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In Nepal, insufficient healthcare infrastructure and limited funding contribute to unmet public healthcare needs and reduced quality of care. While foreign health researchers have stepped in to support local research initiatives, their involvement has sparked ethical concerns regarding the sharing and ownership of data. This study aims to develop a locally governed framework for ethical healthcare data exchange, establish an evidence base to understand local challenges in data transfer, and to identify potential solutions for data sharing with international research teams. This cross-sectional qualitative study was conducted in Kathmandu, Nepal, using 11 multiple-choice and 12 open-ended questionnaire models. We conducted a pre-structured questionnaire survey to best identify local ethics issues related to international data transfer and proposed solutions for these challenges. The key representatives identified from the non-governmental and not-for-profit research institute (n = 14) and the life sciences society (n = 7) were invited to one-to-one blind interviews, and their recorded transcripts were coded using the QDA Miner Lite software (version 3.0) for analysis. The ratio of female to male participants was 2:3, while the ratio of junior-level staff to senior staff (≥3 years of experience in the sector) was 1:9. Approximately 42.86% of participants shared both raw and analytical data, while <5% shared no data with collaborators. Concerning knowledge, attitudes, and practices, most (38.46%) preferred open-access storage, while approximately 23.1% had limited knowledge, and 15.38% opted for confidentiality. Additionally, < 10% were in the learning process and sought training in data transfer procedures. Within this group of key representatives, participants faced main challenges in the data transfer process from four key categories: (i) the lack of standardized guidelines from government or institutes for data transfer, (ii) inadequate awareness and training in data sharing, (iii) problems related to data sharing, and (iv) problems related to biological sample transfer. In summary, this study emphasizes the importance of a standardized data-sharing platform, focusing on protecting intellectual property rights and establishing a centralized data repository in Nepal. It also recommends educational reforms, legal measures, well-defined agreements, and dedicated oversight to ensure data integrity and security, while streamlining sample transfer processes to enhance transparency and scientific progress in Nepal's research landscape.

Introduction T-cell acute lymphoblastic leukemia (T-ALL) is a genetically heterogeneous disease with poor prognosis and inferior outcome. Although multiple studies have been perform on genomics of T-ALL, data from Indian sub-continent is scarce. Methods In the current study we aimed to identify the genetic variability of T-ALL in an Indian cohort of pediatric (age ≤ 12 years) T-ALL patients ( n  = 25) by whole transcriptome sequencing along with whole exome sequencing and correlated the findings with clinical characteristics and disease outcome. Results The median age was 7 years (range 3 -12 years). RNA sequencing revealed a definitive fusion event in 14 cases (56%) (including a novel fusions) with STIL::TAL1 in 4 (16%), followed by NUP21::ABL1, TCF7::SPI1, ETV6::HDAC8, LMO1::RIC3, DIAPH1::JAK2, SETD2::CCDC12 and RCBTB2::LPAR6 in 1 (4%) case each. Significant aberrant expression was noted in RAG1 (64%), RAG2 (80%), MYCN (52%), NKX3-1 (52%), NKX3-2 (32%), TLX3 (28%), LMO1 (20%) and MYB (16%) genes. WES data showed frequent mutations in NOTCH1 (35%) followed by WT1 (23%), FBXW7 (12%), KRAS (12%), PHF6 (12%) and JAK3 (12%). Nearly 88.2% of cases showed a deletion of CDKN2A/CDKN2B/MTAP genes. Clinically significant association of a better EFS and OS ( p =0.01) was noted with RAG2 over-expression at a median follow up of 22 months, while a poor EFS ( p =0.041) and high relapse rate ( p =0.045) was observed with MYB over-expression. Conclusion Overall, the present study demonstrates the frequencies of transcriptomic and genetic alterations from Indian cohort of pediatric T-ALL and is a salient addition to current genomics data sets available in T-ALL.

