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Abstract We analyzed the neutrophil-to-lymphocyte ratio (NLR) in 1508 patients with PV and found that those with an NLR ≥ 5 were generally older, had a longer disease history, and had higher cardiovascular risk factors, more arterial thrombosis, and more aggressive blood counts, indicating a more proliferative disease. NLR was an accurate predictor of mortality, with patients with NLR ≥ 5 having significantly worse overall survival and more than twice the mortality rate compared to those with NLR

Abstract This viewpoint summarizes findings from analyses of large personal patient databases of myeloproliferative neoplasms (MPNs) to assess the impact of thrombosis on mortality, disease progression, and second cancers (SC). Despite advances, the current incidence of arterial and venous thrombosis remains a challenge. These events appear to signal a more aggressive disease course, as evidenced by their association with myelofibrosis progression and mortality using multistate models and time-dependent multivariable analysis. Inflammatory biomarkers, such as the neutrophil-to-lymphocyte ratio (NLR), are associated with the aggressiveness of polycythemia vera (PV) and essential thrombocythemia (ET), linking thrombosis to SC risk. This suggests a common inflammatory pathway likely influencing cardiovascular disease and cancer incidence. Notably, this is observed more frequently in younger patients, likely due to prolonged exposure to MPN and environmental inflammatory triggers. These data underscore the need for new studies to validate these associations, delineate the sequence of events, and identify therapeutic targets to mitigate thrombotic events and potentially improve overall patient outcomes in MPN.

Although allogeneic hematopoietic cell transplantation (HCT) represents a curative approach for many patients with hematological diseases, post-transplantation relapse occurs in 20-50% of cases, representing the primary cause of treatment failure and mortality. Alloreactive donor T cells are responsible for the graft versus leukemia (GvL) effect, which represents the key mechanism for the long-term curative effect of HCT. However, the downside is represented by graft versus host disease (GvHD), largely contributing to transplant-related mortality (TRM). Multiple factors play a role in regulating the delicate balance between GvL and GvHD, such as the optimization of the donor HLA and KIR match, the type of graft source, and the adaptive use of post-transplant cellular therapy. In addition to the standard donor lymphocyte infusion (DLI), several attempts were made to favor the GvL effect without increasing the GvHD risk. Selected DLI, NK DLI, activated DLI and more sophisticated genetically engineered cells can be employed. In this scenario, cytokine-induced killer (CIK) cells represent a suitable tool to boost GvL while minimizing GvHD. CIK cells are T lymphocytes activated in culture in the presence of monoclonal antibodies against CD3 (OKT3), interferon-gamma (IFN-g), and interleukin-2 (IL-2), characterized by the expression of markers typical of NK cells and T cells (CD3+, CD56+, with a prevalent CD8+ phenotype). CIK cells can mediate cytotoxicity through both MHC and non-MHC restricted recognition, which is the so‐called “dual‐functional capability” and display minimum alloreactivity. Allogeneic CIK cells showed a favorable rate of response, especially in the setting of minimal residual disease, with a rate of GvHD not exceeding 25%. Finally, the CIK cell platform can be adapted for chimeric antigen receptor (CAR) cell strategy, showing promising results in both preclinical and clinical settings. In this review, we describe the main immunological basis for the development of the GvL and the possible cellular therapy approaches used to boost it, with a particular focus on the use of CIK cells.

Abstract We report the long-term results of a randomized trial (GITMO, AML-R2), comparing 1:1 the combination of busulfan and cyclophosphamide (BuCy2, n = 125) and the combination of busulfan and fludarabine (BuFlu, n = 127) as conditioning regimen in acute myeloid leukemia patients (median age 51 years, range 40–65) undergoing allogeneic hematopoietic stem cell transplantation. With a median follow-up of 6 years, significantly better non-relapse mortality (NRM) was confirmed in BuFlu recipients, which is sustained up to 4 years after transplant (10% vs. 20%, p = 0.0388). This difference was higher in patients older than 51 years (11% in BuFlu vs. 27% in BuCy2, p = 0.0262). The cumulative incidence of relapse, which was the first cause of death in the entire study population, did not differ between the two randomized arms. Similarly, the leukemia-free survival (LFS) and overall survival (OS) were not different in the two cohorts, even when stratifying patients per median age. Graft-and relapse-free survival (GRFS) in BuFlu arm vs. the BuCy2 arm was 25% vs. 20% at 4 years and 20% vs. 17% at 10 years. Hence, the benefit gained by NRM reduction is not offsets by an increased relapse. Leukemia relapse remains a major concern, urging the development of new therapeutic approaches.

