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The development of Next-Generation Sequencing (NGS) has provided useful diagnostic, prognostic, and therapeutic strategies for individualized management of B-cell precursor acute lymphoblastic leukemia (BCP-ALL) patients. Consequently, NGS is rapidly being established in clinical practice. However, the technology's complexity, bioinformatics analysis, and the different available options difficult a broad consensus between different laboratories in its daily routine introduction. This collaborative study among Spanish centers was aimed to assess the feasibility, pros, and cons of our customized panel and other commercial alternatives of NGS-targeted approaches. The custom panel was tested in three different sequencing centers. We used the same samples to assess other commercial panels (OncomineTM Childhood Cancer Research Assay; Archer®FusionPlex® ALL, and Human Comprehensive Cancer Panel GeneRead Panel v2®). Overall, the panels showed a good performance in different centers and platforms, but each NGS approach presented some issues, as well as pros and cons. Moreover, a previous consensus on the analysis and reporting following international guidelines would be preferable to improve the concordance in results among centers. Our study shows the challenges posed by NGS methodology and the need to consider several aspects of the chosen NGS-targeted approach and reach a consensus before implementing it in daily practice. [ABSTRACT FROM AUTHOR]

Simple Summary: Dysregulation of noncoding RNAs has been described in numerous types of cancers and it has been associated with oncogenic or tumor suppressor activities. However, the signature of clinically relevant noncoding RNAs in pediatric B-cell acute lymphoblastic leukemia is still poorly understood. In a search for long non-coding RNAs that characterize pediatric B-cell acute lymphoblastic leukemia, we found that the long non-coding RNA AL133346.1 and a neighbouring protein-coding mRNA (CCN2) were significantly over-expressed in leukemia samples compared to healthy bone marrow. Survival analysis showed that patients with high CCN2 expression had a significantly better prognosis. These data suggest that AL133346.1/CCN2 could be useful for discriminating subtypes of leukemia and that CCN2 expression could predict the prognosis of pediatric patients with B-cell acute lymphoblastic leukemia. Pediatric acute B-cell lymphoblastic leukemia (B-ALL) constitutes a heterogeneous and aggressive neoplasia in which new targeted therapies are required. Long non-coding RNAs have recently emerged as promising disease-specific biomarkers for the clinic. Here, we identified pediatric B-ALL-specific lncRNAs and associated mRNAs by comparing the transcriptomic signatures of tumoral and non-tumoral samples. We identified 48 lncRNAs that were differentially expressed between pediatric B-ALL and healthy bone marrow samples. The most relevant lncRNA/mRNA pair was AL133346.1/CCN2 (previously known as RP11-69I8.3/CTGF), whose expression was positively correlated and increased in B-ALL samples. Their differential expression pattern and their strong correlation were validated in external B-ALL datasets (Therapeutically Applicable Research to Generate Effective Treatments, Cancer Cell Line Encyclopedia). Survival curve analysis demonstrated that patients with "high" expression levels of CCN2 had higher overall survival than those with "low" levels (p = 0.042), and this gene might be an independent prognostic biomarker in pediatric B-ALL. These findings provide one of the first detailed descriptions of lncRNA expression profiles in pediatric B-ALL and indicate that these potential biomarkers could help in the classification of leukemia subtypes and that CCN2 expression could predict the survival outcome of pediatric B-cell acute lymphoblastic leukemia patients. [ABSTRACT FROM AUTHOR]

To determine whether a lower imatinib dose could minimize toxicity while maintaining the molecular response (MR), imatinib dose was reduced to 300 mg daily in 43 patients with chronic myeloid leukemia (CML) in sustained deep molecular response to first-line imatinib 400 mg daily. At the time of dose reduction, median duration of the deep response was 4.1 (interquartile range (IQR) 2.2-5.9) years; molecular response was MR4, MR4.5, and MR5 of the international scale in 6, 28, and 9 patients, respectively. Toxicity grade was 1, 2, and 3 in 28, 8, and 1 patients, respectively; 6 patients underwent dose reduction without having side effects. With a median of 1.6 (IQR 0.7-3.2) years on imatinib 300 mg daily, only one patient lost the deep molecular response to MR3. At the last follow-up, response was MR3, MR4, MR4.5, and MR5 in 1, 3, 9, and 30 patients, respectively. Toxicity improvement was observed in 23 (62.2 %) of the 37 patients with side effects, decreasing to grade 0 in 20 of them. All but one anemic patients improved (p = 0.01), the median Hb increase in this subgroup of patients being 1 g/dL. In CML patients with sustained deep response to the standard imatinib dose, reducing to 300 mg daily significantly improves tolerability and preserves efficacy. [ABSTRACT FROM AUTHOR]

Failure of second-generation tyrosine kinase inhibitors (2GTKI) is a challenging situation in patients with chronic myeloid leukemia (CML). Asciminib, recently approved by the US Federal Drug Administration, has demonstrated in clinical trials a good efficacy and safety profile after failure of 2GTKI. However, no study has specifically addressed response rates to asciminib in ponatinib pretreated patients (PPT). Here, we present data on responses to asciminib from 52 patients in clinical practice, 20 of them (38%) with prior ponatinib exposure. We analyzed retrospectively responses and toxicities under asciminib and compared results between PPT and non-PPT patients.After a median follow-up of 30 months, 34 patients (65%) switched to asciminib due to intolerance and 18 (35%) due to resistance to prior TKIs. Forty-six patients (88%) had received at least 3 prior TKIs. Regarding responses, complete cytogenetic response was achieved or maintained in 74% and 53% for non-PPT and PPT patients, respectively. Deeper responses such as major molecular response and molecular response 4.5 were achieved in 65% and 19% in non-PPT versus 32% and 11% in PPT, respectively. Two patients (4%) harbored the T315I mutation, both PPT.In terms of toxicities, non-PPT displayed 22% grade 3-4 TEAE versus 20% in PPT. Four patients (20% of PPT) suffered from cross-intolerance with asciminib as they did under ponatinib.Our data supports asciminib as a promising alternative in resistant and intolerant non-PPT patients, as well as in intolerant PPT patients; the resistant PPT subset remains as a challenging group in need of further therapeutic options. [ABSTRACT FROM AUTHOR]

