Computational and mechanistic studies on the effect of galactoxyloglucan: Imatinib nanoconjugate in imatinib resistant K562 cells.

Imatinib mesylate, a BCR/ABL fusion protein inhibitor, is the first-line treatment against chronic myelogenous leukemia. In spite of its advantageous viewpoints, imatinib still has genuine impediments like undesirable side effects and tumor resistance during chemotherapy. Nanoparticles with sustainable release profile will help in targeted delivery of anticancer drugs while minimizing deleterious side effects and drug resistance. The use of biopolymers like galactoxyloglucan (PST001) for the fabrication of imatinib mesylate nanoparticles could impart its use in overcoming multidrug resistance in chronic myelogenous leukemia patients with minimal side effects. This study involved in the synthesis of PST-Imatinib nanoconjugates with appreciable drug payload and excellent cytotoxicity against drug-resistant chronic myelogenous leukemia cell line (K562) in comparison with free drug. The use of bioinformatics tool revealed better binding affinity for the drug-polysaccharide complex than the drug alone with three proteins: 3QX3 (Topoisomerase), 1M17 (EGFR tyrosine kinase domain), and 3QRJ (ABL1 kinase domain). Assessment of the biochemical, hematological, and histopathological parameters in mice upheld the security and adequacy of the nanoconjugate compared to free drug. Although perspective investigations are warranted, in a condition like drug resistance in leukemia, this nanoconjugate would display a productive approach in cancer therapeutics.

Jehovah's Witness and consent for blood transfusion in a child: The Indian scenario.

In India, the clinical guidelines and laws governing consent for blood transfusion in a minor are meager and vague. In an elective situation, whether the parents can make a decision for the child on his/her behalf or whether the doctor has the right to make the decision in the best interests of the child is not clear. We present the case scenario of a child belonging to Jehovah's Witness denomination diagnosed with Burkitt lymphoma. His parents were in a dilemma whether to opt for blood transfusion or not. In the absence of laws and guidelines in this context, and considering the complications that he developed during the treatment period, it was very challenging for us to manage the situation both medically and medico-legally. This situation highlights the need for framing consensus guidelines/laws regarding elective blood transfusion in a minor to make health-care delivery, smooth, transparent and flawless.

Comparative diagnostic test evaluation of serum procalcitonin and C-reactive protein in suspected bloodstream infections in children with cancer.

PURPOSE: To compute diagnostic test properties of C-reactive protein (CRP) and serum procalcitonin (PCT) levels in bloodstream infections in children with cancer and suspected sepsis, in comparison with blood culture as the gold standard. METHODOLOGY: Consecutive paediatric cancer patients, aged ≤14 years, with clinically suspected bloodstream infections were evaluated with blood culture and assay of PCT and CRP levels. Blood culture was taken as the gold standard for comparison. Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), likelihood ratio (LR) and receiver operating characteristic (ROC) with area under ROC curve (AUC) were calculated to assess the diagnostic test performance for PCT and CRP.Results/Key findings. The ROC curve for PCT was better than that for CRP, with an AUC of 0.751 for PCT at a cut-off of 2.25 ng ml-1. The AUC for CRP was 0.638 at a cut-off of 8.0 mg dl-1. Among the three cut-off values of PCT selected from the ROC curve applicable to the patients under study, the cut-off value of ≥0.49 ng ml-1 had the maximum sensitivity of 81.4 % and an NPV of 94.67 %; ≥2.25 ng ml-1 had a sensitivity and specificity of 65.12 and 71.6 %, respectively, and ≥6.47 ng ml-1 had a maximum specificity of 82.10 %. For CRP, the cut-off value of ≥5.3 mg dl-1 had the maximum sensitivity of 72.09 %; ≥8.0 mg dl-1 had a sensitivity and specificity of 58.14 and 68.09 %, respectively, and ≥8.4 mg dl-1 had the maximum specificity of 70.04 %. CONCLUSION: PCT is a better serological marker for excluding bloodstream infections than CRP. The cut-off value of 0.49 ng ml-1 with a negative predictive value of 94.67 % will be ideal in a clinical setting of immune-compromised children with suspected sepsis.

