The HLA-G molecule is expressed in few tissues and cells, such as erythroid precursors, in which it has been associated with cell signaling, maturation, and proliferation. Considering that the daily production of erythrocytes is massive, the bone marrow sHLA-G levels are higher than in peripheral plasma, and little attention has been devoted to the role of HLA-G in disorders presenting with defective erythropoiesis, we hypothesized that bone marrow could be a major source of sHLA-G to plasma. We investigated HLA-G (soluble-sHLA-G and membrane-bound) in patients presenting hematologic disorders with impaired erythropoiesis (n = 61, including myeloproliferative neoplasm-MPN, myelodysplastic neoplasm-MDS, aplastic anemia-AA, inherited bone marrow failure-BMF, secondary erythrocytosis-SE), and in healthy individuals. Indeed, sHLA-G levels (ELISA) in bone marrow of healthy individuals were higher compared to patients (p < 0.001), and we observed an overall positive correlation between bone marrow and peripheral blood sHLA-G (p < 0.05). The logistic regression showed that the lower sHLAG plasma bone marrow levels the greater the probability of developing impaired erythropoiesis (p < 0.001). In MDS patients and controls, HLA-G (flow cytometry) was expressed on myeloid precursor cells, followed by erythroid precursors and proerythroblasts, and was not expressed on differentiated erythroid cells. The use of recombinant erythropoietin (rEPO) in MDS patients (n = 3) increased the expression of HLA-G in myeloid precursors, in which HLA-G was normally expressed. Decreased bone marrow sHLA-G level is a shared feature among patients with impaired erythropoiesis and rEPO may increase HLA-G cell expression in MDS patients, corroborating the role of HLA-G along erythrocyte development.

FLT3 inhibitors have become a cornerstone in the treatment of FLT3-mutated acute myeloid leukemia (AML), however, durable clinical responses are frequently limited by the emergence of acquired resistance. In this study, we established and comprehensively characterized a quizartinib-resistant FLT3-ITD AML model to elucidate the molecular and functional mechanisms underlying therapeutic failure. Prolonged exposure of MV4-11 cells to escalating concentrations of quizartinib resulted in the selection of quizartinib resistant clones (MV4-11QR), displaying an increase in IC50 and a shift from cytotoxic to predominantly cytostatic responses. Resistant cells maintained MAPK signaling despite FLT3 inhibition. Global proteomic profiling revealed extensive reprogramming, with enrichment of pathways related to energy metabolism, RNA processing, and translational regulation, accompanied by enhanced mitochondrial respiration and glycolytic capacity. Whole-genome sequencing identified acquisition of the FLT3D835H mutation and clonal expansion of TP53R248W with loss of the wild-type TP53 allele, indicating strong treatment-driven clonal selection. Functionally, MV4-11QR cells showed broad cross-resistance to clinically relevant agents, including midostaurin, venetoclax, and cytarabine. Importantly, pharmacological targeting of mutant p53 with eprenetapopt or MAPK signaling with trametinib restored sensitivity to quizartinib, inducing synergistic or additive cytotoxic effects and increased apoptosis. Together, these findings define a multilayered resistance program involving genetic, signaling, and metabolic adaptations and support rational combination strategies to overcome FLT3 inhibitor resistance in AML.

Introduction Insulin Receptor Substrate 1 (IRS1) is differentially expressed in hematological neoplasms suggesting a role in hematopoiesis and neoplastic transformation. Irs1 knockout mice represent a tool to investigate IRS1 function. This study compared hematological parameters of wild-type and heterozygous Irs1S57X mice and assessed fetal lethality in homozygous mice. Methods Hematological parameters were analyzed monthly in wild-type and heterozygous mice from 8 to 22 weeks of age. Successive intercrosses failed to yield homozygous knockouts. Fetal lethality was evaluated through timed matings of heterozygous mice, with genotyping performed at various gestational stages (E9.5, E12.5, E15.5 and E18.5). Results Heterozygous mice showed no significant differences in body weight or hematological parameters compared with wild-type mice (all p-value >0.05). Homozygous Irs1S57X mice exhibited fetal or postnatal lethality as fetuses developed until gestational stage E18.5 but were either aborted or died shortly after birth. Conclusion The Irs1S57X mutation in heterozygosis does not alter phenotype, whereas homozygosity for Irs1S57X is associated with markedly reduced perinatal survival, precluding adult hematopoiesis studies. Future research should focus on fetal hematopoiesis.