Simple Summary: The present work comes up after detecting stakeholders' need to test for manifold molecular biomarkers in each sample from an individual diagnosed with myeloid neoplasm or acute leukemia. The development of gene panels based on NGS technology is considered a potentially effective testing alternative with less human effort, compared to that required by other conventional techniques (PCR, FISH, conventional karyotype, etc.). The validation of this panel aims to propose a new solution for hospitals to face the challenges posed by the molecular study of this group of onco-hematological diseases. A suitable diagnostic classification of myeloid neoplasms and acute leukemias requires testing for a large number of molecular biomarkers. Next-generation sequencing is a technology able to integrate identification of the vast majority of them in a single test. This manuscript includes the design, analytical validation and clinical feasibility evaluation of a molecular diagnostic kit for onco-hematological diseases. It is based on sequencing of the coding regions of 76 genes (seeking single-nucleotide variants, small insertions or deletions and CNVs), as well as the search for fusions in 27 target genes. The kit has also been designed to detect large CNVs throughout the genome by including specific probes and employing a custom bioinformatics approach. The analytical and clinical feasibility validation of the Haematology OncoKitDx panel has been carried out from the sequencing of 170 patient samples from 6 hospitals (in addition to the use of commercial reference samples). The analytical validation showed sensitivity and specificity close to 100% for all the parameters evaluated, with a detection limit of 2% for SNVs and SVs, and 20% for CNVs. Clinically relevant mutations were detected in 94% of all patients. An analysis of the correlation between the genetic risk classification of AML (according to ELN 2017) established by the hospitals and that obtained by the Haematology OncoKitDx panel showed an almost perfect correlation (K = 0.94). Among the AML samples with a molecular diagnosis, established by the centers according to the WHO, the Haematology OncoKitDx analysis showed the same result in 97% of them. The panel was able to adequately differentiate between MPN subtypes and also detected alterations that modified the diagnosis (FIP1L1-PDGFRA). Likewise, the cytogenetic risk derived from the CNV plot generated by the NGS panel correlated substantially with the results of the conventional karyotype (K = 0.71) among MDS samples. In addition, the panel detected the main biomarkers of prognostic value among patients with ALL. This validated solution enables a reliable analysis of a large number of molecular biomarkers from a DNA sample in a single assay. [ABSTRACT FROM AUTHOR]

Background: The analysis of molecular haematopoietic chimerisms (HC) has become a well-established method to monitor the transplant evolution and to assess the risk of relapse after allogeneic stem cells transplantation (allo-STC). Different techniques and molecular markers are being used for chimerism surveillance after transplantation, including quantitative real-time PCR (qPCR) and the recently developed digital PCR (dPCR). This study aims to compare the sensitivity and accuracy of both methods to quantify HC and predict early relapse. Methodology: HC was evaluated using custom PCR systems for the specific detection of the Y-chromosome, null alleles and insertion-deletion polymorphisms. A total of 281 samples from 28 adult patients who underwent an allo-SCT were studied. Increasing mixed chimerism was detected prior to relapse in 100% of patients (18 relapses). Results: Compared with conventional qPCR amplification, dPCR predicted relapse with a median anticipation period of 63 days versus 45.5 days by qPCR. Overall, 56% of the relapses were predicted earlier with dPCR whereas 38% of the relapses where detected simultaneously using both techniques and only in 1 case, relapse was predicted earlier with qPCR. Conclusions: In conclusion, chimerism determination by dPCR constitutes a suitable technique for the follow-up of patients with haematological pathologies after allo-STC, showing greater sensitivity to predict an early relapse. [ABSTRACT FROM AUTHOR]

Programmed death 1 (PD-1) activation triggers an immune checkpoint resulting in inhibition of T cells that leads to peripheral tolerance. Some PD-1 polymorphisms have been described and associated with the development of autoimmune diseases or cancer predisposition, but there are few data concerning the relevance of such polymorphisms on the clinical outcome after allogeneic hematopoietic stem cell transplant (alloHSCT). We analyzed the distribution of the SNPs PD-1.1G/A (rs36084323) and PD-1.3G/A (rs11568821) genotypes of the donor in a cohort of 1485 alloHSCT from HLA-identical sibling donors. We found an increased risk of grades II to IV graft-versus-host disease (GvHD) in patients receiving grafts from donors homozygous for the G allele at the rs36084323 SNP (P = 0.033; hazard ratio [HR] 2.2; 95% confidence interval [CI] 1.1 to 4.8) and also from donors homozygous for the A allele at the rs11568821 position (P < 0.001; HR 4.5, 95%CI 2.0 to 10.1). In contrast, the PD-1 genotype of the donor did not show association with overall survival or relapse incidence. These results suggest that the PD-1 genotype of the donor plays an important role for the development of acute GvHD after alloHSCT from HLA-identical sibling donors. [ABSTRACT FROM AUTHOR]