Profiling gene mutations, translocations, and multidrug resistance in pediatric acute lymphoblastic leukemia: a step forward to personalizing medicine.

Precise risk stratification and tailored therapy in acute lymphoblastic leukemia (ALL) can lead to enhanced survival rates among children. Translocations and mutations along with multidrug resistance markers are important factors that determine therapeutic efficacy. Gene mutation profiling of patients at the time of diagnosis can offer accurate clinical decision-making. Multiplex PCR was used to screen for various translocations, mutations, and P-glycoprotein (P-gp) status in pediatric ALL samples. The roles of P-gp were analyzed at the transcriptional and translational levels by using real-time PCR and immunoblotting, respectively. ALL specific cell line Jurkat was used to validate the functional role of P-gp in imparting drug resistance by siRNA knockdown studies. Co-occurrence of translocations and mutations contributes to cellular drug resistance. Presence of any translocation in addition to FLT3/ITD hints for overactive P-gp. Co-occurrence of E2A/PBX and TEL/AML has also been positively correlated with P-gp status. Multiplex PCR provides a rapid and cost effective technique for profiling translocations, mutations, and multidrug resistance status that determines what therapy patients could be administered. Mutation profiling in patients for analyzing genetic lesions along with drug resistance profiling will help improve risk stratification and personalized medicine, thereby increasing the treatment success rates among pediatric patients with leukemia.

DNA repair gene XRCC1 polymorphisms in childhood acute lymphoblastic leukemia.

Defective DNA repair has been reported to be a risk factor for various malignancies. Genetic polymorphisms of DNA repair genes are thought to result in different phenotypic features compared to the wild type. Genetic polymorphisms in XRCC1 gene could, through alteration of protein structure, lead to defective functioning of DNA Polbeta, PARP and LIG3 enzymes resulting in defective DNA repair and increased risk of childhood acute lymphoblastic leukemia (ALL). The role of DNA repair gene XRCC1 in susceptibility to childhood ALL has, however, not been widely studied and no data exists from Indian children. In this pilot study, through the use of PCR and RFLP, further confirmed by DNA sequencing, we have shown an increased risk of ALL among children with XRCC1 codons 194 and 399 variant genotypes. Among the three variants, only the association between codon 399 variant and risk of ALL appeared to be significant. The risk of ALL was higher in males with codons 194 and 399 polymorphisms than in females. However, no relation was found between the presence of these variant genotypes and treatment outcome.

Synchronous Occurance of Acute Myeloid Leukemia and Rhabdomyosarcoma.

Metachronous primary distinct tumors are frequently and increasingly encountered in oncology clinical practice of recent times, but synchronous tumours are still a rarity. We report an unusual case of a 2 year old male child who had synchronous occurrence of rhabdomyosarcoma of pelvis and acute myeloid leukemia.Our search of literature suggests that this may be the first reported case of simultaneous occurrence of these two malignancies.

Argyrophilic nuclear organizer regions (AgNORs) in pediatric acute lymphoblastic leukemia.

Nuclear organizer regions (NORs) code for ribosomal RNA and are associated with non-histone nucleoproteins which can be identified by silver staining (AgNORs). AgNORs have been correlated to proliferative activity of tumors and hence may be prognosis-related. The present study evaluates the possible prognostic importance of the AgNORs in tumor cells of patients with pediatric acute lymphoblastic leukemia (ALL). A significant increase in AgNOR counts was observed in ALL patients as compared to normal controls. Further, a significant positive correlation was observed between AgNOR counts and total WBC count at diagnosis. A negative correlation was also observed between AgNOR counts and age of the patients. Logistic regression analysis revealed further correlation between AgNOR counts and disease with an odds ratio of 1.29 (P = 0.03) as compared to normal controls. These results therefore suggest that AgNOR counts may be significant in the evaluation of pediatric ALL.

Mutant p53 protein, Bcl-2/Bax ratios and apoptosis in paediatric acute lymphoblastic leukaemia.

