. Female preponderance, hyperleukocytosis, and hepatosplenomegaly are common findings in this subgroup. No standard protocol for this rare entity has proven ideal for managing these young infants.

Spectrum of hemoglobinopathies with hematological and biochemical profile: A five year experience from a tertiary care hospital.

Background & Objective: Determination of hemoglobinopathies is significant for epidemiological studies. There is a need to identify burden of hemoglobinopathies at national level to lay down the foundation of appropriate screening and prevention programs. The present study aimed to evaluate the spectrum of hemoglobinopathies along with hematological and biochemical parameters in a tertiary care hospital. Methods: This retrospective study included results of high performance liquid chromatography (HPLC) test from July 2015 - May 2020 in the department of Hematology, Indus Hospital and Health Network, Karachi, Pakistan. Data of all patients collected for red blood cell (RBC) indices, serum iron profile, and vitamin B12 and red cell folate levels. Diagnosis of hemoglobinopathies was done by an automatic analyzer ADAMS A1C Model No. HA-8180T Arkray/Japan. Results: Among 2422 participants, hemoglobinopathy observed in 14.5% (n=352). Beta thalassemia trait is observed as the most common hemoglobinopathy (6.4%). Severe anemia (Hb=5.1-5.5 g/dl) found in beta thalassemia major (BTM) and HbE disease. Red cell parameters showed significant association with different types of hemoglobinopathies. Mean ferritin level was high in E-beta thalassemia (687.8±591.9) followed by sickle cell disease (615.7±543.5). Conclusion: Apparently, overall frequency is static however, results of this study are not applicable to general population due to sample bias. Moreover, true figures are difficult to identify due to high incidence of iron deficiency anemia that masks the diagnosis by conventional techniques. Molecular characterization by DNA analysis is the most reliable tool of diagnosis. However, this method is not widely available in our country due to lack of expertise and cost issues.

T-cell Acute Lymphoblastic Leukemia in an Infant: A Case Report and Review of Literature.

A high index of suspicion of acute leukemia in infants presenting with atypical clinical and laboratory features is of paramount importance to avoid the delay in diagnosis, and treatment of this rare, yet extremely aggressive entity. We report a case of a 10 months infant presenting with a one-month history of fever and restlessness along with complete blood count (CBC) findings of bicytopenia, borderline leukocytosis, leucoerythroblastic picture, and reactive lymphocytes. Considering infection, the patient was kept on antibiotics and symptomatic treatment. However, the persistence of findings led to bone marrow aspirate which revealed T-cell acute lymphoblastic leukemia (T-ALL) on flow cytometry. Infant ALL is almost B cell phenotype and T-ALL is hardly ever reported. Knowledge and recognition of these cases can help to improve awareness regarding consideration of this rare phenotype of leukemia irrespective of clinical presentation and age of the patient. Key Words: Acute lymphoblastic leukemia, Infant leukemia, Flow cytometry.

Comparison of Clinical and Diagnostic Features of Pediatric Oncology Patients With or Without COVID-19 Infection: A Retrospective Chart Review.

There is a scarcity of data summarizing the clinical picture, laboratory, and imaging findings and outcome in children with malignancy and coronavirus disease 2019 (COVID-19) infection. This study characterizes a detailed comparison of pediatric oncology patients with and without COVID infection. A retrospective study was conducted at The Indus Hospital, Karachi, from March 2020 to June 2020. Clinical presentation, laboratory and imaging findings, disease severity, and outcome were compared between cohorts. The mean age of children with and without COVID was 8.0±4.9 and 7.4±4.1 years, respectively. Hematologic malignancy comprised the largest number of patients, followed by solid tumors. Lymphocytosis and low neutrophil-lymphocyte ratio was observed in the COVID positive group. Cardiac dysfunction (1.4% vs. 0%), acute respiratory distress syndrome (8% vs. 0%) and lower peripheral capillary oxygen saturation/fraction of inspired oxygen ratio (473 vs. 486) found to be associated with severe disease in COVID positive group (P<0.05). Overall mortality in children with COVID was 6.8% versus 2.7% in children without COVID. Pediatric patients with malignancy have different clinical features and laboratory parameters as compared with children without malignancy. Acute respiratory distress syndrome, absolute lymphocytosis and low neutrophil-lymphocyte ratio is associated with severe disease in children with malignancy and COVID infection. In contrast to adults, biochemical markers and complete blood count parameters do not help recognize COVID infection in pediatric patients with malignancy.