Minor histocompatibility antigen (miHA) mismatches have been related to graft-versus-host disease (GVHD) after allogeneic stem cell transplantation, but this association remains controversial due to the lack of consistency in the results obtained by different groups. The CTLA-4 genotype of the donor has been reported to be relevant in the appearance of acute GVHD. We explored the effect of the donor's CTLA-4 genotype in the incidence of acute GVHD associated with HA-1, HA-8, or H-Y miHA mismatches in a large cohort of 1295 patients receiving an allogeneic transplant from an HLA-identical sibling donor. The incidence of acute GVHD was higher if the donor and recipient were mismatched for HA-1, HA-8, or H-Y, but only when the donor had the CTLA-4 rs231775 AA genotype (hazard ratio [HR], 2.18; 95% confidence interval [CI], 1.27 to 3.75; P  = .005; HR, 2.11, 95% CI, 1.06 to 4.18; P  = .033; and HR, 1.50; 95% CI, 1.05 to 2.15; P  = .025, respectively). In contrast, this increased risk of developing acute GVHD was not found when the donor presented the CTLA-4 rs231775 AG or GG genotypes. We conclude that the immune response to specific miHA mismatches is modulated by the CTLA-4 genotype of the donor. [ABSTRACT FROM AUTHOR]

**Introduction:** Primary myelofibrosis (MF) is a rare hematologic disease belonging to the group of Philadelphia-negative chronic myeloproliferative neoplasms. Identification of the Janus Kinase (JAK) gene mutations inaugurated a new era in the targeted therapy of myeloproliferative diseases. Ruxolitinib is the first JAK1/JAK2 inhibitor specifically approved for the treatment of disease-related splenomegaly or symptoms in adult patients with primary myelofibrosis. The objective of this study was to assess the cost-effectiveness of ruxolitinib vs best available therapy (BAT) in MF patients in Spain. **Methods:** A decision-tree and Markov model were adapted to the Spanish setting to assess the cost-effectiveness of ruxolitinib vs. BAT on a lifetime horizon (≤15 years) from the societal perspective, while healthcare system perspective was included in the one-way sensitivity analysis. The population was assumed to be similar to that of the COMFORT-II clinical trial (CT), which was also the source of treatment efficacy data. BAT composition was derived from the same CT and validated with Spanish experts. Utilities were derived from the COMFORT-I CT. Costs included treatment, management, hospitalizations, emergency and outpatient visits, as well as adverse events and end-of-life costs. Additionally, costs associated to productivity loss were taken into account. Resource use was validated with experts and costs were extracted from Spanish sources. A probabilistic sensitivity analysis was also performed to evaluate the consistency of the results under the uncertainty or variability of the input data. **Results:** Patients on ruxolitinib accumulated 6.1 life years gained (LYGs), resulting in 73% extra life-years compared to patients treated with BAT (3.5LYs gained). Ruxolitinib provided 4.4 quality-adjusted life years (QALYs), with a 99% improvement compared to BAT (2.2 QALYs). This analysis gave an incremental cost of €47 199 per LYG and an incremental cost of €55 616 per QALY gained from the societal perspective. **Conclusions:** Ruxolitinib would be cost-effective in Spain according to the end-of-life criteria defined by the NICE and commonly referred for Spain (cost-effectiveness threshold of €61 500/QALY), in line with results published for other European countries.

In chronic myeloid leukemia (CML) patients, 3-month BCR-ABL1 levels have consistently been correlated with further outcomes. Monitoring molecular responses in CML using the GeneXpert (Cepheid) platform has shown an optimal correlation with standardized RQ-PCR (IS) when measuring BCR-ABL1 levels lower than 10%, as it is not accurate for values over 10%. The aim of the present study was to determine the predictive molecular value at three months on different outcome variables using the Xpert BCR-ABL1 MonitorTM assay (Xpert BCR-ABL1). We monitored 125 newly diagnosed consecutive CML patients in the chronic phase (CML-CP) using an automated method: Xpert BCR-ABL1. Only 5% of patients did not achieve an optimal response at 3 months, and the 10% BCR-ABL1 cutoff defined by RQ-PCR (IS) methods was unable to identify significant differences in the probabilities of achieving a complete cytogenetic response (CCyR) (50% vs. 87%, p = 0.1) or a major molecular response (MMR) (60% vs. 80%, p = 0.29) by 12 months. In contrast, a cutoff of 1.5% more accurately identified differences in the probabilities of achieving CCyR (98% vs. 54%, p

In chronic myeloid leukemia (CML) patients, 3-month BCR-ABL1 levels have consistently been correlated with further outcomes. Monitoring molecular responses in CML using the GeneXpert (Cepheid) platform has shown an optimal correlation with standardized RQ-PCR (IS) when measuring BCR-ABL1 levels lower than 10%, as it is not accurate for values over 10%. The aim of the present study was to determine the predictive molecular value at three months on different outcome variables using the Xpert BCR-ABL1 MonitorTM assay (Xpert BCR-ABL1). We monitored 125 newly diagnosed consecutive CML patients in the chronic phase (CML-CP) using an automated method: Xpert BCR-ABL1. Only 5% of patients did not achieve an optimal response at 3 months, and the 10% BCR-ABL1 cutoff defined by RQ-PCR (IS) methods was unable to identify significant differences in the probabilities of achieving a complete cytogenetic response (CCyR) (50% vs. 87%, p = 0.1) or a major molecular response (MMR) (60% vs. 80%, p = 0.29) by 12 months. In contrast, a cutoff of 1.5% more accurately identified differences in the probabilities of achieving CCyR (98% vs. 54%, p<0.001) and MMR (88% vs. 56%, p<0.001) by 12 months, as well as probabilities of treatment changes (p = 0.005). Therefore, when using the Xpert BCR-ABL1 assay, a cutoff of 1.5% at 3 months could with high probability identify patients able to achieve an optimal response at 12 months. [ABSTRACT FROM AUTHOR]