PURPOSE: The Bcl-2 family of proteins regulates a late step in the apoptosis pathway. Bcl-2 protein is believed to be involved in imparting resistance to programmed cell death or apoptosis induced by chemotherapeutic agents and radiation. The anti-apoptotic function of the Bcl-2 protein appears to be modulated by its ability to heterodimerize with other members of the gene family, predominantly Bax, a protein favouring induction of apoptosis. Susceptibility to undergoing apoptosis may, therefore, be dependent on the ratio between Bcl-2 and Bax. Both Bax and Bcl-2 are regulated by the tumour-suppressor protein p53. The present study therefore aims to study the significance of the Bcl-2:Bax ratio, p53 expression and apoptosis in paediatric acute lymphoblastic leukaemia (ALL). METHODS: Expression of Bax, Bcl-2 and p53 was determined by immunocytochemistry, and apoptosis was evaluated by an enzymatic end-labelling technique using biotin-dUTP and further confirmed by annexin binding. The presence of mutant p53 was determined using a mutant-p53-specific enzyme-linked immunosorbent assay (ELISA). RESULTS: A total of 32 cases and 20 controls were evaluated. Bcl-2 was found to be expressed in 22/32 of the ALL cases. Pretreatment (spontaneous) apoptosis was observed in 23/32 cases. The mean pretreatment apoptotic index was 11.34 +/- 2.04% with a median value of 7.5%. CONCLUSIONS: There was a positive correlation between apoptosis and Bax expression (r = 0.5044; P = 0.0038). There was good correlation between the immunoreactivity of p53 and detection of mutant p53 by ELISA (r = 0.4605; P = 0.0079). The apoptosis index showed a negative borderline correlation to the expression of Bcl-2 protein (r = -0.3181; P = 0.076). There was an inverse correlation between extent of apoptosis and the presence of mutant p53 protein (r = -0.4732; P = 0.006). p53 protein expression also showed a correlation with both Bcl-2 (r = 0.4647; P = 0.007) and Bax (r = 0.4128; P = 0.018). The Bcl-2/Bax ratio, however, showed no significant correlation with apoptosis (r = -0.3131; P = 0.08) or with p53 expression. No significant association was evident between clinical and laboratory parameters with the Bcl-2/Bax protein expression except lymphadenopathy (r = 0.5774; P = 0.03). However, Bax expression showed a borderline correlation with the immediate tumour response to chemotherapy (r = -0.338; P = 0.0628). These patients are being followed-up to look for any association between clinical outcome, Bcl-2/Bax ratio and apoptosis.

Increased levels of mutagen-induced chromosome breakage in Down syndrome children with malignancy.

Even though an association between Down syndrome (DS) and malignancies has been established, the mechanism behind this is still unclear. We therefore investigated constitutional chromosomal abnormalities and bleomycin-induced chromosome sensitivity in 12 DS children, 8 DS children with malignancies, and 10 normal controls to explore whether these factors play any role in cancer predisposition. Trisomy 21 was the only constitutional cytogenetic abnormality observed in all the DS children. But there was significant variation between the patients and controls with regard to bleomycin sensitivity. Compared to the normal controls, all the DS patients expressed significantly higher chromosomal breaks per cell (b/c) values indicating sensitivity to bleomycin. Furthermore, DS children with malignancies demonstrated significantly higher b/c values than DS children with malignancies. These results permit us to assume that DS children showing mutagen hypersensitivity may be having defective DNA repair competence and hence may be predisposed to malignancies.

Bcl-2 protein and apoptosis in pediatric acute lymphoblastic leukemia.

Many chemotherapeutic agents are thought to exert their genotoxic effects through induction of programmed cell death (PCD) (apoptosis) in tumor cells. The bcl-2 is an anti-apoptotic oncoprotein and can confer a survival advantage to tumor cells by preventing apoptosis. Overexpression of bcl-2 may therefore be implicated in resistance to chemotherapy. We studied the significance of bcl-2 expression and the PCD index in pediatric acute lymphoblastic leukemia. Evaluation of bcl-2 by immunocytochemistry and PCD by an enzymatic end labelling technique using biotin-dUTP was carried out in a total of 55 cases and 40 controls. Bcl-2 was found to be expressed in 47% (26/55) of the acute lymphoblastic leukemia cases. The positive cells varied from 0-49% among individual samples. Pre-treatment (spontaneous) apoptosis was observed in 62% (34/55) cases. The mean pre-treatment PCD index was 8.27 1.3%, while the median PCD index was 5. The PCD value for the leukemic samples analyzed were then classified as either high apoptosis values ( 5) and low apoptosis values (<5). PCD index was high in 53% (29/55) and low in 47% (26/55). However, 23% (13/55) of cases did not show presence of either apoptosis or bcl-2. There was no association between clinical and laboratory parameters with the apoptotic index or bcl-2 protein expression. However, evaluation of apoptotic index and bcl-2 expression on day 7 of induction chemotherapy showed a borderline correlation between these markers and initial WBC count, presence of mediastinal mass and hepatosplenomegaly. Follow-up of these patients is being done to look for any association between treatment response and apoptosis.