Primary myelofibrosis (PMF) is a chronic myeloproliferative neoplasm characterized by the activation of the JAK-STAT pathway. Previous evidence showed that metformin might be a possible therapeutic option for treating JAK2-mediated myeloproliferative neoplasms. In vitro and in vivo studies demonstrated that metformin inhibits the JAK-STAT pathway, induces apoptosis in JAK2V617F-positive cell lines and reduces tumor burden and splenomegaly in Jak2V617F knock-in-induced mice. The FIBROMET trial, an open label phase II study, evaluated metformin effects on 10 primary myelofibrosis patients over 2 years of treatment. Primary endpoint was bone marrow fibrosis reduction. Secondary endpoints were constitutional symptoms, blood counts, spleen size modulation and exploratory evaluation of protein and gene expression. Metformin treatment reduced bone marrow collagen deposits, downregulated the STAT pathway and reduced the p85 subunit of PI3K enzymatic complex, together with endothelial maintenance genes, in PMF patients. These results raise new evidence regarding metformin, a cheap and widely available drug, as a possible adjuvant for the treatment of PMF patients.

Next-generation sequencing (NGS) has transformed the diagnostic and prognostic landscape of acute myeloid leukemia (AML) by enabling the simultaneous detection of multiple clinically relevant mutations. However, access to this technology remains limited in many Latin American countries due to economic and infrastructural constraints. This study reports the implementation and validation of an NGS workflow for AML molecular profiling within the International Consortium on Acute Leukemias (ICAL), emphasizing feasibility and reproducibility across reference laboratories in Brazil. iSeq100 sequencers (Illumina) were donated to ICAL centers. Laboratory personnel received training through the American Society of Hematology's Visitor Training Program at Erasmus MC (Rotterdam). Genomic DNA from 15 AML patients enrolled in the ICAL-2015 study was analyzed using a custom amplicon-based panel targeting FLT3, NPM1, IDH1, IDH2, ASXL1, CEBPA, TP53, and RUNX1. Results were compared between the 2 Brazilian laboratories, a European reference laboratory, and conventional single-gene testing. NGS demonstrated positive interlaboratory concordance for variant allele frequencies (Pearson r = 0.72). Concordance between NGS and PCR-based methods reached 77%, with discordant cases involving FLT3-ITD detection and the assessment of ASXL1 mutations. Mutation frequencies were consistent with those reported in European and North American cohorts. The present study demonstrates that simplified NGS workflows can be successfully established and quality-controlled in resource-constrained settings through international collaboration, training, and standardized protocols.

Autophagy induction has recently emerged as a mechanism of resistance to FLT3 inhibitors (FLT3i) in patients with FLT3-ITD mutant acute myeloid leukemia (AML). Here, we assessed the molecular mechanisms of autophagy inhibition associated with FLT3i and its impact on cell survival and pharmacological resistance. In FLT3-ITD AML cell lines (MOLM13 and MV4-11), treatment with first- and second-generation FLT3i (midostaurin and quizartinib, respectively) induced autophagy. Combining FLT3i with autophagy inhibitors further decreased cell viability and increased cell apoptosis in both cell lines and in primary patient samples. Label-free quantification proteomics of MOLM13 cells revealed that RFC4 (Replication Factor C Subunit 4), an autophagy regulator linked to increased chemosensitivity, and GATD3/C21orf33 (Glutamine Amidotransferase Class 1 Domain Containing 3) proteins were upregulated only in the combined group, while 11 proteins mostly associated with chemoresistance were downregulated. In vivo, the combination of midostaurin and autophagy inhibition improved overall survival in MOLM13-transplanted mice. ATG5- (Autophagy Related 5) and ATG7-knockdown (Autophagy Related 7) increased sensitivity to first- and second-generation FLT3i in MOLM13 cells. To investigate the potential of autophagy inhibition in overcoming FLT3i resistance, we generated MV4-11 cells resistant to quizartinib (MV4-11QR). The resistant cell line exhibited higher basal levels of autophagy compared to the parental cell line. The combination of quizartinib and chloroquine demonstrated a synergistic effect in MV4-11QR cells and this effect was associated with greater inhibition of the FLT3 receptor compared to the monotherapies. Therefore, combining FLT3i with autophagy inhibition enhances the FLT3i antileukemic efficacy and overcomes pharmacological resistance.