An Outcome Analysis of Childhood Acute Promyelocytic Leukemia Treated with Atra and Arsenic Trioxide, and Limited Dose Anthracycline.

The overall survival of Acute Promyelocytic Leukemia (APL), reported in recent studies, is approaching to 90% wherein, arsenic trioxide (ATO) and all-trans retinoic acid (ATRA) are used as the mainstay of treatment with either limited or no use of anthracycline and cytarabine. This study is aimed to ascertain the outcome of children with APL using similar approach. A total of 30 patients with APL, registered from January 2015 to December 2018, were reviewed. Diagnosis was established on bone marrow aspirate and confirmed by the presence of PML-RARA translocation. Treatment protocol was based on Australian APML 4 study performed by Australian Leukemia Lymphoma Group (ALLG). Lumbar puncture was not performed as it was not part of the protocol due to the risk of bleeding. The mean age in current cohort was 9 years with 53% males. Seven (23.3%) patients died and three (10%) abandoned treatment during induction. Twenty patients completed the intensive phase of chemotherapy and all (100%) of them attained molecular remission (MR). One patient dropped out after MR whereas, 19 remain on follow up with no evidence of disease, reflecting disease free survival (DFS) of 95%. With a median follow up of 2.5 years (range 2.1-4.8 years) the 5 years Kaplan-Meier estimate of OS was 63% and 73%, with and without abandonment, respectively. Analysis of outcome according to risk groups revealed inferior outcome of high risk (HR) group (38% and 50% with and without abandonment, respectively) in contrast to standard risk (SR) group which showed better outcome (82% and 88% with and without abandonment, respectively). The attainment of 100% molecular remission and absence of relapse supports the effectiveness of this regimen. Moreover, it is found to be less toxic and therefore, can be conveniently managed in day-care settings.

Philadelphia Chromosome Positive B-lymphoblastic Leukemia in an Infant.

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Distribution of subtypes and immunophenotypic characterization of 1379 cases of paediatric acute leukaemia.

OBJECTIVES: Acute leukaemia is the most common and highly curable childhood malignancy; subtyping and identification of antigens via immunophenotyping helps in treatment plan as well as minimal residual disease monitoring. METHODS: This retrospective study was conducted at the Haematology section of the clinical laboratories of Ziauddin University Hospital and The Indus Hospital, Karachi conducted at January 1st, 2012 to December 31st, 2017. The study included 1379 cases of de novo acute leukemia from 2012 to 2017. Among these, 80% were diagnosed by using four color flowcytometry (FACS Calibur), 9% and 11% via immunohistochemistry on bone marrow trephine biopsy samples and morphological examination respectively. RESULTS: The mean age of patients was 7.4 ± 4.3 years while male to female ratio was 1.75:1. Lymphoblastic leukaemia accounted for 77.2% and myeloid leukaemia 21.2%. Amongst lymphoblastic lineage, B-ALL was 80.4% while T-ALL was 19.6%. Among the phenotypic expression of B-ALL, CD79a (99.8%) had the highest positivity. In B-ALL, CD13 (29.8%) was the most common aberrant myeloid marker. Aberrant expression of CD79a observed in 11.1% of T-ALL cases. In non APL AML, aberrant expression of CD79a and CD19 was observed in 6.6% and 5.5% of cases respectively. CONCLUSION: Overall immunophenotypic profile, expression of aberrant phenotypes and subtype distribution in our patients was similar to international literature except for a relatively high frequency of T-ALL which was discordant from the western data.

Safety and Efficacy of Aminophylline in Intrathecal Methotrexate-related Neurological Toxicity in Large Pediatric Oncology Centre.

