Real-life use of ropeg-interferon α2b in polycythemia vera: patient selection and clinical outcomes.

Ropeginterferon-alfa2b (ropegIFNα2b) is a long-acting IFN formulation with broad FDA/EMA approval as a therapy of polycythemia vera (PV) with no symptomatic splenomegaly. There is currently lack of information on the real-world patient selection, including the impact of local reimbursement policies, and drug management, particularly: type/timing of screening and follow-up tests; absolute/relative contraindications to therapy; ropegIFNα2b dose and combinations with hydroxyurea. As a sub-analysis of the PV-ARC retrospective study (NCT06134102), we here report our monocenter experience with ropegIFNα2b in the period from January 2021, corresponding to drug availability outside clinical trial, and December 2023. Among the 149 patients with EMA/FDA indication, only 55 (36.9%) met the local reimbursement criteria and 18 (12.1%) received ropegIFNα2b. Thanks to appropriate screening, relative/absolute contraindications to ropegIFNα2b were detected and managed in a multidisciplinary manner. Efficacy and safety of ropegIFNα2b was confirmed, with 3 cases of early molecular response. General use of low ropegIFNα2b dose, with frequent need for hydroxyurea combinations, was noted. This real-world experience suggests a significant impact of local regulations on drug prescription and the need for greater real-world data collection on ropegIFNα2b in PV patients. Also, it describes appropriate multidisciplinary screening and monitoring procedures during ropegIFNα2b therapy.

A venetoclax and azacitidine bridge-to-transplant strategy for NPM1-mutated acute myeloid leukaemia in molecular failure.

NPM1-mutated acute myeloid leukaemia (NPM1mut AML) represents a mostly favourable/intermediate risk disease that benefits from allogeneic haematopoietic stem cell transplantation (HSCT) in case of measurable residual disease (MRD) relapse or persistence after induction chemotherapy. Although the negative prognostic role of pre-HSCT MRD is established, no recommendations are available for the management of peri-transplant molecular failure (MF). Based on the efficacy data of venetoclax (VEN)-based treatment in NPM1mut AML older patients, we retrospectively analysed the off-label combination of VEN plus azacitidine (AZA) as bridge-to-transplant strategy in 11 NPM1mut MRD-positive fit AML patients. Patients were in MRD-positive complete remission (CRMRDpos ) at the time of treatment: nine in molecular relapse and two in molecular persistence. After a median number of two cycles (range 1-4) of VEN-AZA, 9/11 (81.8%) achieved CRMRD -negative (CRMRDneg ). All 11 patients proceeded to HSCT. With a median follow-up from treatment start of 26 months, and a median post-HSCT follow-up of 19 months, 10/11 patients are alive (1 died from non-relapse mortality), and 9/10 patients are in MRDneg status. This patient series highlights the efficacy and safety of VEN-AZA to prevent overt relapse, achieve deep responses and preserve patient fitness before HSCT, in patients with NPM1mut AML in MF.

Concurrent chromothripsis events in a case of TP53 depleted acute myeloid leukemia with myelodysplasia-related changes.

Acute myeloid leukemia with myelodysplasia-related changes (AML-MRC) is a heterogeneous hematological disorder defined by morphological, genetic, and clinical features. Patients with AML-MRC often show cytogenetic changes, which are associated with poor prognosis. Straightforward criteria for AML-MRC diagnosis and a more rigorous characterization of the genetic abnormalities accompanying this disease are needed. Here we describe an informative AML-MRC case, showing two separate, but concurrent, chromothripsis events, occurred at the onset of the tumor, and originating an unbalanced t(5;7) translocation and a derivative chromosome 12 with a highly rearranged short arm. Conversely, despite chromothripsis has been often associated with genomic amplification in cancer, in this case a large marker chromosome harboring amplified sequences from chromosomes 19 and 22 arose from a stepwise mechanism. Notably, the patient also showed a TP53 mutated status, known to be associated with an increased susceptibility towards chromothripsis and a poor prognosis. Our results indicate that multiple chromothripsis events may occur early in neoplastic transformation and act in a synergistic way with progressive chromosomal alterations to determine a dramatic impact on disease outcome, as suggested by the gene expression profile analysis.