Introduction: Allogeneic hematopoietic stem cell transplantation is a viable therapeutic option for several serious diseases however it is a high-risk procedure because it involves high-toxicity protocols with many adverse effects. Existing factors, such as the underlying disease and nutritional status, may influence the outcome. The objective of this study was to evaluate the Nutritional Risk Index as a prognostic tool by correlating it with body mass index, nutritional status, and clinical outcomes in patients undergoing hematopoietic stem cell transplantation. Methods: This single center retrospective study was conducted collected sociodemographic, anthropometric, biochemical, and clinical data before conditioning and 30 days post-transplantation. Statistical analyses were performed using the Mann-Whitney test and Spearman's correlation. Overall survival was estimated using the Kaplan-Meier method, with comparisons conducted via the Gehan-Breslow-Wilcoxon test. A Cox Proportional Hazards regression analysis was employed to identify factors associated with mortality; variables demonstrating a p-value ≤0.20 in the univariate analysis were included in the multivariate model. For all analyses, statistical significance was defined as a p-value <0.05. Results: Seventy-seven participants were included, with an average age of 41 years. According to the nutritional risk index, the entire sample was classified as having severe nutritional risk. The body mass index showed that 6.4 % were malnourished, 19.4 % were obese, and 12.9 % had hypoalbuminemia. The estimated survival curve identified a significant difference for patients aged <45 years with survival being significantly longer (p-value = 0.01). Higher albumin levels (≥3.5) after transplantation were associated with longer survival (p-value = 0.04). Sex, body mass index, albumin level before conditioning, and graft-versus-host disease showed no significant differences in terms of survival. Albumin levels ≥3.5 g/dL after transplantation were marginally associated with a lower mortality risk and malignant disease showed a trend toward increased mortality. Conclusion: These findings underscore the clinical utility of prognostic indices, such as the Nutritional Risk Index and albumin levels, during the pre-transplant period, emphasizing the necessity for early nutritional interventions in hematopoietic stem cell transplantation patients.

The tumor microenvironment plays an important role in cancer onset and progression. Its significance has been increasingly addressed in myeloid neoplasms. Such a shift marks a departure from the usual tumor-centered approach to a more comprehensive and integrated understanding of the interplay between myeloid tumors and their surroundings.On this note, we aimed to summarize, in this review, an up-to-date take on how a pro-inflammatory milieu influences clonal selection of genetically altered hematopoietic stem cells towards myeloid malignancies at the expense of their healthy counterpart; the role of NLRP3 inflammasome, a major component of the innate immunity and source of interleukin-1β, over acute myeloid leukemia development and performance; and, alongside post-translational modifications, how autophagy represents a major NLRP3 inflammasome regulator.