Children with acute lymphoblastic leukemia, receiving intrathecal methotrexate (IT-MTX), develop acute and subacute neurological dysfunction in 3-15% of cases. Altered level of consciousness, seizure, and stroke-like manifestations are among the most common presentations. MRI of the brain in an early stage is consistent with white matter leukoencephalopathy. There are no specific guidelines for the treatment of such a syndrome. An elevated cerebrospinal fluid (csf) adenosine concentration causes vasodilatation in the brain and leads to cerebral ischemia, which may be reduced by aminophylline infusion. The study is a retrospective data analysis in which electronic data records of 30 patients, collected from September 2017 - August 2019. The primary objective of the study was to evaluate the safety and efficacy of aminophylline in IT-MTX induced neurotoxicity. Aminophylline infusion was used in 30 patients with IT-MTX neurotoxicity. Twenty-five patients (83.33%) showed dramatic improvement of neurologic signs and symptoms within 48 hours. Aminophylline was, hence, coined as a salvageable therapy. No noticeable side effects were observed during treatment with aminophylline infusion. Key Words: Intrathecal methotrexate, Neurotoxicity, Children, Cancer, Aminophylline.

The Effects of Voriconazole on Metabolism of All-Trans Retinoic Acid in the Treatment of Acute Promyelocytic Leukemia: A Case Report.

All-trans retinoic acid (ATRA) is a derivative of vitamin A and is the mainstay treatment of acute promyelocytic leukemia (APL). Hypercalcemia is a rare yet important side-effect of ATRA, especially when it is used concomitantly with a medication that impedes its metabolism by inhibiting cytochrome P-450 in the liver and thus increasing the duration of exposure to ATRA. Azole antifungal drugs such as voriconazole are frequently used in patients undergoing chemotherapy due to a high incidence of fungal infections. These medications inhibit two vital enzymes of cytochrome P-450, CYP2C9 and CYP3A4, potentiating the effects of ATRA on calcium metabolism. We present a case of a nine-year-old girl who underwent chemotherapy with all-trans retinoic acid for acute promyelocytic leukemia. The patient was given an anti-fungal cover with voriconazole for extensive fungal chest infection simultaneously. She was found to have asymptomatic hypercalcemia on routine follow-up during the consolidation phase. Both medications were stopped. Subsequently, she was admitted to the ward and managed conservatively with hydration. Serum calcium levels were returned to normal within six days after stopping the combination of ATRA and voriconazole. We underscore that the use of anti-fungal medications should be limited while using ATRA. However, strict monitoring must be done when a combination of these drugs is started, if necessary.

Central Nervous System Involvement in Childhood Acute Lymphoblastic Leukemia: An Analysis of Day-One Versus Day-Eight Lumbar Punctures in Remission Induction Therapy.

Introduction Traumatic lumbar puncture (TLP+) can lead to the iatrogenic infiltration of the central nervous system (CNS) by circulating leukemic blast cells in childhood acute lymphoblastic leukemia (ALL). The risk of TLP+ is increased by a number of factors at the time of presentation of the disease, such as a high white cell count (WCC), T-ALL phenotype, and unstable clinical condition of the patient. For this reason, the first lumbar puncture (LP) was deferred until Day Eight of prednisolone prophase during remission induction therapy in one set of patients. The objective was to compare the historical cohort of Day-One LP with Day-Eight LP with respect to the incidence of TLP+ and de novo CNS leukemia. Methods A retrospective comparative data analysis of 1,185 childhood ALL patients aged 1-16 years was conducted based on the electronic medical records of the pediatric hematology-oncology department of The Indus Hospital (TIH), Karachi, from January 2010 to August 2018. A total of 600 patients whose LP was done on Day One (January 2010-May 2015) were placed in cohort A, whereas 585 patients whose LP was performed on Day Eight (June 2015-August 2018) were placed in cohort B. After the examination of the cerebrospinal fluid (CSF), the status of CNS infiltration was classified as CNS-1, CNS-2, CNS-3, and TLP+. Results A total of 1,185 patients were included in the study, of whom 600 patients were in cohort A and 585 patients in cohort B. The incidence of TLP+ was found to be lower in cohort B (1.7%) as compared with the incidence in cohort A (4.3%) (p-value=0.009). However, there was an increase in the incidence of CNS-3 cases in cohort B (8%) as compared to cohort A (3%) (p-value: <0.001). When the CNS status of both the cohorts was compared with that of the internationally published data, a low incidence of TLP+ cases was noted in patients with LP on Day Eight. Conclusion The modified approach of performing the first LP on Day Eight significantly reduced the incidence of TLP+ cases. However, an unusual finding of a significant increase in the CNS-3 leukemia was noted. More prospective studies are needed to investigate this significant increase in CNS-3 cases.