The current consensus on the diagnosis, prognosis, and treatment of essential thrombocythemia (ET) is based on experts' recommendations. However, several aspects of the diagnosis of, prognosis of, and therapy for ET are still controversial. The Delphi method was employed with an expert panel of members of the Spanish Group of Ph-negative Myeloproliferative Neoplasms in order to identify the degree of agreement on the diagnosis, prognosis, and treatment of ET. Nine leading experts selected a total of 41 clinical hematologists with well-known expertise in ET. An electronic questionnaire was used to collect the questions rated in a four-step scale. The questions were grouped into four blocks: diagnosis, risk stratification, goals of therapy, and treatment strategy. After the first round consisting of 80 questions, a second round including 14 additional questions focused on the recommendations advocated by experts of the European LeukemiaNet in 2011 was analyzed. The median and mean values for the first and second rounds were calculated. A summary of the conclusions considered as the most representative of each block of questions is presented. The Delphi method is a powerful instrument to address the current approaches and controversies surrounding ET. [ABSTRACT FROM AUTHOR]

Allo-SCT has a strong curative potential for AML patients mainly due to a GVL effect. Unfortunately, GvL and GVHD are intimately linked. IFN regulatory factor-3 (IRF3), by modulating innate immune reactions, could impact on the incidence and intensity of GVL and GVHD. We analyzed two gene variants in IRF3 (rs7251 and rs2304205) on the clinical outcome of 249 AML patients submitted to HLA-identical sibling allo-SCT. Patients with a donor carrying the dominant GG gene variant in rs7251 had, as compared with GC and CC variants, a lower acute GVHD (aGVHD) III-IV incidence (4% vs 11% vs 27%; P=0.0078), a higher relapse incidence (49% vs 35% vs 26%; P=0.018), and lower TRM (7% vs 24% vs 18%; P=0.0065). In functional studies, the GG variant was associated with lower production of IFN-γ, decreased lymphocyte proliferation after antigen presentation by DCs, and lower cytotoxic response of mature natural killer cells. Patients carrying the AA dominant variant in rs2304205 had higher relapse incidence (50% vs 39% vs 18%, P=0.0068). The presence of both variants (GG in rs7251 and AA in rs2304205) in donors and patients resulted in a stronger clinical impact. [ABSTRACT FROM AUTHOR]

The single nucleotide polymorphism (SNP) rs16754 of the WT1 gene has been previously described as a possible prognostic marker in normal karyotype acute myeloid leukemia (AML) patients. Nevertheless, the findings in this field are not always reproducible in different series. One hundred and seventy-five adult de novo AML patients were screened with two different methods for the detection of SNP rs16754: high-resolution melting (HRM) and FRET hybridization probes. Direct sequencing was used to validate both techniques. The SNP was detected in 52 out of 175 patients (30 %), both by HRM and hybridization probes. Direct sequencing confirmed that every positive sample in the screening methods had a variation in the DNA sequence. Patients with the wild-type genotype ( WT1) for the SNP rs16754 were significantly younger than those with the heterozygous WT1 genotype. No other difference was observed for baseline characteristic or outcome between patients with or without the SNP. Both techniques are equally reliable and reproducible as screening methods for the detection of the SNP rs16754, allowing for the selection of those samples that will need to be sequenced. We were unable to confirm the suggested favorable outcome of SNP rs16754 in de novo AML. [ABSTRACT FROM AUTHOR]

Abstract: The aim of our study was to determine the potential mechanism(s) implicated in Imatinib resistance in patients with Ph+ ALL. Resistance of Ph+ ALL cells to Imatinib-induced apoptosis was associated with lack of inhibition of Akt phosphorylation. Addition of the PI3K inhibitor LY294002 to Imatinib significantly increased apoptosis of Ph+ ALL cells. Interestingly, expression of PTEN was reduced in Ph+ ALL cells which was due to PTEN promoter hypermethylation. Treatment of Ph+ ALL cells with 5-Aza-2′-deoxycytidine was associated with an increased expression of PTEN and an increase in cell apoptosis. These results suggest that Imatinib resistance in patients with ALL may be dependent at least in part to PTEN down-regulation due to the abnormal promoter hypermethylation and support the potential role of de-methylating agents for the treatment of patients with Ph+ ALL. [Copyright &y& Elsevier]

Abstract: In this study, we report the case of a Philadelphia (Ph) positive chronic myelogenous leukemia (CML) patient with the presence of p190 and p210 BCR-ABL1 mRNA fusion transcripts derived from e1a2 and b3a2 BCR-ABL1 genomic rearrangements, respectively. The presence of e1a2 BCR-ABL1 genomic rearrangement was seen in 2 different clones, one with the rearrangement and another one with the rearrangement and deletion of the BCR gene of the non-rearranged chromosome 22. After treatment with imatinib, the p210 transcript could not be detected, whereas p190 was still present 6 months after initiation of imatinib therapy and progression to blast phase. The absence of p210 transcript post treatment indicates that the clone with b3a2 responded to imatinib and that the observed resistance was associated to cells harboring the e1a2 genomic rearrangement. Despite resistance of this patient to imatinib, no evidence of mutations in the kinase domain of ABL1 was found. Loss of normal BCR in one cell clone may contribute to the resistance to imatinib due to the lack of BCR mediated inhibition of BCR-ABL1. [Copyright &y& Elsevier]