Leukodystrophies represent a large and complex group of inherited disorders affecting the white matter of the central nervous system. Adult-onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP) is a rare leukodystrophy which still needs the proper identification of diagnostic, prognostic, and monitoring biomarkers. The aim of this study was to determine the diagnostic and prognostic value of chitinases and neurofilament light chain as biomarkers for ALSP. A cross-sectional study was performed to analyze cerebrospinal fluid levels of chitinases (chitotriosidase and chitinase 3-like 2) and neurofilament light chain in five different groups: (i) normal health individuals; (ii) patients with definitive diagnosis of ALSP and genetic confirmation; (iii) asymptomatic patients with CSF1R variants; (iv) patients with other adult-onset leukodystrophies; and (v) patients with amyotrophic lateral sclerosis (external control group). Chitinase levels showed a statistical correlation with clinical assessment parameters in ALSP patients. Chitinase levels were also distinct between ALSP and the other leukodystrophies. Significant differences were noted in the levels of chitinases and neurofilament light chain comparing symptomatic (ALSP) and asymptomatic individuals with CSF1R variants. This study is the first to establish chitinases as a potential biomarker for ALSP and confirms neurofilament light chain as a good biomarker for primary microgliopathies.

Abstract There is an increased risk of GVHD and of non-relapse mortality (NRM) after allogeneic stem cell transplantations (alloSCT) when mismatched unrelated donors (MMUD) are used. In Europe, it is standard practice to use rabbit anti-thymocyte globulin (rATG) to reduce the high NRM and GVHD risks after MMUD alloSCT. As an alternative to rATG, post-transplantation Cyclophosphamide (PTCy) is in increasing clinical use. It is currently impossible to give general recommendations regarding preference for one method over another since comparative evidence from larger data sets is lacking. To improve the evidence base, we analyzed the outcome of rATG vs. PTCy prophylaxis in adult patients with hematologic malignancies undergoing first peripheral blood alloSCT from MMUD (9/10 antigen match) between Jan 2018 and June 2021 in the database of the European Society for Blood and Marrow Transplantation (EBMT). We performed multivariate analyses using the Cox proportional-hazards regression model. We included 2123 patients in the final analyses (PTCy, n = 583; rATG, n = 1540). p values and hazard ratios (HR) presented here are multivariate outcomes. Two years after alloSCT we found a lower NRM in the PTCy group of 18% vs. 24.9% in the rATG group; p = 0.028, HR 0.74. Overall survival in the PTCy cohort was higher with 65.7% vs. 55.7% in the rATG cohort; p

We performed a monocentric, retrospective real-world analysis (RWA) to evaluate the feasibility and the main results of an autologous stem cell transplantation (ASCT)-based program in a cohort of 90 newly diagnosed multiple myeloma patients aged > 65 years who were deemed eligible to such program on the basis of a simplified frailty score. Seventy-six patients received at least one ASCT and 14 patients received also the second. Median progression-free survival was 42 months and overall survival was 84 months. These results compare well with those of other RWAs, and pave the way to further improvements upon incorporation of an anti-CD38+ monoclonal antibody in the ASCT programs.

IntroductionUrinary incontinence (UI) consists of involuntary leakage of urine during the storage phase of urination.MethodsAn anonymous survey was given to Spanish and Italian veterinarians about canine UI treated cases, diagnosis, treatment, follow-up, and professional interest.Results and discussionMost veterinarians treated ≤3 cases/quarter, resulting in the percentage of incontinence males being lower than that of females (1-4% vs 0-24%). The percentage of spayed incontinent females was lower in Spain (0-24%) than in Italy (75-100%). Most diagnoses were based on a diagnostic algorithm (Spain: 88.7%; Italy: 65.3%); patient report and history, blood work, urinalysis and abdominal ultrasound. Urethral/bladder pressure measurement was unusual (Spain: 0.2%; Italy: 2.4%). In Spain, radiology with contrast medium and CT urography (26.3% and 34.4%, respectively) were more frequent than in Italy (11.6% and 22.7%, respectively). When suspecting urethral sphincter mechanism incompetence pharmacological trial (Spain: 93.2%; Italy: 78.9%). The first-choice medical treatment was Phenylpropanolamine, followed by Ephedrine and Deslorelin. When pharmacotherapy failed, the most frequent option was drug change, followed by increased drug dosage/frequency of administration, surgical therapy and colposuspension. A review was completed after the first week of treatment followed by periodic reviews. Most of the respondents participated in continuing education only if UI occurred in their everyday practice (Spain: 63.0%; Italy: 55.4%) and about 30% responders did it regardless of the number of UI cases treated (Spain: 30.5%; Italy: 37.4%).ConclusionSome recommendations in clinical practice were made. UI can be underestimated by owners; therefore, a complete history should be obtained by veterinarians. Veterinarians should carefully evaluate if spaying is advisable considering it could increase UI risk. A step-by-step approach is recommended and a specific diagnostic-therapeutic algorithm for UI in dogs is provided. Conservative approaches (regular exercise, weight loss in overweight dogs and observing an “incontinence diary” to identify abnormal patterns of urination) are advisable.

