LATS1 controls CTCF chromatin occupancy and hormonal response of 3D-grown breast cancer cells.

The cancer epigenome has been studied in cells cultured in two-dimensional (2D) monolayers, but recent studies highlight the impact of the extracellular matrix and the three-dimensional (3D) environment on multiple cellular functions. Here, we report the physical, biochemical, and genomic differences between T47D breast cancer cells cultured in 2D and as 3D spheroids. Cells within 3D spheroids exhibit a rounder nucleus with less accessible, more compacted chromatin, as well as altered expression of ~2000 genes, the majority of which become repressed. Hi-C analysis reveals that cells in 3D are enriched for regions belonging to the B compartment, have decreased chromatin-bound CTCF and increased fusion of topologically associating domains (TADs). Upregulation of the Hippo pathway in 3D spheroids results in the activation of the LATS1 kinase, which promotes phosphorylation and displacement of CTCF from DNA, thereby likely causing the observed TAD fusions. 3D cells show higher chromatin binding of progesterone receptor (PR), leading to an increase in the number of hormone-regulated genes. This effect is in part mediated by LATS1 activation, which favors cytoplasmic retention of YAP and CTCF removal.

Germline assessment for alloHSCT candidates over 50 years: A 'Fast-Track' screening in myeloid neoplasms.

Patients aged 50 or above diagnosed with myeloid neoplasms (MNs) are typically not candidates for germline testing. However, approximately 8% carry pathogenic germline variants. Allogeneic haematopoietic stem cell transplantation (alloHSCT) remains an option for those aged over 50; neglecting germline testing could mask the risk for relative donor cell-derived MN. We propose a germline-augmented somatic panel (GASP), combining MN predisposition genes with a myeloid somatic panel for timely germline variant identification when initial testing is not indicated. Out of our 133 whole-exome-sequenced MN cases aged over 50 years, 9% had pathogenic/likely variants. GASP detected 92%, compared to 50% with somatic-only panel. Our study highlights the relevance of germline screening in MN, particularly for alloHSCT candidates without established germline-testing recommendations.

Glycolysis inhibition affects proliferation and cytotoxicity of Vγ9Vδ2 T cells expanded for adoptive cell therapy.

BACKGROUND AIMS: Vγ9Vδ2 T cells are under investigation as alternative effector cells for adoptive cell therapy (ACT) in cancer. Despite promising in vitro results, anti-tumor efficacies in early clinical studies have been lower than expected, which could be ascribed to the complex interplay of tumor and immune cell metabolism competing for the same nutrients in the tumor microenvironment. METHODS: To contribute to the scarce knowledge regarding gamma delta T-cell metabolism, we investigated the metabolic phenotype of 25-day-expanded Vγ9Vδ2 T cells and how it is intertwined with functionality. RESULTS: We found that Vγ9Vδ2 T cells displayed a quiescent metabolism, utilizing both glycolysis and oxidative phosphorylation (OXPHOS) for energy production, as measured in Seahorse assays. Upon T-cell receptor activation, both pathways were upregulated, and inhibition with metabolic inhibitors showed that Vγ9Vδ2 T cells were dependent on glycolysis and the pentose phosphate pathway for proliferation. The dependency on glucose for proliferation was confirmed in glucose-free conditions. Cytotoxicity against malignant melanoma was reduced by glycolysis inhibition but not OXPHOS inhibition. CONCLUSIONS: These findings lay the groundwork for further studies on manipulation of Vγ9Vδ2 T-cell metabolism for improved ACT outcome.

Immune reconstitution after single-unit umbilical cord blood transplantation using anti-thymoglobulin and myeloablative conditioning in adults with hematological malignancies.