Se presentan los resultados en el niño, hasta el primer mes de nacidos, y los costos, de la evaluación tecnológica de tres modelos de Atención del Parto, con diferente nivel de complejidad, en 2.313 gestantes de bajo riesgo obstétrico y similar nivel socioeconómico, en Cali, Colombia. Son estos resultados, en conjunto con los de la madre, objeto de otra publicación, los que determinan la escogencia del mejor modelo. Se hace énfasis en que las conclusiones comprenden a modelos de atención en una población urbana y a muy corta distancia (5-10 minutos) del hospital de referencia.

ABSTRACT Objective: To determine the prevalence and factors associated with syphilis, human immunodeficiency virus (HIV), hepatitis B (HBV) and herpes type 2 (HSV2) among women in the prison of San Sebastian in Cochabamba (Bolivia). Material and methods: We carried out a cross-sectional study including a standardized questionnaire to assess socio-demographics characteristics and risk factors (sexual practices and exposure to blood); and serological tests for syphilis, HSV2, VIH, and HBV. We performed bivariate and multivariate analyses to test the associations between variables of interest and infections. Results: A total of 219 out of 220 prisoners (99.5%) participated in the study. For syphilis, 12.8% of participants had both reactive tests (RPR+/TPPA+). The prevalence of HSV2 and VIH was 62.6% and 1.4%, respectively. Anti-HBc, indicating a resolved or chronic HBV, was positive in 11.9% of participants and 0.5% had active HBV (HBsAg positive). A low level of education was associated with syphilis, HSV2 and HBV. Having occasional sexual partners was associated with syphilis and HSV2. Being over 36 years old and having more than 3 children were associated with HBV. The number of sexual partners, history of prostitution and rape, having sexual intercourses in prison and detention time were not associated with any of these infections. Discussion: The prevalence of syphilis, HIV, HSV2 and HBV was higher in this vulnerable female population than in the general population in Bolivia. Control measures in detention are needed to limit the spread of these infections both in prisons and in the community.

Patients with chronic myeloid leukaemia in chronic phase (CML-CP) who have a sustained deep molecular response (DMR) are eligible to discontinue treatment and attempt treatment-free remission (TFR). In the DASFREE study (ClinicalTrials.gov; NCT01850004), the 2-year TFR rate after dasatinib discontinuation was 46%; here we present the 5-year update. Patients with a stable DMR after ≥2 years of dasatinib therapy discontinued treatment and were followed for 5 years. At a minimum follow-up of 60 months, in 84 patients discontinuing dasatinib, the 5-year TFR rate was 44% (n = 37). No relapses occurred after month 39 and all evaluable patients who relapsed and restarted dasatinib (n = 46) regained a major molecular response in a median of 1.9 months. The most common adverse event during the off-treatment period was arthralgia (18%, 15/84); a total of 15 withdrawal events were reported in nine patients (11%). At the 5-year final follow-up, almost half of the patients who discontinued dasatinib after a sustained DMR maintained TFR. All evaluable patients who experienced a relapse quickly regained a DMR after restarting dasatinib, demonstrating that dasatinib discontinuation is a viable and potentially long-term option in patients with CML-CP. The safety profile is consistent with the previous report., (© 2023 The Authors. British Journal of Haematology published by British Society for Haematology and John Wiley & Sons Ltd.)

(1) Background: Despite the prognostic improvements achieved with tyrosine kinase inhibitors (TKIs) in chronic myeloid leukemia (CML), a minority of patients still fail TKIs. The recent introduction of asciminib may be a promising option in intolerant patients, as it is a first-in-class inhibitor with a more selective mechanism of action different from the ATP-competitive inhibition that occurs with TKIs. Therefore, our goal was to analyze toxicities shown with asciminib as well as to study cross-toxicity with previous TKIs. (2) Methods: An observational, multicenter, retrospective study was performed with data from 77 patients with CML with therapeutic failure to second-generation TKIs who received asciminib through a managed-access program (MAP) (3) Results: With a median follow-up of 13.7 months, 22 patients (28.5%) discontinued treatment: 32% (7/22) due to intolerance and 45% (10/22) due to resistance. Fifty-five percent of the patients reported adverse effects (AEs) with asciminib and eighteen percent grade 3-4. Most frequent AEs were: fatigue (18%), thrombocytopenia (17%), anemia (12%), and arthralgias (12%). None of the patients experienced cardiovascular events or occlusive arterial disease. Further, 26%, 25%, and 9% of patients required dose adjustment, temporary suspension, or definitive discontinuation of treatment, respectively. Toxicities under asciminib seemed lower than with prior TKIs for anemia, cardiovascular events, pleural/pericardial effusion, diarrhea, and edema. Cross-toxicity risk was statistically significant for thrombocytopenia, anemia, neutropenia, fatigue, vomiting, and pancreatitis. (4) Conclusion: Asciminib is a molecule with a good safety profile and with a low rate of AEs. However, despite its new mechanism of action, asciminib presents a risk of cross-toxicity with classical TKIs for some AEs.