Autophagy has recently been postulated as a potential mechanism that may contribute to the development of resistance to FLT3 inhibitors (FLT3i). Our aim was to investigate the role of pharmacologic autophagy inhibition in enhancing the efficacy of FLT3i in the FLT3-ITD acute myeloid leukemia (AML) resistant model and its impact on the survival and molecular response to FLT3i. In MOLM13 and MV4-11 (FLT3-ITD mutated cell lines), the administration of FLT3i first (midostaurin) or second (quizartinib) generation significantly elevates autophagic flux by inducing the formation of acidic vesicular organelles visualized by flux cytometry. We also observed a reduction in negative autophagy regulators, evidenced by the inhibition of AKT (p-Ser473), mTOR (p-Ser2448), and its downstream protein p70S6K (p-Thr421/Ser424), as well as a reduction of ULK1 (p-Ser757) by western blot assay. FLT3i treatment resulted in the conversion of LC3B-I to LC3B-II, accompanied by the degradation of LC3B-II and p62, indicating autophagy induction. The combination of FLT3i with autophagy inhibitors chloroquine (CQ), bafilomycin A1 and ROC-325 significantly reduced cell viability and survival in comparison with vehicle and monotherapies in vitro in MOLM13 and MV4-11 cells and ex vivo in primary cells from bone marrow of FLT3-ITD AML patients. To elucidate the role of inhibition of autophagy in FLT3i-resistant cells, we generated MV4-11 cells resistant to quizartinib (MV4-11 QR) trough selective pressure and we noted that MV4-11 QR cells exhibited a higher basal level of autophagy compared to conventional MV4-11 cells, as indicated by the increased formation of acidic vesicular organelles, and confirmed by the reduced expression of LC3B and p62 proteins. In MV4-11 QR cells, we observed that CQ effectively reduced the IC50 of quizartinib and increased FLT3i-induced apoptosis, suggesting its use to mediate resistance to FLT3i in AML. We also observed an increased molecular effect of quizartinib when associated with CQ, manifested by a significant increase in Cleaved-Caspase3 and a reduction of STAT5 (p-Tyr694) and P70S6K in comparison with vehicle and quizartinib monotherapy. The combination of quizartinib with CQ showed a strong synergistic effect in the resistant lineage demonstrated by a ZIP score value = 10.8, while in the non-resistant lineage it demonstrated an additive effect (ZIPscore = 8.4), suggesting that CQ is autophagy-dependent and has its effect potentiated in conditions of high autophagic flux, as observed in MV4-11 QR, being effective, mainly, in conditions of resistance generated by the induction of autophagy. In conclusion, FLT3i induces autophagy flux, limiting drug inhibitory activity, and autophagy inhibition can enhance the antileukemic effect of FLT3i and reverts cellular resistance. Analysis of the autophagic profile in AML can play a crucial role in individualized treatment. Manuela Albuquerque de Melo, Diego A. Pereira-Martins, Brunno G. Macedo, Priscila S. Scheucher, João A. Machado-Neto, Fabíola Traina. Autophagy inhibition overcomes resistance to FLT3 inhibitors in FLT3-ITD acute myeloid leukemia models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4447.

BACKGROUND:Hereditary angioedema (HAE) is a rare autosomal dominant disorder with a prevalence of 1:50,000 individuals. Delayed diagnosis and deaths from asphyxia still occur. OBJECTIVE:To identify knowledge and management gaps regarding clinical, genetic, and therapeutic aspects of HAE in Brazil, aiming to improve patient care and outcomes. METHODS:A Brazilian multicenter HAE registry was established, with patients' data included by treating physicians using the REDCap (Research Electronic Data Capture) platform. RESULTS:Of the 820 patients with HAE enrolled, 68.8% were female. Most (72.4%) experienced HAE due to C1 inhibitor deficiency (HAE-C1INH), whereas 19.4% had HAE with normal C1INH caused by variants in the F12 gene (HAE-FXII). Onset of symptoms occurred earlier in HAE-C1INH as compared with HAE-FXII (mean 11.2 years vs 19.4 years, respectively), and time for diagnosis was shorter in patients younger than 18 years, as compared with those 18 years and older (mean 1.8 years vs 14.5 years, respectively). Regarding treatment, 52.8% received first-line on-demand therapies (icatibant or plasma-derived C1INH [pdC1INH]). Only 4.8% used first-line options for long-term prophylaxis (LTP), such as lanadelumab or subcutaneous/intravenous pdC1INH. Attenuated androgens were used for LTP in 52% of patients, with adverse effects reported for 34.8%. CONCLUSIONS:Brazilian patients with HAE share common aspects with global patients, including predominance in women, and HAE-C1INH as the most common subtype. Available genetic testing allowed for identification of a notable proportion of HAE-FXII (19.4% of the patients). Despite recent advances, access to first-line therapies for LTP of HAE attacks remains limited.