This study aimed to investigate the kinetics of immune recovery following umbilical cord blood transplantation (UCBT) in adults who received a myeloablative conditioning (MAC) regimen and antithymocyte globulin (ATG). While the immune recovery kinetics has been extensively studied in pediatric UCBT recipients, limited data exist for adults. We conducted a comprehensive analysis of 221 consecutive adult patients who underwent UCBT with MAC and ATG at a single institution. Our objective was to evaluate the influence of patient, disease, and transplant factors, along with acute graft-versus-host disease (aGVHD), on immune reconstitution and overall survival. Our findings confirm a delayed recovery of T cells, while B and NK cell reconstitution exhibited rapid progress, with NK cell counts reaching normal levels within 3 months post-transplantation and B cells within 6 months. Within CD3+ T cells, CD8+ T cells also experienced a delayed recovery (12 months), but to a lesser extent compared to CD4+ T cells (18 months). Delayed immune recovery of T-cell subsets was associated with the development of aGVHD grade II-IV, older age, CMV negativity, and a female donor. Patients with lymphoproliferative diseases showed slower NK cell recovery. Our study demonstrates that adult patients undergoing MAC with ATG and receiving a single unit UCBT for hematologic malignancies experienced rapid reconstitution of NK and B cells. However, T cell recovery, particularly CD4+ T cells, was significantly delayed. To enhance T cell recovery, it may be crucial to consider UCB units with higher cellularity and optimize ATG doses in conditioning.

Design and Validation of a Custom Next-Generation Sequencing Panel in Pediatric Acute Lymphoblastic Leukemia.

The molecular landscape of acute lymphoblastic leukemia (ALL) is highly heterogeneous, and genetic lesions are clinically relevant for diagnosis, risk stratification, and treatment guidance. Next-generation sequencing (NGS) has become an essential tool for clinical laboratories, where disease-targeted panels are able to capture the most relevant alterations in a cost-effective and fast way. However, comprehensive ALL panels assessing all relevant alterations are scarce. Here, we design and validate an NGS panel including single-nucleotide variants (SNVs), insertion-deletions (indels), copy number variations (CNVs), fusions, and gene expression (ALLseq). ALLseq sequencing metrics were acceptable for clinical use and showed 100% sensitivity and specificity for virtually all types of alterations. The limit of detection was established at a 2% variant allele frequency for SNVs and indels, and at a 0.5 copy number ratio for CNVs. Overall, ALLseq is able to provide clinically relevant information to more than 83% of pediatric patients, making it an attractive tool for the molecular characterization of ALL in clinical settings.

The lipidomic profile of the tumoral periprostatic adipose tissue reveals alterations in tumor cell's metabolic crosstalk.

BACKGROUND: Periprostatic adipose tissue (PPAT) plays a role in prostate cancer (PCa) progression. PPAT lipidomic composition study may allow us to understand the tumor metabolic microenvironment and provide new stratification factors. METHODS: We used ultra-high-performance liquid chromatography-mass spectrometry-based non-targeted lipidomics to profile lipids in the PPAT of 40 patients with PCa (n = 20 with low-risk and n = 20 high-risk). Partial least squares-discriminant analysis (PLS-DA) and variable importance in projection (VIP) analysis were used to identify the most relevant features of PPAT between low- and high-risk PCa, and metabolite set enrichment analysis was used to detect disrupted metabolic pathways. Metabolic crosstalk between PPAT and PCa cell lines (PC-3 and LNCaP) was studied using ex vivo experiments. Lipid uptake and lipid accumulation were measured. Lipid metabolic-related genes (SREBP1, FASN, ACACA, LIPE, PPARG, CD36, PNPLA2, FABP4, CPT1A, FATP5, ADIPOQ), inflammatory markers (IL-6, IL-1B, TNFα), and tumor-related markers (ESRRA, MMP-9, TWIST1) were measured by RT-qPCR. RESULTS: Significant differences in the content of 67 lipid species were identified in PPAT samples between high- and low-risk PCa. PLS-DA and VIP analyses revealed a discriminating lipidomic panel between low- and high-risk PCa, suggesting the occurrence of disordered lipid metabolism in patients related to PCa aggressiveness. Functional analysis revealed that alterations in fatty acid biosynthesis, linoleic acid metabolism, and ß-oxidation of very long-chain fatty acids had the greatest impact in the PPAT lipidome. Gene analyses of PPAT samples demonstrated that the expression of genes associated with de novo fatty acid synthesis such as FASN and ACACA were significantly lower in PPAT from high-risk PCa than in low-risk counterparts. This was accompanied by the overexpression of inflammatory markers (IL-6, IL-1B, and TNFα). Co-culture of PPAT explants with PCa cell lines revealed a reduced gene expression of lipid metabolic-related genes (CD36, FASN, PPARG, and CPT1A), contrary to that observed in co-cultured PCa cell lines. This was followed by an increase in lipid uptake and lipid accumulation in PCa cells. Tumor-related genes were increased in co-cultured PCa cell lines. CONCLUSIONS: Disturbances in PPAT lipid metabolism of patients with high-risk PCa are associated with tumor cell metabolic changes.