Introduction Pediatric T-cell acute lymphoblastic leukemia (T-ALL) poses significant challenges due to its aggressive nature and resistance to standard treatments. Long non-coding RNAs (lncRNAs) have emerged as potential biomarkers and therapeutic targets in leukemia. This study aims to characterize the lncRNA landscape in pediatric T-ALL, identify specific lncRNAs signatures, and assess their clinical relevance. Methods RNA sequencing was performed on T-ALL patient and control samples. Differential expression analysis identified dysregulated lncRNAs and mRNAs. Functional enrichment analysis revealed potential roles of these lncRNAs in cancer pathogenesis. Validation of candidate lncRNAs was conducted using real-time PCR. Clinical correlations were assessed, including associations with patients’ clinical characteristics and survival outcomes. Results Analysis identified 674 dysregulated lncRNAs in pediatric T-ALL, with LINC01221 and CRNDE showing the most interactions in cancer progression pathways. Functional enrichment indicated involvement in apoptosis, survival, proliferation, and metastasis. Top 10 lncRNAs based on adjusted p value < 0.05 and Fold Change > 2 were selected for validation. Seven lncRNAs LINC01221, PCAT18, LINC00977, RP11-620J15.3, RP11-472G21.2, CTD-2291D10.4, and CRNDE showed correlation with RNA sequencing data. RP11-472G21.2 and CTD-2291D10.4 were highly expressed in T-ALL patients, with RP11-620J15.3 correlating significantly with better overall survival ( p  = 0.0007) at a median follow up of 32 months. The identified lncRNAs were further analysed in B-ALL patients. Distinct lncRNAs signatures were noted, distinguishing T-ALL from B-ALL and healthy controls, with lineage-specific overexpression of LINC01221 ( p  < 0.0001), RP11-472G21.2 ( p  < 0.001) and CRNDE ( p  = 0.04) in T-ALL. Conclusion This study provides insights into the lncRNA landscape of pediatric T-ALL, offering potential diagnostic and prognostic markers. RP11-620J15.3 emerges as a promising prognostic marker, and distinct lncRNAs signatures may aid in the differentiation of T-ALL subtypes. Further research with larger cohorts is warranted to validate these findings and advance personalized treatment strategies for pediatric T-ALL patients.

T-cell Acute Lymphoblastic Leukemia (T-ALL) is a highly aggressive form of ALL with at least 25% relapse rates. The high relapse rates are often linked to poor prognoses. More detailed studies for novel therapeutic targets for the treatment of T-ALL are required as the genetic and transcriptomic data currently available on T-ALL pathophysiology is insufficient. Long non-coding RNAs are emerging as important players in the regulation of tumour proliferation and metastasis. Studies on various cancers have revealed their potential as biomarkers and therapeutic targets in treatment. This review describes the characterization, biosynthesis, and role of long non-coding RNA in T-ALL and highlights their potential as next generation molecule in development of promising diagnostic, prognostic and/or therapeutic markers.

Background Polymorphisms in thiopurine methyltransferase (TPMT) and Nudix hydrolase-15 (NUDT15) have been implicated as the predominant cause of thiopurine induced leukopenia in the Western countries and East Asia respectively. Exact role of these polymorphisms in South Asian population with inflammatory bowel disease (IBD) is uncertain. Methods We included consecutive patients with IBD who were initiated on thiopurines at a center in North India. The dosage of thiopurines was titrated using regular monitoring of hemogram and liver function tests. Three TPMT polymorphisms (c.238 G > C, c.460 G > A, and c.719A > G) and one NUDT15 polymorphism (c.415 C > T) were assessed. Comparison regarding incidence of leukopenia and maximum tolerated thiopurine dosage was performed between those with wild polymorphism and those with TPMT and NUDT15 polymorphisms, respectively. Results Of the 119 patients (61 males, mean age 36.8 ± 13.5 years), 105 (88.2%) had ulcerative colitis and 14 (11.8%) had Crohn’s disease. Leukopenia was noted in 33 (27.7%), gastrointestinal intolerance in 5 (4.2%) and pancreatitis in 2 (1.6%). TPMT polymorphisms were detected amongst five patients of whom 1 developed leukopenia. NUDT15 polymorphism was noted in 13 patients of whom 7 had leukopenia. The odds of developing leukopenia in TPMT polymorphism were non-significant (0.77, 95% CI:0.0822 to 7.2134, P  = 0.819) but were significantly higher in those with NUDT15 polymorphism (3.5933, 1.1041 to 11.6951, P value: = 0.0336). Conclusion NUDT15 polymorphism was more frequent than TPMT polymorphisms and was associated with thiopurine induced leukopenia. However, the tested polymorphisms account for only 24.2% of the risk of thiopurine induced leukopenia.