Venetoclax is a promising alternative for patients with acute myeloid leukemia who are considered unfit for conventional chemotherapy; however, its employment still faces challenges mostly related to drug resistance. Here, we provide further biological mechanisms underlying the previously described and potentially novel intrinsic sources of poor response to venetoclax departing from ex vivo response data. Acute myeloid leukemia data including FLT3 mutation status, gene expression data, and ex vivo response data were extracted from the publicly available BeatAML 1.0 study database and aided sample categorization that supported differential gene expression analysis that, in turn, supported gene set enrichment analysis. CIBERSORTx-based bulk RNA sequencing deconvolution of BeatAML 1.0 data allowed us to categorize samples according to their cell type content. We observed that inflammation-related gene sets, such as cytokines and inflammatory response, NLRP3 inflammasome activation, and activation of adaptive immune response, were concordantly positively enriched across all the conditions reported to be associated with poor ex vivo venetoclax response, whereas samples from good ex vivo responders’ mostly enriched gene sets related to mitochondrial activity, and early myeloid progenitor cell molecular programs. Besides the alternative reliance on BCL2A1, we highlight inflammation as a common element present across multiple sources of venetoclax ex vivo response modulation in acute myeloid leukemia samples. Hence, a potential key modulator for venetoclax response.

Metabolic reprogramming is a hallmark of cancer, with acute myeloid leukemia (AML) being no exception. Mitochondrial function, particularly its role in protecting tumor cells against chemotherapy, is of significant interest in AML chemoresistance. In this study, we identified mitochondrial DNA content (mtDNAc), measured by quantitative PCR, as a simple and precise marker to stratify the metabolic states of AML patients. We show that patients with high mtDNAc are associated with increased mitochondrial metabolism and a higher dependency on oxidative phosphorylation (OXPHOS), often correlating with chemoresistance. Clinically, patients receiving cytarabine and an anthracycline-based regimen (7 + 3 regimen) experienced inferior relapse-free survival and a higher overall rate of leukemia recurrence. Ex vivo experiments using primary AML samples confirmed cytarabine resistance in high mtDNAc patients, which could be overcome by inhibiting mitochondrial complex I. The FDA-approved drug metformin, which targets mitochondrial metabolism, significantly enhanced apoptosis in response to chemotherapy or targeted agents, such as venetoclax, in AML models. However, metformin-treated cells adapted by increasing glycolysis and NAD+ production, a resistance mechanism that could be bypassed by targeting the nicotinamide phosphoribosyltransferase (NAMPT) enzyme. In summary, we demonstrated that mtDNAc is an effective tool for assessing the metabolic state of AML cells. This method can be easily implemented in clinical practice to identify chemoresistant patients and guide personalized treatment strategies, including novel combination therapies for those with a high reliance on mitochondrial metabolism.