A Virtual Community of Practice to Improve Primary Health Care Professionals' Attitudes Toward Patient Empowerment (e-MPODERA): A Cluster Randomized Trial.

PURPOSE: We aimed to evaluate the effectiveness of a virtual community of practice (vCoP) in improving primary health care professionals' (HCPs') attitudes toward empowering patients with chronic disease. METHODS: We conducted a cluster randomized controlled trial. Practices were units of randomization, and primary HCPs and patients were units of analysis. Sixty-three practices in Madrid, Catalonia, and the Canary Islands were randomly allocated to the intervention or control groups. Randominzation of practices was performed after HCP and patient recruitment. The patients and statistician were anonymized to group allocation; it was not possible to anonymize HCPs. The intervention was a 12-month multicomponent tailored vCoP built on the Web 2.0 concept and focused on skills toward patient empowerment. The primary outcome was Patient-Provider Orientation Scale (PPOS) score at baseline and at 12 months. The secondary outcome was the Patient Activation Measure (PAM) score. RESULTS: A total of 321 HCPs and 1,921 patients were assessed. The intervention had a positive effect on PPOS total score (0.14 points higher in the vCoP arm; 95% CI, 0.03-0.25; P = .011) and the PPOS Sharing subscale (0.3 points higher in the vCoP arm; 95% CI, 0.15-0.44; P < .001). No effect was found for the PPOS Caring subscale, and no significant differences were found for PAM scores. CONCLUSIONS: A vCoP led to a minor increase in the PPOS Sharing component and the total score but not in the Caring component. However, considerable uncertainty remains, given the observed attrition and other limitations of the study. Further research is needed on the effectiveness of the vCoP model and on how to improve HCP engagement.VISUAL ABSTRACT.

Flow-Cytometric Method for Viability Analysis of Mycoplasma gallisepticum and Other Cell-Culture-Contaminant Mollicutes.

Mycoplasma is the smallest self-replicating bacteria, figuring as common contaminant of eukaryotic cell cultures. Production inputs and operator's manipulation seem to be the main sources of such contamination. Many analytical approaches have been applied for mycoplasma detection in cell cultures and also in biological products. However, unless they were validated, only indicator cell culture and bacteriological culture are considered as compendial methods for quality control of biological products. Nano-flow cytometry has been pointed out as an alternative technique for addressing prokaryotic and eukaryotic cell viability being a substantial tool for reference material production. In this study, a viability-flow-cytometry assay was standardized for M. gallisepticum and then applied to other cell-culture-contaminant mycoplasmas. For this, M. galliseticum's growth rate was observed and different treatments were evaluated to establish low viability cultures (cell death-induced control). Distinct viability markers and their ideal concentrations (titration) were appraised. Ethanol treatment showed to be the best death-inducing control. CFDA and TOPRO markers revealed to be the best choice for detecting live and dead mycoplasma frequencies, respectively. The standardized methodology was applied to Mycoplasma arginini, M. hyorhinis, M. orale, Spiroplasma citri and Acholeplasma laidlawii. Significant statistical difference was observed in the percentage of viable cells in comparison to ethanol treatment for A. laidlawii in CFDA and in both markers for M. gallisepticum, M. hyorhinis and S. citri. In summary, we standardized a flow cytometry assay for assessing M. gallisepticum - and potentially other species - viability and ultimately applied for reference material production improving the quality control of biological products.

Troubleshooting Complex Vascular Cases in the Kidney Transplant Recipient: Vascular Anomalies, Challenging Vessel Diseases, and Procedural Disasters.