Introduction: Optimal DNA and RNA quantity and purity is essential for downstream molecular biology experimentation and to avoid re-processing of sample. Despite availability of different kits and automated systems for nucleic acid isolation there is limited data on their performance evaluation, more so with pediatric blood samples, that are usually compromised in quantity. Hence, we evaluated the performance of automated QIAcube platform using pediatric blood samples in parallel with manual Qiagen extraction kits. Materials and Methods: A total of 500 samples were analyzed based on groups of PBMC and direct blood input. The isolated DNA and RNA were surveyed for quantity and quality tests by spectrophotometric and downstream analysis. Results: There was no significant difference in the DNA quantity (ng/ul) between manual and automated method based on similar sample input but quality (260/280) was significantly better with the QIAcube platform when direct blood and or PBMCs were used for extraction respectively (1.82 ± 004 Vs. 1.84.002; P-0.000008 and 1.859 ± 005 Vs. 1.843 ± 0.003; P-0.02). Moreover, the standard error mean was low for both quantity and quality in the QIAcube method suggesting uniformity. Comparison of quality assessment by spectrophotometer and qubit fluorimeter showed that QIAcube sheared DNA less (P- 0.038) as compared to manual method (P-0.013). Also, time taken to process the samples in QIAcube was 23% less than the kit-based method. Conclusion: Overall analysis of QIAcube platform suggests that it yields more better, uniform, and less-sheared quality of nucleic acid in a relatively less time as compared to manual extraction kits.

Carrageenan has been proved as potent growth promoting substance in its depolymerized form. However, relatively little is known about its role in counteracting the adverse effects of drought stress on plants. In a pot experiment, lemongrass (Cymbopogon flexuosus Steud.), grown under different water stress regimes [(100% field capacity (FC), 80% FC and 60% FC)], was sprayed with 40, 80 and 120 mg L-1 of gamma irradiated carrageenan (ICA). Foliar application of ICA mitigated the harmful effects of drought stress to various extents and improved the biochemical characteristics, quality attributes and active constituents (citral and geraniol) of lemongrass significantly. Among the applied treatments, ICA-80 mg L-1 proved the best in alleviating detrimental effects of drought. However, drought stress (80 and 60% FC), irrespective of the growth stages, had an adverse impact on most of the studied attributes. Generally, 60% FC proved more deleterious than 80% FC. At 80% FC, application of ICA-80 mg L-1 elevated the essential oil (EO) content by 18.9 and 25%, citral content by 7.33 and 8.19% and geraniol content by 9.2 and 8.9% at 90 and 120 days after planting (DAP), respectively, as compared to the deionized-water (DW) spray treatment (80% FC+ DW). Whereas, at 60% FC, foliar application of 80 mg L-1 ICA significantly augmented the EO content by 15.4 and 17.8% and active constituents viz. citral and geraniol, by 5.01 and 5.62% and by 6.06 and 5.61% at 90 and 120 DAP, respectively, as compared to the control (water-spray treatment).