Gemtuzumab ozogamicin (GO) is a conjugate of a monoclonal antibody and calicheamicin, which has been reapproved for the treatment of acute myeloid leukemia (AML). AML patients with the CD33 rs12459419 CC genotype might benefit from the addition of GO to intensive treatment in contrast to patients with CT/TT genotypes. Nevertheless, contradictory results have been reported. We sought to shed light on the prediction of GO response in AML patients with rs12459419 polymorphism who were treated with GO in the consolidation ( n = 70) or reinduction ( n = 20) phase. The frequency distribution of the rs12459419 polymorphism in the complete cohort of patients was 44.4% ( n = 40), 50% ( n = 45), and 5.6% ( n = 5) for CC, CT, and TT genotypes, respectively. Regarding the patients treated with GO for consolidation, we performed a Kaplan-Meier analysis of overall survival and relapse-free survival according to the rs12459419 polymorphism (CC vs. CT/TT patients) and genetic risk using the European Leukemia Net (ELN) 2010 risk score. We also carried out a Cox regression analysis for the prediction of overall survival, with age and ELN 2010 as covariates. We found no statistical significance in the univariate or multivariate analysis. Additionally, we performed a global Kaplan-Meier analysis for the patients treated with GO for reinduction and did not find significant differences; however, our cohort was too small to draw any conclusion from this analysis. The use of GO in consolidation treatment is included in the approval of the compound; however, evidence regarding its efficacy in this setting is lacking. Rs12459419 polymorphism could help in the selection of patients who might benefit from GO. Regrettably, in our cohort, the rs12459419 polymorphism does not seem to be an adequate tool for the selection of patients who might benefit from the addition of GO in consolidation cycles., Competing Interests: T.C.B. is a PhD candidate at Universidad Autónoma de Madrid (UAM). No other conflicts of interest were declared., (Copyright © 2022 Tamara Castaño-Bonilla et al.)

Standardized monitoring of BCR::ABL1 mRNA levels is essential for the management of chronic myeloid leukemia (CML) patients. From 2016 to 2021 the European Treatment and Outcome Study for CML (EUTOS) explored the use of secondary, lyophilized cell-based BCR::ABL1 reference panels traceable to the World Health Organization primary reference material to standardize and validate local laboratory tests. Panels were used to assign and validate conversion factors (CFs) to the International Scale and assess the ability of laboratories to assess deep molecular response (DMR). The study also explored aspects of internal quality control. The percentage of EUTOS reference laboratories (n = 50) with CFs validated as optimal or satisfactory increased from 67.5% to 97.6% and 36.4% to 91.7% for ABL1 and GUSB, respectively, during the study period and 98% of laboratories were able to detect MR 4.5 in most samples. Laboratories with unvalidated CFs had a higher coefficient of variation for BCR::ABL1 IS and some laboratories had a limit of blank greater than zero which could affect the accurate reporting of DMR. Our study indicates that secondary reference panels can be used effectively to obtain and validate CFs in a manner equivalent to sample exchange and can also be used to monitor additional aspects of quality assurance., (© 2022. The Author(s).)

Robust and applicable risk-stratifying genetic factors at diagnosis in pediatric T-cell acute lymphoblastic leukemia (T-ALL) are still lacking, and most protocols rely on measurable residual disease (MRD) assessment. In our study, we aimed to analyze the impact of NOTCH1, FBXW7, PTEN , and RAS mutations, the measurable residual disease (MRD) levels assessed by flow cytometry (FCM-MRD) and other reported risk factors in a Spanish cohort of pediatric T-ALL patients. We included 199 patients treated with SEHOP and PETHEMA consecutive protocols from 1998 to 2019. We observed a better outcome of patients included in the newest SEHOP-PETHEMA-2013 protocol compared to the previous SHOP-2005 cohort. FCM-MRD significantly predicted outcome in both protocols, but the impact at early and late time points differed between protocols. The impact of FCM-MRD at late time points was more evident in SEHOP-PETHEMA 2013, whereas in SHOP-2005 FCM-MRD was predictive of outcome at early time points. Genetics impact was different in SHOP-2005 and SEHOP-PETHEMA-2013 cohorts: NOTCH1 mutations impacted on overall survival only in the SEHOP-PETHEMA-2013 cohort, whereas homozygous deletions of CDKN2A / B had a significantly higher CIR in SHOP-2005 patients. We applied the clinical classification combining oncogenetics, WBC count and MRD levels at the end of induction as previously reported by the FRALLE group. Using this score, we identified different subgroups of patients with statistically different outcome in both Spanish cohorts. In SHOP-2005, the FRALLE classifier identified a subgroup of high-risk patients with poorer survival. In the newest protocol SEHOP-PETHEMA-2013, a very low-risk group of patients with excellent outcome and no relapses was detected, with borderline significance. Overall, FCM-MRD, WBC count and oncogenetics may refine the risk-stratification, helping to design tailored approaches for pediatric T-ALL patients., Competing Interests: MT reports travel and accommodation support from Novartis (outside the submitted work); travel and accommodation support from Jazz Pharma and Shire/Servier (outside the submitted work); and travel and accommodation support from Amgen (outside the submitted work). JLF is a consultant/advisory member for Amgen, Jazz Pharmaceuticals, and Novartis (outside the submitted work), receives honoraria for speaking at symposia from Amgen, Servier, Jazz Pharmaceuticals, and Pfizer (outside the submitted work) and support for attending symposia from Servier and Jazz Pharmaceuticals (outside the submitted work). AM discloses talk fees from Jazz Pharma and Shire/Servier outside the presented work and reports travel and accommodation support from Jazz Pharma, Shire/Servier, and Novartis (outside the submitted work). JD reports advisory board honorarium, speaker fees, and travel and accommodation support from Jazz Pharma and Shire/Servier; personal fees, advisory board honorarium, speaker fees, and travel and accommodation support from Novartis (outside the submitted work); advisory board honorarium, and travel and accommodation support from Amgen (outside the submitted work); advisory board honorarium, speaker fees, and travel and accommodation support from Celgene (outside the submitted work); advisory board honorarium, speaker fees, and travel and accommodation support from Sobi (outside the submitted work). SR reports advisory board honorarium, speaker fees, and travel and accommodation support from Jazz Pharma and Shire/Servier; personal fees, advisory board honorarium, speaker fees, and travel and accommodation support from Novartis (outside the submitted work); advisory board honorarium, and travel and accommodation support from Amgen (outside the submitted work); advisory board honorarium, speaker fees, and travel and accommodation support from Celgene (outside the submitted work). MC discloses talk fees from Shire/Servier outside the presented work. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Vega-García, Perez-Jaume, Esperanza-Cebollada, Vicente-Garcés, Torrebadell, Jiménez-Velasco, Ortega, Llop, Abad, Vagace, Minguela, Pratcorona, Sánchez-Garcia, García-Calderón, Gómez-Casares, Martín-Clavero, Escudero, Riñón Martinez-Gallo, Muñoz, Velasco, García-Morin, Català, Pascual, Velasco, Fernández, Lassaletta, Fuster, Badell, Molinos-Quintana, Molinés, Guerra-García, Pérez-Martínez, García-Abós, Robles Ortiz, Pisa, Adán, Díaz de Heredia, Dapena, Rives, Ramírez-Orellana, Camós and on behalf of the Biological Committee of the Group of Leukaemia of the Spanish Society of Paediatric Haematology Oncology (SEHOP Group).)