PURPOSE OF REVIEW: To update the most relevant literature regarding complex cases during kidney transplant setting that recipient presents by himself, especially during implantation surgery due to vascular diseases and/or urinary tract anomalies. RECENT FINDINGS: Increasing age of donors and recipients is leading to an increased complexity of kidney transplant implantation surgery. In addition, the high peripheral vascular disease prevalence worldwide increases difficulty of surgery and decreases long-term outcomes as well. Moreover, it also increases transplant morbidity and mortality, both overall and cardiovascular, and finally clearly decreases graft survival. However, dialysis alternative has even worse outcomes in terms of mortality, with a proportional risk of death 2.66 higher compared with transplanted patients. Aorto-iliac prosthesis and 3rd and 4th transplants in occupied iliac fossae do also represent a challenging situation with a clearly increased morbidity and mortality. In some of those particular conditions, orthotopic kidney transplant technique is an alternative with good functional and survival outcomes, but not exempt of complications. Kidney transplant in vascular complex recipients has worse outcomes compared with conventional non-risky population. It remains a challenging surgical and medical procedure with higher morbidity and mortality, and decreased graft survival. However, dialysis mortality is still even greater and a transplant attempt might be justified. Orthotopic kidney transplant technique might play a role in selected patients with aorto-iliac unworkable segments or even in patients with special urinary tract conditions.

Troubleshooting Complex Vascular Cases in the Kidney Graft: Multiple Vessels, Aneurysms, and Injuries During Harvesting Procedures.

PURPOSE OF THE REVIEW: To update the most relevant literature regarding complex vascular cases in kidney transplant setting involving the graft, especially during the harvesting procedure and back-table preparation from the subsequent implant. RECENT FINDINGS: Challenging situations affecting the kidney graft such as multiple vessels, renal artery aneurysms, kidney anatomical anomalies, or major injuries do not contraindicate the transplant, but require an exhaustive graft viability assessment and several bench surgery techniques. Graft vessel conditioning in the back-table might include simple anastomosis between them, enlarging with venous patch or reconstruction with donor or synthetic grafts. Compared with conventional transplant, literature reports longer warm ischemia time (40 vs 32 min) and slightly increased rates of delayed graft function (10.3% vs 8.2%) and vascular complications (10.8% vs 8.1%), but similar graft and patient survival. Kidney graft vascular complex cases require exhaustive assessment, meticulous harvesting, good surgical technique in the bench table, and proper surgery in the recipient. Despite its complexity, vascular complex kidney transplant offers comparable outcomes in the long term to conventional population when technically well performed, with slightly increased rates of vascular complications and delayed graft function.

Noninfectious Neurologic Complications after Allogeneic Hematopoietic Stem Cell Transplantation.

Although allogeneic hematopoietic stem cell transplantation (allo-HSCT) can be associated with neurologic complications, data on noninfectious etiologies are scanty. Therefore, we analyzed the incidence, clinical characteristics, risk factors, and influence on outcomes of noninfectious neurologic complications (NCs) in 971 consecutive patients with hematologic malignancies undergoing allo-HSCT at our center between January 2000 and December 2016. We evaluated NCs affecting the central nervous system (CNS) and peripheral nervous system (PNS). The median duration of follow-up of survivors was 71 months (range, 11 to 213 months). A total of 467 patients received a matched sibling donor (MSD) transplant, 381 received umbilical cord blood (UCB), 74 received a haploidentical transplant, and 49 received a matched unrelated donor (MUD) transplant. One hundred forty-nine (15.3%) NCs were documented at a median of 78 days after transplantation (range, 5 days before to 3722 days after). The cumulative incidence risk of developing NC was 7.5% (95% confidence interval, 6% to 8.2%) at day +90 and 13% at 5 years. The 5-year cumulative incidence of NCs was 10.8% after MSD allo-HSCT and 15.3% after alternative donor (UCB, MUD, haploidentical) allo-HSCT (P = .004). There were 101 (68%) CNS complications, including encephalopathy, n = 46 (31%); headache, n = 20 (13%); stroke, n = 15 (10%); seizures, n = 9 (6%), posterior reversible encephalopathy syndrome, n = 6 (4%), and myelopathy, n = 5 (3%). PNS complications (32%) included neuropathies, n = 25 (17%), and myopathies and neuromuscular junction disorders, n = 23 (17%), with 17% of the total PNS complications being immune-related. In multivariable analysis, donor type other than MSD, age ≥40 years, development of acute graft-versus-host disease (GVHD) grade II-IV (hazard ratio [HR], 3.3; P < .00001), and extensive chronic GVHD (HR, 3.2; P = .0002) were independently associated with increased risk of NCs. The 5-year overall survival (OS) was 21% in patients who developed NCs and 41% for those who did not (P < .0001). This difference in OS was observed in patients developing CNS NCs, but not in those developing PNS complications. In conclusion, our study reveals NCs as a frequent and heterogeneous complication that, when affecting CNS, is associated with poor prognosis following allo-HSCT.