Extragonadal germ-cell tumors: a ten-year experience.

Extragonadal germ-cell tumors (EGCT) are uncommon and biologically distinct from their gonadal counterparts. Thirty-seven patients who had EGCT were treated over a ten-year period at the Regional Cancer Centre, Trivandrum, India. There were 26 men and boys and 11 women and girls. The sites of primary tumor were mediastinum (n=18), central nervous system (n=5), sacrococcygeal region (n=4), retroperitoneum (n=2), and other sites (n=8). After combined modality therapy, 13 of 18 patients who had mediastinal EGCT--1 of 2 with retroperitoneal, 1 of 4 with sacrococcygeal, 0 of 5 with central nervous system, and 2 of 8 patients with tumor in other sites-were alive with no evidence of disease at a median follow-up of 16 months. The overall 5-year survival rate was 40%. Histologic subtype and elevated marker levels were the significant prognostic factors on univariate analysis.

Disseminated Langerhans cell histocytosis in children: treatment outcome.

Langerhans cell histiocytosis is an interesting disorder with a variety of presentations and variable outcomes. This study evaluates response to treatment, recurrence, and survival in disseminated Langerhans cell histiocytosis treated at Regional Cancer Centre, Trivandrum, India from 1983 through 1994. Thirty-five patients with disseminated Langerhans cell histiocytosis were seen. Six had group A disease, 21 had group B disease, and eight had group C disease. In group A, five of six patients are disease free at a median follow-up of 48 months. Two had recurrence after initial treatment, which was salvaged. In group B, 13 of 15 patients had complete response after chemotherapy, nine of whom experienced recurrence later. Three of five patients who received irradiation alone experienced recurrence. One died of progressive disease. Two patients were lost to follow-up. Seventeen of 20 are alive with median follow-up of 67 months. In group C, one of eight patients are alive after multiple recurrences. Of the surviving patients, 29% had significant sequelae. In summary, group A and B patients do well with treatment, and most of the recurrences can be salvaged. A significant proportion of patients have sequelae. Newer aggressive protocols must be developed for treating group C patients. Measures to prevent sequelae must also be developed.

Expression of folate sensitive and aphidicolin induced chromosomal fragile sites in familial neuroblastoma.

The expression of folate sensitive and aphidicolin induced fragile sites in the blood lymphocyte chromosomes of affected and unaffected members from 2 neuroblastoma families were studied. The subjects included 4 neuroblastoma patients, and 9 of their clinically healthy first degree relatives and corresponding number of age and sex matched controls. Lymphocytes cultured in folate deprived culture medium showed rare fragile sites at band p13.1 of chromosome 1, in a frequency of 3%-5% in all the 4 neuroblastoma patients. In aphidicolin treated cultures, the patients and unaffected members in neuroblastoma families, showed hypersensitivity to aphidicolin, as evidenced by the significant increase in percentage of aberration/cell (ab/c) and damaged cells (dc), over that of controls (P < 0.01). Aphidicolin induced fragile sites were more pronounced in chromosomes 1 and 2. A larger number of subjects have to be studied to prove whether altered fragile site expression may be a cytogenetic evidence for an individual or familial cancer predisposing genetic constitution.

Clear cell sarcoma of kidney (CCSK)--a clinicopathological study of 4 cases.