The dysregulation of m6A-related genes recognized as ‘writers’, ‘readers’, and ‘erasers’ is reported to be involved in the initiation, progression, and drug resistance of acute myeloid leukemia (AML). In the present study, we investigated the expression levels of various readers, writers, and erasers in pediatric AML patients. Additionally, we categorized the patients according to the molecular subtyping of common mutations and recurrent fusions and correlated the expression of m6A-associated genes with different molecular subtypes and evaluated their prognostic and clinical implications. A total of fifty-seven patients with pediatric de novo AML were enrolled in the study. The study cohort consisted of 41 males and 16 females with a median age of 7 years (range 1 to 12 years). A high expression of m6A RNA modification complex genes was noted in AML patients. Among the writers, METTL3 and METTL14 were found to be upregulated in 19 and 17 patients, the readers YTHDF1 and YTHDF2 showed higher expression in 6 and 10 patients, while a high expression of erasers FTO and ALKBH5 was found in 28 patients and 1 patient, respectively. Further, the expression of m6A regulators showed a significant association with genetic alterations including FLT3-ITD, RBM15::MKL fusions and NPM1 mutations. Additionally, while evaluating the prognostic implications, both the readers YTHDF1 and YTHDF2 showed a significant correlation with TLC at diagnosis (p < 0.05). Further, Kaplan–Meier estimation showed a poor event-free survival in cases with the overexpression of YTHDF1 (log-rank p = 0.028). Additionally, we noted a strong correlation between YTHDF1 overexpression and treatment-related mortality (log-rank p < 0.001), and a nearly significant correlation with YTHDF2 expression in such patients (log-rank p = 0.053) at a median follow-up of 8 months. Thus, our data suggest that m6A genes, especially readers YTHDF1 and YTHDF2, are involved in the disease prognosis of AML and probably function in an integrated manner with other m6A-modifying genes to subsequently play a role in AML pathogenesis.

Methods: Forty pediatric (0–12 years) B-ALL DNA samples (20 paired Diagnosis-Relapse) and an additional six B-ALL DNA samples (without relapse at 3 years post treatment), as the non-relapse arm, were retrieved from the biobank for advanced genomic analysis. Deep sequencing (1050–5000X; mean 1600X) was performed using a custom NGS panel of 74 genes incorporating unique molecular barcodes. Results: A total 47 major clones (>25% VAF) and 188 minor clones were noted in 40 cases after bioinformatic data filtering. Of the forty-seven major clones, eight (17%) were diagnosis-specific, seventeen (36%) were relapse-specific and 11 (23%) were shared. In the control arm, no pathogenic major clone was noted in any of the six samples. The most common clonal evolution pattern observed was therapy-acquired (TA), with 9/20 (45%), followed by M-M, with 5/20 (25%), m-M, with 4/20 (20%) and unclassified (UNC) 2/20 (10%). The TA clonal pattern was predominant in early relapses 7/12 (58%), with 71% (5/7) having major clonal mutations in the NT5C2 or PMS2 gene related to thiopurine-dose response. In addition, 60% (3/5) of these cases were preceded by an initial hit in the epigenetic regulator, KMT2D. Mutations in common relapse-enriched genes comprised 33% of the very early relapses, 50% of the early and 40% of the late relapses. Overall, 14/46 (30%) of the samples showed the hypermutation phenotype, of which the majority (50%) had a TA pattern of relapse. Conclusions: Our study highlights the high frequency of early relapses driven by TA clones, demonstrating the need to identify their early rise during chemotherapy by digital PCR.

Here, we have studied in p53 null H1299 lung carcinoma cells, the dominant-negative effect of human p53 (h-p53) on buffalo p53 (b-p53) induced nuclear transactivation-dependent function. Recently, we have isolated and cloned the full-length cDNA of buffalo p53. Buffalo and human p53 proteins exhibit a high degree of structural and functional similarities. In transiently transfected H1299 cell line b-p53 appeared to be more sensitive to Mdm2-mediated degradation as compared to h-p53, although its ability to transactivate p21 promoter was stronger than that of the human counterpart. This higher transactivation ability of b-p53 was lost in the presence of h-p53 suggesting, a dominant-negative effect of h-p53 on b-p53’s transactivation of p21 promoter. Both human and buffalo p53 proteins could hetero-oligomerize but the b-p53 could tetramerize much faster than the h-p53. A chimeric cDNA construct of human p53 was made where the 1–260 bp N-terminus was replaced with buffalo p53 counterpart and expressed in H1299 cell line. The tetramerization ability of the chimeric p53 protein was comparable to that of h-p53. Properties of b-p53 like stronger p21 transactivation and super sensitivity to Mdm2 mediated degradation were lacking in the chimeric protein. Thus, it is suggested that faster ability of tetramerization as well as higher transactivation property of buffalo p53 is determined by the interplay of N- and C-terminal domains through macromolecular interactions.