Factors influencing platelet transfusion refractoriness in patients undergoing allogeneic hematopoietic stem cell transplantation.

Hematopoietic stem cell transplantation has been considered a risk factor for development of platelet transfusion refractoriness. The objective of this study was to assess the platelet transfusion refractoriness rate in patients undergoing allogeneic hematopoietic stem cell transplantation from different sources. We retrospectively reviewed the charts and transfusion records of patients who underwent allogeneic stem cell transplantation at our institution between 2013 and 2015. The evaluation of post-transfusion platelet count was assessed for each transfusion given, from day of progenitor infusion to day 30 after transplantation. Of 167 patients included in this study, 101 received peripheral blood stem cell transplantation (PBSCT) and 66 received umbilical cord blood transplantation (UCBT). Overall, the percentage of platelet transfusions with a 14-h CCI lower than 5000 was 59.3%, being these data significantly higher for UCBT (67.6%) than for PBSCT (31.0%). Seventy-eight percent of patients underwent UCBT become refractory, while 38.6% of patients who received PBSCT were refractory. Factors associated to platelet refractoriness were lower CD34+ cell dose infused, higher number of antibiotics used, presence of anti-HLA I antibodies, and reduced-intensity conditioning regimen. Platelet refractoriness is a frequent and complex adverse event and remains a therapeutic challenge in the management of patients undergoing HSCT. There is a higher rate of platelet refractoriness in patients who received UCBT as compared to patients who received PBSCT.

Anti-D Alloimmunization after RhD-Positive Platelet Transfusion in RhD-Negative Women under 55 Years Diagnosed with Acute Leukemia: Results of a Retrospective Study.

BACKGROUND: Anti-D alloimmunization can occur when platelets from RhD-positive donors are transfused to RhD-negative patients, due to red blood cell residues in the platelet concentrates. METHODS: Our objective was to analyze the anti-D alloimmunization rate in a selected group of women under 55 years of age diagnosed with acute leukemia over an 18-year period. We focused the analysis on RhD-negative patients who received RhD-positive platelet transfusions. RESULTS: From January 1998 to October 2016, 382 women under 55 years were diagnosed with acute leukemia. A total of 56 patients were RhD-negative, and 48 (85.7%) received RhD-positive platelets. The median number of platelet concentrates transfused per patient was 23, and 48% of all platelet transfusions were RhD-positive. The 48 RhD-negative patients received a total of 949 RhD-positive platelet concentrates. Two patients developed anti-D: a 36-year-old woman with M3 acute myeloblastic leukemia and a 52-year-old patient with a secondary acute myeloblastic leukemia. CONCLUSION: We conclude that there is a need for agreement in the transfusion guidelines on the recommendation of anti-D alloimmunization prophylaxis. We suggest a possible benefit in favor of anti-D prophylaxis in childbearing women with acute leukemia.

Severe feline sporotrichosis associated with an increased population of CD8low cells and a decrease in CD4⁺ cells.