Abstract Background Few studies explored the effect of the combination of glucose sodium-cotransporter-2 inhibitors (SGLT-2i) and glucagon-like peptide-1 receptor agonists (GLP-1RA) on the incidence of cardiovascular events in patients with type 2 diabetes (T2D) and acute myocardial infarction (AMI). Methods We recruited patients with T2D and AMI undergoing percutaneous coronary intervention, treated with either SGLT-2i or GLP-1RA for at least 3 months before hospitalization. Subjects with HbA1c 50% was higher in the SGLT-2i/GLP-1RA group compared with both SGLT-2i and GLP-1RA groups. Interpretation The combination of SGLT-2i and GLP-1RA is associated with a reduced incidence of cardiovascular events in patients with T2D and AMI compared with either drug used alone, with a significant effect also on peri-infarcted myocardial rescue in patients without a second event. Trial registraition ClinicalTrials.gov ID: NCT06017544.

Abstract We applied a parametric Markov five-state model, on a well-characterized international cohort of 1,545 patients with polycythemia vera (PV; median age 61 years; females 51%), in order to examine the impact of incident thrombosis on the trajectory of death or disease progression. At a median follow-up of 6.9 years, 347 (23%) deaths, 50 (3%) blast phase (BP), and 138 (9%) fibrotic (post-PV MF) transformations were recorded. Incident thrombosis occurred at a rate of 2.62% pt/yr (arterial 1.59% and venous 1.05%). The probability of death, in the first 10 years, for 280 (18%) patients who developed thrombosis during follow-up was 40%, which was two-fold higher than that seen in the absence of thrombosis or any other transition state (20%; p

Abstract Indicine cattle, also referred to as zebu (Bos taurus indicus), play a central role in pastoral communities across a wide range of agro-ecosystems, from extremely hot semiarid regions to hot humid tropical regions. However, their adaptive genetic changes following their dispersal into East Asia from the Indian subcontinent have remained poorly documented. Here, we characterize their global genetic diversity using high-quality whole-genome sequencing data from 354 indicine cattle of 57 breeds/populations, including major indicine phylogeographic groups worldwide. We reveal their probable migration into East Asia was along a coastal route rather than inland routes and we detected introgression from other bovine species. Genomic regions carrying morphology-, immune-, and heat-tolerance-related genes underwent divergent selection according to Asian agro-ecologies. We identify distinct sets of loci that contain promising candidate variants for adaptation to hot semi-arid and hot humid tropical ecosystems. Our results indicate that the rapid and successful adaptation of East Asian indicine cattle to hot humid environments was promoted by localized introgression from banteng and/or gaur. Our findings provide insights into the history and environmental adaptation of indicine cattle.

Strategies to increase the anti-tumor efficacy of cytokine-induced killer cells (CIKs) include genetic modification with chimeric antigen receptors (CARs) or the addition of soluble T-cell engaging bispecific antibodies (BsAbs). Here, CIKs were modified using a transposon system integrating two distinct anti-CD19 CARs (CAR-MNZ and CAR-BG2) or combined with soluble CD3xCD19 BsAb blinatumomab (CIK + Blina). CAR-MNZ bearing the CD28-OX40-CD3ζ signaling modules, and CAR-BG2, designed on the Tisagenlecleucel CAR sequence (Kymriah®), carrying the 4-1BB and CD3ζ signaling elements, were employed. After transfection and CIK expansion, cells expressed CAR-CD19 to a similar extent (35.9% CAR-MNZ and 17.7% CAR-BG2). In vitro evaluations demonstrated robust proliferation and cytotoxicity (~50% cytotoxicity) of CARCIK-MNZ, CARCIK-BG2, and CIK + Blina against CD19+ target cells, suggesting similar efficacy. All effectors formed an increased number of synapses, activated NFAT and NFkB, and secreted IL-2 and IFN-ɣ upon encountering targets. CIK + Blina displayed strongest NFAT and IFN-ɣ induction, whereas CARCIK-BG2 demonstrated superior synapse formation. All the effectors have shown therapeutic activity in vivo against the CD19+ Daudi tumor model, with CARCIK cells showing a more durable response compared to CIK + Blina, likely due to the short half-life of Blina in this model.