FLT3 mutations contribute to leukemogenesis and poor prognosis in acute myeloid leukemia (AML). Despite FLT3-targeted therapies, relapse rates remain high, suggesting other mechanisms of persistence. FLT3-ITD promotes leukemic stem cell survival and may alter the bone marrow microenvironment, leading to immune evasion. We hypothesized that FLT3-ITD allelic burden and clonal dominance affect immune status and treatment response. We analyzed NK and T cell immunophenotypes in a Flt3-ITD mouse model and in human AML samples to investigate this dysfunction. In the murine model, the Flt3 mutation was associated with reduction in NK cell (CD45hiCD19-CD3-NK1.1+) content characterized by predominance of immature phenotypes (CD27+CD11b-) and a reduction in cytotoxic subpopulations (CD27-CD11b+). Although these alterations were observed in both heterozygous and homozygous animals as compared to wild-type controls, they were more pronounced in the homozygous group. These results were corroborated by an extended phenotypic analysis using CD122, NK1.1, CD49b, and NKp46 expression to identify four NK cell more primitive maturation stages. Among these, a significant reduction in the frequency of more mature subsets (NKII to NKIV) was observed in Flt3-mutated homozygous mice, whose NK cells exhibited an increased expression of DNAM-1, a key activating receptor, and TIM-3, an inhibitory checkpoint receptor involved in immune regulation. In the T cell compartment, a significant increase in short-lived effector cells (CD127-KLRG1+) was observed among CD4⁺ T cells specifically in homozygous animals. In the CD8⁺ T cell population, homozygous animals showed an increased frequency of central memory (CD62L+CD44+) and memory precursor effector cells (CD127+KLRG1-), accompanied by elevated expression of activation and inhibitory receptors, including NKG2D, TIM-3, and CTLA-4, highlighting altered immune regulation within this subset. Furthermore, γδ T cells were expanded in homozygous mutant animals compared to wild-type mice. These results suggested that the Flt3 mutation content influenced the immune profile and inspired us to study NK cells in AML samples of different stages of leukemia arrest (HSC-L, MPP-L, CMP-L, GMP-L, MP-L, and GP-L) driven by their FLT3-ITD/NPM1 mutational status. Multiparametric flow cytometry analysis of bone marrow samples from 148 de novo AML patients with normal karyotype and known FLT3/NPM1 status revealed that FLT3mut/NPM1wtpatients (n=36) were predominantly associated with CMP-L (CD34+/-CD117+CD13+CD33+/-HLA-DR+MPO+) and GMP-L (CD34+/-CD117+/-CD13+/-CD33+HLA-DR+MPO+) leukemias, whereas FLT3wt/NPM1mut patients (n=80) predominantly presented GP-L (CD34-CD117+/-CD13+CD33+HLA-DR-MPO+) leukemias. Double-mutant patients (FLT3mut/NPM1mut, n=32) exhibited a similar profile to FLT3wt/NPM1mut patients, with a predominance of MP-L and GP-L stages. As compared to other maturation stages of leukemia arrest, GMP-L leukemias were associated with disease persistence after the first induction therapy and a trend toward earlier relapse, although impact on overall or relapse-free survival was not observed. In agreement, in silico analysis using the Beat AML 2.0 dataset indicated higher resistance to cytarabine, midostaurin, and azacitidine in patients with GMP-L FLT3mut/NPM1wt cells. No differences were observed in the frequencies of CD56+ NK cells, CD56bright, or CD56dim subsets among the FLT3wt/NPM1mut, FLT3mut/NPM1mut, and FLT3mut/NPM1wt groups. However, in FLT3-mutated patients, high AR showed a trend toward fewer CD56dim and more CD56bright NK cells compared to low AR. Regarding leukemic maturation, a trend toward increased CD56bright and decreased CD56dim populations was observed in more differentiated leukemias (GMP-L, MP-L, and GP-L), suggesting functional dysregulation linked to stage of leukemia arrest. Collectively, our findings show that FLT3-ITD promotes dysregulation in maturation and activation of cytotoxic immune cells, enabling immune evasion. Immature leukemic subsets are linked to impaired cytotoxic differentiation and altered receptor expression, possibly contributing to resistance and poor outcomes. Immunophenotypic alterations in mouse and human models support the potential of targeted immunotherapy and emphasize the need to consider leukemic maturation and immune status in therapeutic strategies.

The Hodgkin lymphoma International Study for Individual Care (HoLISTIC) Consortium's A-HIPI model, developed in 2022 for advanced-stage classical Hodgkin lymphoma (cHL), predicts survival within 5 years amongst newly diagnosed patients. This study validates its performance in the Brazilian Hodgkin lymphoma registry. By 2022, the Brazilian HL registry included 1357 cHL patients, with a median 5-year follow-up. Probabilities for 5-year progression-free survival (PFS) and overall survival (OS) were calculated using A-HIPI-model equations. Discrimination (Harrell C -statistic/Uno C -statistic) and calibration measures assessed external validation and calibration. Lab values beyond the allowed range were excluded, mirroring the initial A-HIPI analysis. A total of 694 advanced-stage cHL patients met the original inclusion criteria (age 18–65 years, Stage IIB-IV). Median age was 31 years; 46.3% were females. Stage distribution was IIB (33.1%), III (27.4%), IV (39.5%). Bulky disease in 32.6%. Five-year PFS and OS were 68.4% and 86.0%, respectively. Harrell C -statistics were 0.60 for PFS and 0.69 for OS, and Uno C -statistics were 0.63 for PFS and 0.72 for OS. Calibration plots demonstrated well-calibrated predictions with calibration slopes of 0.91 and 1.03 for 5-year OS and PFS, respectively. Despite differing patient, clinical characteristics, and socioeconomic factors, the baseline prediction tool performed well in the Brazilian cohort, demonstrating adequate discrimination and calibration. This supports its reliability in diverse settings.