Burkitt Lymphoma With Aberrant Immunophenotype Imposing Diagnostic Challenge.

BACKGROUND: Burkitt lymphoma (BL) exhibits a characteristic immunophenotype that is positive for pan-B-cell antigens and germinal center markers while negative for immature markers. A deviation from classic immunophenotype can cause diagnostic confusion and might result in false exclusion of BL. In some cases, overlapping clinical, morphological and immunophenotypic features of BL and B lymphoblastic lymphoma (B-LL) can be of diagnostic challenge. However, definitive delineation is of paramount importance due to difference in treatment. We describe a case of BL in a child with atypical features including absence of L3 morphology in diagnostic tissue and aberrant expression of CD34, CD99 and BCL2 on immunohistochemistry. These findings led to the interpretation of B-LL which was later on excluded by detection of t (8;14). This unorthodox case not only highlights the importance of cytogenetic testing but also emphasizes the correlation of all the diagnostic tools before making a definitive diagnosis. Therefore, reporting this case will help in eliciting the high index of suspicion among pathologists for this exceptionally unusual immunophenotype.

Hypercalcemia and Disseminated Osteolytic Lesions With Normal Blood Counts and Absence of Circulating Blasts: A Rare Presentation of Childhood B-Lymphoblastic Leukemia.

Hypercalcemia and disseminated osteolytic bone lesions are a rare presentation of pediatric acute lymphoblastic leukemia (ALL). The authors report a 3-year-old boy who presented with hypercalcemia and diffuse osteolytic lesions involving axial and appendicular bones. He had normal complete blood count and the absence of blasts in peripheral smear; however, bone marrow aspirate and trephine were consistent with B-cell ALL. A review of the literature highlights the variable clinical outcome of this rare presentation depending on the presence of hypercalcemia and osteolytic lesions with or without chromosomal translocation t(17;19) and coagulation abnormalities. The patient had no coagulopathy and normal karyotype, and showed excellent response to initial treatment in terms of complete remission and negative minimal residual disease after standard-risk induction chemotherapy. Hypercalcemia with diffuse osteolytic lesions warrants bone marrow examination to rule out leukemia even in the absence of any abnormality in complete blood count. The case was reported for awareness of this rare presentation of ALL so that delays can be avoided for this potentially curable but life-threatening disease.

Outcome of Core Binding Factor Acute Myeloid Leukemia in Children: A Single-Center Experience.

Childhood acute myeloid leukemia (AML) harboring core binding factor (CBF)-associated translocations are considered as a favorable cytogenetic subgroup. The 2 major subtypes of CBF-AML include t(8;21) and inversion of chromosome 16, accounting for ∼25% of patients. Because of expensive and toxic treatment, which may require hospitalization during the entire course of induction chemotherapy, most of the centers in Pakistan neither workup for this low-risk entity nor offer curative treatment. Therefore, we adopted an approach of screening AML cases for the presence of CBF with the rationale of offering curative treatment to this subgroup. Data of 244 cases were reviewed, and translocations were found in 72 (34%) patients among them, 59 (82%) had t(8;21) and 13 (18%) showed inversion of chromosome 16. The event-free survival with and without abandonment was 36% and 40%, respectively. Among 44 patients who completed treatment, 26 (59%) are leukemia-free, while 18 (41%) relapsed. None of the relapsed patients received salvage chemotherapy or hematopoietic stem cell transplant. Treatment-related mortality and abandonment was found in 24% and 10% of patients, respectively. The frequency of CBF-AML is higher in our study; however, poor outcome demands holistic measures in supportive care to improve the survival.

8p11 associated myeloid/lymphoid neoplasms: An illustrative case report of this rare entity and review of literature.