Sensitivity and uncertainty analysis hold significant importance across a range of applications, spanning from industrial problems to climate change, financial risk assessment, as well as mathematical and computational models. These analyses involve identifying influential input parameters in models to comprehend their impact on the output. Sensitivity analysis can be performed locally, examining parameter effects at a fixed value, or globally, evaluating the model across a range of parameter values. The Sobol method stands as a robust approach for global sensitivity analysis, employing a Sobol sequence to create samples more uniformly within the input parameter space, thus enabling efficient exploration of model inputs. This paper aims to introduce a computational implementation in Scilab to generate the Sobol sequence for utilization in sensitivity analysis through the Sobol method. A test case was applied to generate Sobol sequences and discuss the obtained results.

Abstract Aberrant DNA methylation at CpG dinucleotides is a cancer hallmark that is associated with the emergence of resistance to anti cancer treatment, though molecular mechanisms and biological significance remain elusive. Genome scale methylation maps by currently used methods are based on chemical modification of DNA and are best suited for analyses of methylation at CpG rich regions (CpG islands). We report the first high coverage whole-genome map in cancer using the long read nanopore technology, which allows simultaneous DNA-sequence and -methylation analyses on native DNA. We analyzed clonal epigenomic/genomic evolution in Acute Myeloid Leukemias (AMLs) at diagnosis and relapse, after chemotherapy. Long read sequencing coupled to a novel computational method allowed definition of differential methylation at unprecedented resolution, and showed that the relapse methylome is characterized by hypermethylation at both CpG islands and sparse CpGs regions. Most differentially methylated genes, however, were not differentially expressed nor enriched for chemoresistance genes. A small fraction of under-expressed and hyper-methylated genes at sparse CpGs, in the gene body, was significantly enriched in transcription factors (TFs). Remarkably, these few TFs supported large gene-regulatory networks including 50% of all differentially expressed genes in the relapsed AMLs and highly-enriched in chemoresistance genes. Notably, hypermethylated regions at sparse CpGs were poorly conserved in the relapsed AMLs, under-represented at their genomic positions and showed higher methylation entropy, as compared to CpG islands. Analyses of available datasets confirmed TF binding to their target genes and conservation of the same gene-regulatory networks in large patient cohorts. Relapsed AMLs carried few patient specific structural variants and DNA mutations, apparently not involved in drug resistance. Thus, drug resistance in AMLs can be mainly ascribed to the selection of random epigenetic alterations at sparse CpGs of a few transcription factors, which then induce reprogramming of the relapsing phenotype, independently of clonal genomic evolution.

Although allogeneic hematopoietic cell transplantation (HCT) represents a curative approach for many patients with hematological diseases, post-transplantation relapse occurs in 20-50% of cases, representing the primary cause of treatment failure and mortality. Alloreactive donor T cells are responsible for the graft versus leukemia (GvL) effect, which represents the key mechanism for the long-term curative effect of HCT. However, the downside is represented by graft versus host disease (GvHD), largely contributing to transplant-related mortality (TRM). Multiple factors play a role in regulating the delicate balance between GvL and GvHD, such as the optimization of the donor HLA and KIR match, the type of graft source, and the adaptive use of post-transplant cellular therapy. In addition to the standard donor lymphocyte infusion (DLI), several attempts were made to favor the GvL effect without increasing the GvHD risk. Selected DLI, NK DLI, activated DLI and more sophisticated genetically engineered cells can be employed. In this scenario, cytokine-induced killer (CIK) cells represent a suitable tool to boost GvL while minimizing GvHD. CIK cells are T lymphocytes activated in culture in the presence of monoclonal antibodies against CD3 (OKT3), interferon-gamma (IFN-g), and interleukin-2 (IL-2), characterized by the expression of markers typical of NK cells and T cells (CD3+, CD56+, with a prevalent CD8+ phenotype). CIK cells can mediate cytotoxicity through both MHC and non-MHC restricted recognition, which is the so‐called "dual‐functional capability" and display minimum alloreactivity. Allogeneic CIK cells showed a favorable rate of response, especially in the setting of minimal residual disease, with a rate of GvHD not exceeding 25%. Finally, the CIK cell platform can be adapted for chimeric antigen receptor (CAR) cell strategy, showing promising results in both preclinical and clinical settings. In this review, we describe the main immunological basis for the development of the GvL and the possible cellular therapy approaches used to boost it, with a particular focus on the use of CIK cells. [ABSTRACT FROM AUTHOR]