Despite considerable advances in our understanding of the pathophysiology of graft-versus-host disease (GVHD), its prediction remains unresolved and depends mainly on clinical data. The aim of this study is to build a predictive model based on clinical variables and cytokine gene polymorphism for predicting acute GVHD (aGVHD) and chronic GVHD (cGVHD) from the analysis of a large cohort of HLA-identical sibling donor allogeneic stem cell transplant (allo-SCT) patients. A total of 25 SNPs in 12 cytokine genes were evaluated in 509 patients. Data were analyzed using a linear regression model and the least absolute shrinkage and selection operator (LASSO). The statistical model was constructed by randomly selecting 85% of cases (training set), and the predictive ability was confirmed based on the remaining 15% of cases (test set). Models including clinical and genetic variables (CG-M) predicted severe aGVHD significantly better than models including only clinical variables (C-M) or only genetic variables (G-M). For grades 3-4 aGVHD, the correct classification rates (CCR1) were: 100% for CG-M, 88% for G-M, and 50% for C-M. On the other hand, CG-M and G-M predicted extensive cGVHD better than C-M (CCR1: 80% vs. 66.7%, respectively). A risk score was calculated based on LASSO multivariate analyses. It was able to correctly stratify patients who developed grades 3-4 aGVHD ( P < .001) and extensive cGVHD ( P < .001). The novel predictive models proposed here improve the prediction of severe GVHD after allo-SCT. This approach could facilitate personalized risk-adapted clinical management of patients undergoing allo-SCT., (© 2018 by The American Society of Hematology.)

Minor histocompatibility Ags (mHags) have been implicated in the pathogenesis of GVHD after allogeneic hematopoietic stem cell transplantation (HSCT). Uridine diphospho-glucuronosyltransferase 2B17 (UGT2B17) gene deletion may act as a mHag and its association with acute GVHD (aGVHD) has been described. We retrospectively studied the clinical impact of a UGT2B17 mismatch in a cohort of 1127 patients receiving a HSCT from an HLA-identical sibling donor. UGT2B17 mismatch was present in 69 cases (6.1%). Incidence of severe aGVHD was higher in the UGT2B17 mismatched pairs (22.7% vs 14.6%), but this difference was not statistically significant (P: 0.098). We did not detect differences in chronic GVHD, overall survival, relapse-free survival, transplant-related mortality or relapse. Nevertheless, when we analyzed only those patients receiving grafts from a male donor (616 cases), aGVHD was significantly higher in the UGT2B17 mismatched group (25.1% vs 12.8%; P: 0.005) and this association was confirmed by the multivariate analysis (P: 0.043; hazard ratio: 2.16, 95% confidence interval: 1.03-4.57). Overall survival was worse for patients mismatched for UGT2B17 (P: 0.005). We conclude that UGT2B17 mismatch has a negative clinical impact in allogeneic HSCT from HLA-identical sibling donors only when a male donor is used. These results should be confirmed by other studies.