Background: Sickle cell disease (SCD), characterized by vaso-occlusive crises, hemolysis, endothelial damage, and chronic inflammation, is the most common inherited monogenic disorder worldwide. The mortality rate among patients with SCD in Brazil remains high, despite improvements in treatment over the past decades. Allogeneic hematopoietic stem cell transplantation (HSCT) remains, to date, the only curative therapy available for both children and adults in Brazil. The best outcomes, with overall survival (OS) and event-free survival (EFS) rates of 90%-100%, are mainly achieved in children, using HLA-identical sibling donors and a myeloablative conditioning regimen. In adults and in patients with comorbidities, reduced intensity and non-myeloablative conditioning regimens have been used to minimize both early and late HSCT-related toxicity. This study aims to assess the clinical outcomes of HLA-identical sibling HSCT using a reduced toxicity myeloablative (RTM) conditioning regimen in children and adults with sickle cell disease. Methods: This is a single center, observational and retrospective analysis of 60 patients (30 children and 30 adults), who underwent HLA-identical sibling HSCT for SCD after a RTM conditioning regimen, between 2008 and 2025. The conditioning regimen consisted of fludarabine (120-150mg/m2, busulfan (9.6-12.8mg/kg) and ATG (4.5-10.0mg/kg). Methotrexate and cyclosporin were used as graft-versus-host disease (GVHD) prophylaxis. Data was collected from medical records. The primary endpoint was 2-year overall survival (OS). Secondary endpoints included 2-year EFS, defined as survival without death or graft failure, and graft-versus-host disease-free, relapse-free survival (GRFS), defined as survival without grade III-IV acute and chronic GVHD requiring immunosuppressive therapy, relapse, or death. Cumulative incidence (CI) of relapse and grade III-IV acute or chronic GVHD requiring immunosuppression were also analyzed, with death considered a competing event. Continuous variables were summarized as median (range), and categorical variables as percentages. OS, EFS, and GRFS probabilities were estimated using the Kaplan–Meier method and comparisons were performed using the log-rank test. Risk factors were studied using Cox regression models. Multivariate analyses were not performed due to the low number of events. The analysis was performed using R statistical software (version 4.4.1). P values were considered statistically significant. Results: The median age at HSCT was 15 years (range, 7-35 years); 53% were male; 80% had sickle cell anemia (HbSS); cerebrovascular disease was present in 58% of patients (77% in children) and 22% had a previous history of erythrocyte alloimmunization. Prior to HSCT, 36% of patients were receiving hydroxyurea and chronic transfusion therapy. Bone marrow was the progenitor stem cell source in 97% of the patients (the others received peripheral blood). The mean (DP) dose of total nucleated cells was 3.88 x 108/kg (±1.48); 60% of HSCT were isogroups in the ABO system and 65% of donors had sickle cell trait. Median (IIQ) follow-up of survivors was 73 (30-122) months. All patients engrafted, except one who died due to multiple organ failure resulting from complications associated with transfusion refractoriness (severe alloimmunization) at day+15, before the expected period of engraftment. At day+30 and 1 year post-HSCT, mixed chimera was observed in 84% and 79%, respectively. Two-year OS, EFS and GRFS were 93% (95% CI 86-100); 80% (95% CI 70-91); and 72% (95% CI 85-61), respectively, and were not different between children and adults. Patients with prior erythrocyte alloimmunization had worse OS than those without alloimmunization (p=0.01). Cumulative incidence of secondary graft failure, grade III-IV acute GVHD and chronic GVHD requiring immunosuppressive therapy at 2 years post-HSCT were 10%, 5% and 5%, respectively. Variables associated with inferior overall survival was erythrocyte alloimmunization, age of patient and donor at HSCT (hazard ratio [HR], 9.39, p=0.007; [HR], 1.10, p=0.045; [HR], 1.07, p=0.046, respectively). Conclusion: HSCT with RTM conditioning regimen resulted in similar outcomes in both adult and pediatric populations and is a viable and safe treatment option for SCD treatment in adults and children.