Sporotrichosis is a subcutaneous mycosis with worldwide distribution, especially in tropical and subtropical areas. Zoonotic transmission is described with cats being the main animal species involved. The occurrence of severe feline sporotrichosis with high fungal levels demonstrates the susceptibility of cats to this disease and the importance of studying its pathogenesis. This study describes the leukocytes profile in blood of cats with sporotrichosis by flow cytometry and its correlation with histopathology and fungal load. The cats with sporotrichosis were separated into groups L1, L2, and L3 (lesions at one, two, and three or more noncontiguous skin locations, respectively) and were classified as good, fair, or poor general conditions. The highest percentage of CD4+ cells was associated to L1 (P = .04) and to good general condition (P = .03). The percentage of CD8+ cells was greater in L2 and L3 (P = .01). CD8(low) expression occurred in 20 animals with sporotrichosis, mainly in L3 (P = .01) and was not observed in healthy controls. This expression was related to macrophage granulomas (P = .01) and predominated in cases with high fungal load. Altogether, the results indicated that control over feline sporotrichosis, with maintenance of a good general condition, fixed lesions, well-organized response and lower fungal load, is associated with increased CD4+ cells percentages. In contrast, a poor general condition, disseminated lesions and high fungal load were related to increased CD8+ cell percentages and increased expression of CD8(low). As conclusion these results point to an important role of the CD4:CD8 balance in determining the clinical outcome in feline sporotrichosis.

Treatment of gastric marginal zone B-cell lymphoma of the mucosa-associated lymphoid tissue with rituximab, cyclophosphamide, vincristine and prednisone.

There is no standard treatment for patients with gastric marginal zone B-cell lymphoma of the mucosa-associated lymphoid tissue (MALT lymphoma) who are resistant to, or ineligible for, anti-Helicobacter pylori (anti-HP) therapy. In this study, we investigated the activity of the rituximab, cyclophosphamide, vincristine and prednisone (R-CVP) regimen in patients with gastric MALT lymphoma. Patients were included provided they had untreated gastric MALT lymphoma (except for anti-HP therapy) and were resistant to, or ineligible for, anti-HP therapy. Treatment plan consisted of six to eight 21-day cycles of the R-CVP chemotherapy regimen. Toxicity, response, relapse and survival were evaluated. Twenty patients (12 women and 8 men) were included in the analyses with median age of 59 years. Thirteen patients (65%) had stage I tumours, and seven patients (35%) had stages II-IV tumours. The overall response rate was 100%, with 19 (95%) complete responses and one (5%) partial response. Regimen toxicity was mild and mainly hematological, and no cases of gastric bleeding or perforation occurred. After a median follow-up of 56.3 months, three patients had relapsed, and 19 patients remained alive (specific lymphoma survival 100%), of whom 17 had no evidence of disease. In our experience, the R-CVP regimen is a well-tolerated and effective treatment for patients with gastric MALT lymphoma who are resistant to, or ineligible for, anti-HP therapy.

Therapeutic approaches in patients with newly diagnosed follicular lymphoma.

The prognosis of follicular lymphoma (FL) has significantly improved over the last decade, particularly following the introduction of the anti-CD20 monoclonal antibody rituximab, which has challenged the old concept of FL as an incurable disease. However, the decision whether to start treatment in a patient with advanced FL or adopt a watch-and-wait policy remains a subject of controversy. Furthermore, the optimal first-line treatment for FL remains a clinical challenge owing to the numerous different therapeutic options available. In this review, the authors focus on the initial management of patients with newly diagnosed FL, consider the different treatment options for every stage, paying special consideration to the therapeutic approaches for each clinical scenario, and discuss future directions.

Unraveling germline predisposition in hematological neoplasms: Navigating complexity in the genomic era.

Genomic advancements have yielded pivotal insights into hematological neoplasms, particularly concerning germline predisposition mutations. Following the WHO 2016 revisions, dedicated segments were proposed to address these aspects. Current WHO 2022, ICC 2022, and ELN 2022 classifications recognize their significance, introducing more mutations and prompting integration into clinical practice. Approximately 5-10% of hematological neoplasm patients show germline predisposition gene mutations, rising with risk factors such as personal cancer history and familial antecedents, even in older adults. Nevertheless, technical challenges persist. Optimal DNA samples are skin fibroblast-extracted, although not universally applicable. Alternatives such as hair follicle use are explored. Moreover, the scrutiny of germline genomics mandates judicious test selection to ensure precise and accurate interpretation. Given the significant influence of genetic counseling on patient care and post-assessment procedures, there arises a demand for dedicated centers offering specialized services.

Neutrophil Oxidative Burst Profile Is Related to a Satisfactory Response to Itraconazole and Clinical Cure in Feline Sporotrichosis.