The clinical and histopathological features of four cases of clear cell sarcoma of kidney (CCSK) or Bone metastasising renal tumour of childhood (BMRTC) are presented. These cases were identified among 107 primary renal tumours in childhood over a period of 15 years (1973-1987) in the Trivandrum Medical College. Of the 107 cases 96 (89.7 percent) were nephroblastomas and 7 (6.6 percent) were Mesoblastic Nephromas. The incidence of Clear cell sarcoma was 3.7 percent. Abdominal mass and haematuria were the most common clinical features. All the four cases occurred in male children with no predilection for the right or the left kidney. At the time of presentation bone metastasis was not present in any of the four cases. Metastasis to scapula and skull was detected ten months after nephrectomy in one case. Of the four patients three were in stage I disease at the time of diagnosis. All the four cases showed the typical gross morphology and the classic microscopic pattern of Clear Cell sarcoma kidney. The treatment was similar in all the four cases with Surgery followed by radiotherapy and chemotherapy (Vincristine, Adriamycin Actinomycin D and cyclophosphamide). Only one of the four patients is alive and well 12 months after surgery. The literature is reviewed along with a discussion of the gross pathology, histology and histogenesis of clear cell sarcoma of kidney.

Malignancies in Down syndrome.

Down Syndrome (DS) is associated with an increased incidence of malignancies, especially leukaemias. We came across 8 DS children presenting with malignancies and having trisomy 21 as the sole cytogenetic abnormality. Of these 8 DS cases, 4 presented with acute lymphocytic leukaemia, 2 with acute myeloid leukaemia and one case each with Hodgkin's disease and Wilms' tumour. There are contradictory reports regarding the distribution of myeloid versus lymphoid malignancies in DS children and their response to therapy. The exact mechanism by which patients with DS are predisposed to develop malignancies is unclear. However, presence of the extra chromosome no. 21 is presumed to disrupt the genetic balance which increases generalized susceptibility to genetic and environmental trauma. Furthermore, an increased methotrexate toxicity observed in these patients should also be taken into consideration in designing treatment for DS children with malignancies.

Pattern and outcome of neuroblastoma. A 10 year study.

OBJECTIVE: To determine the clinical pattern and outcome of children with neuroblastoma. DESIGN: Retrospective analysis. SETTING: Hospital based information from the case sheets. SUBJECTS: 91 children with age upto 14 years treated in the Regional Cancer Center, Trivandrum. METHODS: Clinical presentation, metastatic pattern and treatment outcome were analyzed. RESULTS: Median age of the study group was 2.5 years with a male:female ratio of 1.6:1. Fifteen per cent children had early stage disease and 85% advanced disease. Five children with stage III and 22 with stage IV disease did not receive may active treatment. The remaining 64 children were evaluated for this analysis with a survival rate of 25%. Two of the four (50%) stage I, four of the six (83.3%) stage II, five of the 18 (27.7%) stage III, three of the 32 (9%) stage IV and two of the four (50%) stage IVs patients are long term survivors. CONCLUSIONS: Majority of the children (> 80%) presented with advanced disease and outcome remained poor. Only 27% of stage III and 9% of stage IV patients are long term survivors in our series. In future, approaches to detect disease at an early stage and aggressive therapeutic strategies in selected patients may improve survival.

Rare presentation of pediatric acute promyelocytic leukemia as multiple lytic bone lesions: case report and review of literature.

Acute promyelocytic leukemia (APL) is an uncommon malignancy in the pediatric population, accounting for only 5-10% of pediatric acute myeloid leukemias, and for this disease to present with bone lesions at diagnosis is extremely unusual. We wish to convey that very rarely, in a pediatric cancer patient presenting with multiple extensive lytic bone lesions, the diagnosis can be APL. The treatment protocol and prognostic implications are vastly different. Histopathology is the gold standard in arriving at a correct diagnosis and delivering proper treatment in such cases. This patient had excellent response to chemotherapy.

Precocious puberty as initial presentation in mediastinal tumour.

Numerous disorders can cause precocious puberty in children, and germ cell tumours (GCT) are one of the rare causes . We report two cases of mediastinal malignant GCTs who presented with precocious puberty. Both patients had bulky and advanced disease, were aggressively treated with neo-adjuvant chemotherapy and surgery, and are surviving and free of disease.