This report describes a unique case of 8p11 myeloproliferative syndrome (EMS), also known as stem cell leukaemia-lymphoma syndrome. A 13 years old male was referred from a tertiary care hospital after cervical lymph node biopsy. The disease mechanism of this neoplasm is to either evolve into acute myeloid leukemia or mixed lineage leukaemia, and less frequently of T or B lymphoid lineage. However, here we show a case of a rare simultaneous presentation of T lymphoblastic lymphoma on cervical lymph node and B lymphoblastic leukaemia on bone marrow biopsy along with t (8; 13) on karyotype testing.

Minimal residual disease in childhood B Lymphoblastic Leukemia and its correlation with other risk factors.

OBJECTIVE: To determine frequency of post induction and post consolidation minimal residual disease (MRD) in pediatric B-lymphoblastic leukemia (B-ALL) patients and its association with clinical risk factors. METHODS: This is a retrospective, cross sectional study carried out at the Indus Hospital on paediatric patients (1-17 years) was performed from May 2015 to January 2018. On day 35, MRD testing was done on bone marrow aspirate using four color flow cytometer with 0.01% cut off. Positive cases were retested at post consolidation. Data was collected for demographics, total leukocyte count (TLC), central nervous system status (CNS), Cytogenetics for BCR-ABL, MLL, TEL-AML by FISH and prophase response then analyzed in association to MRD status. RESULTS: Out of 362 patients, 133 (37%) were post induction MRD positive, with no statistically significant association to age, gender, TLC, CNS status, prophase response, BCR-ABL and TEL-AML1. However, MLL showed closely significant association (p-value=0.05). Post consolidation, 49 (44%) were MRD positive; age, National cancer institute (NCI) risk groups and CNS status showed statistical significance (p-value <0.05). CONCLUSION: Despite high frequency of MRD positivity, significant association is not observed between post induction MRD and risk factors. However, post consolidation MRD has a significant association with NCI risk groups, age and CNS status.

Childhood Core Binding Factor (CBF) acute myeloid leukaemia and its association with French American British (FAB) classification.

OBJECTIVE: To find the frequency of core binding factor acute myeloid leukaemia in our population, and to determine its association with morphological subtypes. METHODS: The retrospective study was conducted at The Indus Hospital, Karachi, and comprised data of patients aged 1-17 years who were diagnosed with acute myeloid leukaemia from July 2013 to June 2017. Data was analysed using SPSS 21. RESULTS: Of the 237 patients, 137(58%) were males and 100(42%) were females. The overall mean age was 8±4.34 years. Cytogenetic testing had been performed in 212(89.45%) cases, and core binding factor was detected in 72(34%) cases. There was significant difference between the mean values of white cell count and the subtypes (p=0.000). Also the difference between core binding factor and the subtypes was significant (p=0.000). CONCLUSIONS: There was found to be a significant association of core binging factor with specific subgroups of acute myeloid leukaemia.

Prednisolone Prophase for a Week Versus Upfront Multiagent Chemotherapy in Childhood Acute Lymphoblastic Leukemia: An Analysis With Reference to Induction Mortality in a Developing Country.

In childhood acute lymphoblastic leukemia, high treatment-related mortality, especially in the induction phase of treatment, is a major challenge for developing countries. The reasons are multifactorial, including a late presentation with higher disease burden, malnourishment, and limited support services. These factors may aggravate the toxic effects of upfront multiagent chemotherapy in terms of severe neutropenic sepsis and tumor lysis. Therefore, instead of upfront chemotherapy, we offered prednisolone prophase for 1 week with the objective of balancing the antileukemic versus the toxic effect of treatment. The data of 538 patients who received induction with this approach (cohort B) are compared for induction mortality with previous records of 438 patients (cohort A) treated with upfront chemotherapy. In the presence of similar clinical characteristics including age, sex, risk group, and phenotype in both cohorts, a significant difference was found in overall induction mortality of 9% in cohort B versus 14% in cohort A (P<0.05). This difference was also significant in the high-risk and T-cell phenotype, which strengthens our hypothesis that patients with higher burden of disease may experience more fatal toxic effects with upfront intensive chemotherapy. Therefore, we suggest that the prednisolone prophase approach is beneficial to control the disease with less severe toxic effects in our settings.