The FOXP3 gene encodes for a protein (Foxp3) involved in the development and functional activity of regulatory T cells (CD4+/CD25+/Foxp3+), which exert regulatory and suppressive roles over the immune system. After allogeneic stem cell transplantation, regulatory T cells are known to mitigate graft versus host disease while probably maintaining a graft versus leukemia effect. Short alleles (≤(GT)15) for the (GT)n polymorphism in the promoter/enhancer of FOXP3 are associated with a higher expression of FOXP3, and hypothetically with an increase of regulatory T cell activity. This polymorphism has been related to the development of auto- or alloimmune conditions including type 1 diabetes or graft rejection in renal transplant recipients. However, its impact in the allo-transplant setting has not been analyzed. In the present study, which includes 252 myeloablative HLA-identical allo-transplants, multivariate analysis revealed a lower incidence of grade III-IV acute graft versus host disease (GVHD) in patients transplanted from donors harboring short alleles (OR = 0.26, CI 0.08-0.82, p = 0.021); without affecting chronic GVHD or graft versus leukemia effect, since cumulative incidence of relapse, event free survival and overall survival rates are similar in both groups of patients.

Background: Granulocyte colony-stimulating factor mobilized peripheral blood stem cells are increasingly used instead of bone marrow as a stem cell source for transplantation. Whereas this change is almost complete for autologous transplantation, there are some concerns when considering allogeneic transplants., Design and Methods: We performed a retrospective case-control study including 820 adult patients who had received an allogeneic stem cell transplant from an HLA-identical sibling donor. Quality of life (QoL) was assessed in 150 patients using the EORTC Quality of Life Questionnaire C30 (QLQ-C30)., Results: There were no statistically significant differences in overall survival at ten years (bone marrow: 48.9% vs. peripheral blood stem cells: 39.8%; p=0.621), transplant-related mortality (bone marrow: 28.9% vs. peripheral blood stem cells: 34.4%; p=0.682) or relapse incidence at 9 years (29.4% vs. 35.2%, respectively; p=0.688). Similar outcomes were maintained independently of the phase of the disease. However, multivariate analysis identified a higher incidence of acute graft-versus-host disease grades II-IV (p: 0.023; Hazard ratio [HR]: 1.41; 95% confidence interval [CI]: 1.05-1.89) and grades III-IV (p: 0.006; HR: 1.89; 95% CI: 1.20-2.98), in the peripheral blood stem cells-stem cell transplant group. As previously described, extensive chronic graft-versus-host disease was also more frequent in the peripheral blood stem cells group (28% vs. 15.6%; p<0.001). Patients transplanted with peripheral blood stem cells had significant impairment of role and social functioning., Conclusions: Although overall survival was not affected by the stem cell source, peripheral blood stem cell transplants were associated with a higher risk of both acute and chronic GvHD. Global quality of life was similar in both groups, but patients transplanted with peripheral blood stem cells showed worse role and social functioning scores, probably related to the increased incidence of chronic graft-versus-host disease.

The most frequent KIT mutations reported in core-binding factor acute myeloid leukemia are point mutations and insertions/deletions in exons 17 and 8. The vast majority of KIT mutation detection procedures are time-consuming, costly, or with a high lower limit of detection. High-resolution melting (HRM) is a gene scanning method that combines simplicity and rapid identification of genetic variants. We describe an HRM method for the simultaneous screening of exons 8 and 17 KIT mutations and report the results obtained in 69 core-binding factor acute myeloid leukemia patients. Mutation detection was compared with sequencing as the gold standard. The HRM method used high-resolution melting master reagents (Roche) and the LightCycler 480 (Roche) platform. HRM was reproducible, showed a lower limit of detection of 1%, and discriminated all patients with mutated KIT from controls without false positive or false negative results. Additionally, most of the mutations were differentiated from the other mutations. KIT mutations were present in 15.9% of patients, showing a higher incidence in inv(16) (25.8%) than in t(8;21) (7.9%). The presence of a KIT mutation was associated with a high white blood cell count, and adult patients with an exon 17 mutation had a higher incidence of relapse. These findings verify that HRM is a reliable, rapid, and sensitive method for KIT mutation screening. Furthermore, our study corroborates the unfavorable prognosis associated with exon 17 KIT mutations.

Background and Objectives: The role of hematopoietic chimerism after allogeneic stem cell transplantation (SCT) for acute leukemia remains controversial. We studied the relationship between hematopoietic chimerism and several prognostic variables on the outcome of SCT in patients with acute leukemia., Design and Methods: Chimerism was determined by a semiquantitative method, based on polymerase chain reaction (PCR) amplification of variable number tandem repeat (VNTR) minisatellites, in 133 consecutive patients who underwent allogeneic SCT for acute leukemia (68 myeloid, 58 lymphoid and 7 biphenotypic), all receiving a myeloablative conditioning regimen., Results: The median follow-up for the surviving patients was 44.8 months (range: 12.0-129.0). Recipient hematopoiesis (mixed chimerism, MC) was detected in 40 cases (30.1%). Two types of patients could be distinguished in this MC group: 29 with increasing MC and 9 with decreasing MC. The remaining 93 cases maintained complete donor chimerism (CC) over the whole follow-up period. Patients with increasing MC showed a significantly higher (p<0.001) rate of relapse (93.1%) and death (89.7%) in comparison to both those with CC (26.9% relapse, 44.1% dead) or decreasing MC (11.1% relapse, 44.4% dead). The detection of increasing MC preceded relapse by a median of 74 days (range: 5-434) and was significantly related with the absence of chronic graft-versus-host disease. Univariate and multivariate analysis confirmed that chimerism was the most significant variable involved in relapse, leukemia-free survival and overall survival after SCT., Interpretation and Conclusions: These results demonstrate that sequential determination of chimerism allows the prediction of relapse and death after SCT for acute leukemia. The interval between detection of increasing MC and relapse may permit timely implementation of therapeutic measures.