Despite the central role of cats in the transmission and amplification of Sporothrix, studies regarding immune response in feline sporotrichosis are scarce. In cats with sporotrichosis, neutrophil-rich lesions are usually associated to good general condition and lower fungal burden. However, the role of neutrophils in anti-Sporothrix immunity has been little explored in cats. Thus, the aim of this study was to evaluate the neutrophil oxidative burst in the blood of cats with sporotrichosis. Cats with sporotrichosis included in the study were treated with itraconazole (ITZ) alone or combined with potassium iodide (KI). The neutrophil oxidative burst was evaluated through a flow-cytometry-based assay using dihydrorhodamine 123 (background) and stimulation with Zymosan and heat-killed Sporothrix yeasts. The cure rate was 50.0% in cats under treatment with ITZ monotherapy and 90.9% in cats treated with ITZ + KI (p = 0.014), endorsing the combination therapy as an excellent alternative for the treatment of feline sporotrichosis. Higher percentages of Sporothrix-stimulated neutrophils were associated with good general condition (p = 0.003). Higher percentages of Sporothrix- (p = 0.05) and Zymosan-activated (p = 0.014) neutrophils before and early in the treatment were related to clinical cure in ITZ-treated cats. The correlation between oxidative burst and successful use of KI could not be properly assessed given the low number of failures (n = 2) in this treatment group. Nasal mucosa involvement, typically linked to treatment failure, was related to lower percentages of activated neutrophils in the background at the treatment outcome (p = 0.02). Our results suggest a beneficial role of neutrophils in feline sporotrichosis and a positive correlation between neutrophil activation and the cure process in ITZ-treated cats.

Comprehensive detection of CRLF2 alterations in acute lymphoblastic leukemia: a rapid and accurate novel approach.

Introduction: Acute lymphoblastic leukemia (ALL) is a prevalent childhood cancer with high cure rate, but poses a significant medical challenge in adults and relapsed patients. Philadelphia-like acute lymphoblastic leukemia (Ph-like ALL) is a high-risk subtype, with approximately half of cases characterized by CRLF2 overexpression and frequent concomitant IKZF1 deletions. Methods: To address the need for efficient, rapid, and cost-effective detection of CRLF2 alterations, we developed a novel RT-qPCR technique combining SYBR Green and highresolution melting analysis on a single plate. Results: The method successfully identified CRLF2 expression, P2RY8::CRLF2 fusions, and CRLF2 and JAK2 variants, achieving a 100% sensitivity and specificity. Application of this method across 61 samples revealed that 24.59% exhibited CRLF2 overexpression, predominantly driven by IGH::CRLF2 (73.33%). High Resolution Melting analysis unveiled concurrent CRLF2 or JAK2 variants in 8.19% of samples, as well as a dynamic nature of CRLF2 alterations during disease progression. Discussion: Overall, this approach provides an accurate identification of CRLF2 alterations, enabling improved diagnostic and facilitating therapeutic decision-making.

Longitudinal outcome over four decades of allogeneic stem cell transplantation: a single center experience.

This 45-year study (1978-2022) at a single institution evaluated HSCT outcomes and complications, emphasizing recent advances, with to provide insights into HSCT's evolving field and ongoing efforts to enhance patient outcomes. Involving 1707 patients, the study revealed an initial phase (1978-1987) with a limited activity that yielded modest outcomes, a nearly three-decade span (1988-2016) with a substantial increase in transplant activity, emphasizing umbilical cord blood transplantation (UCBT) for patients lacking a suitable matched sibling donor. In addition to a gradual increase in recipient age, significant improvement in outcomes emerged in the recent period (2017-2022), marked by UCBT replacement with haploidentical transplants, introduction of PTCY-based GVHD prophylaxis for all type of transplants, and increased use of conditioning regimens with thiotepa, busulfan, and fludarabine. In this period, reductions in GVHD, non-relapse mortality, and relapse rates significantly contributed to improved overall survival, event-free survival, and GVHD-free/relapse-free survival. The study identified specific factors, including GVHD prophylaxis and donor selection changes, associated with these positive trends. This four-decade study provides a unique perspective on allogeneic HSCT, showcasing the dynamic evolution of transplantation practices and their impact on outcomes, offering valuable insights for personalized treatment approaches and emphasizing continual innovation in this critical therapeutic modality.