Mutations and long-term outcome of 217 young patients with essential thrombocythemia or early primary myelofibrosis.

We investigated the influence of molecular status on disease characteristics and clinical outcome in young patients (⩽ 40 years) with World Health Organization (WHO)-defined essential thrombocythemia (ET) or early/prefibrotic primary myelofibrosis (early-PMF). Overall, 217 patients with ET (number 197) and early-PMF (number 20) were included in the analysis. Median follow-up time was 10.2 years. The cumulative incidence of thrombosis, hemorrhages and disease evolution into myelofibrosis/acute leukemia were 16.6%, 8.6% and 3% at 15 years, respectively. No differences were detectable between ET and early-PMF patients, although the latter cohort showed a trend for worse combined-event free survival (EFS). Mutation frequency were 61% for JAK2V617F, 25% for CALR and 1% for MPLW515K, and were comparable across WHO diagnosis; however, JAK2V617F allele burden was higher in the early-PMF group. Compared with JAK2V617F-positive patients, CALR-mutated patients displayed higher platelet count and lower hemoglobin level. CALR mutations significantly correlated with lower thrombotic risk (9.1% versus 21.7%, P = 0.04), longer survival (100% versus 96%, P = 0.05) and better combined-EFS (86% versus 71%, P = 0.02). However, non-type 1/type 2 CALR mutations ('minor' mutations) and abnormal karyotype were found to correlate with increased risk of disease evolution. At last contact, six patients had died; in five cases, the causes of death were related to the hematological disease and occurred at a median age of 64 years (range: 53-68 years). Twenty-eight patients (13%) were unmutated for JAK2, CALR and MPL: no event was registered in these 'triple-negative' patients.

Profiling of drug-metabolizing enzymes/transporters in CD33+ acute myeloid leukemia patients treated with Gemtuzumab-Ozogamicin and Fludarabine, Cytarabine and Idarubicin.

Genetic heterogeneity in drug-metabolizing enzyme/transporter (DMET) genes affects specific drug-related cancer phenotypes. To investigate the relationships between genetic variation and response to treatment in acute myeloid leukemia (AML), we genotyped 1931 variants on DMET genes in 94 CD33-positive AML patients enrolled in a phase III multicenter clinical trial combining Gemtuzumab-Ozogamicin (GO) with Fludarabine-Cytarabine-Idarubicin (FLAI) regimen, with the DMET Plus platform. Two ADH1A variants showed statistically significant differences (odds ratio (OR)=5.68, P=0.0006; OR=5.35, P=0.0009) in allele frequencies between patients in complete/partial remission and patients without response, two substitutions on CYP2E1 (OR=0.13, P=0.001; OR=0.09, P=0.003) and one on SLCO1B1 (OR=4.68, P=0.002) were found to differently influence liver toxicity, and two nucleotide changes on SULTB1 and SLC22A12 genes correlated with response to GO (OR=0.24, P=0.0009; OR=2.75, P=0.0029). Genetic variants were thus found for the first time to be potentially associated with differential response and toxicity in AML patients treated with a combination of GO-FLAI regimen.

Gut microbiota as a candidate for lifespan extension: an ecological/evolutionary perspective targeted on living organisms as metaorganisms.

An emerging central concept in evolutionary biology suggests that symbiosis is a universal characteristic of living organisms that can help in understanding complex traits and phenotypes. During evolution, an integrative circuitry fundamental for survival has been established between commensal gut microbiota and host. On the basis of recent knowledge in worms, flies, and humans, an important role of the gut microbiota in aging and longevity is emerging. The complex bacterial community that populates the gut and that represents an evolutionary adapted ecosystem correlated with nutrition appears to limit the accumulation of pathobionts and infections in all taxa, being able of affecting the efficiency of the host immune system and exerting systemic metabolic effects. There is an urgent need to disentangle the underpinning molecular mechanisms, which could shed light on the basic mechanisms of aging in an ecological perspective. Thus, it appears possible to extend healthy aging and lifespan by targeting the host as a metaorganism by manipulating the complex symbiotic ecosystem of gut microbiota, as well as other possible ecosystems of the body.

Inflammatory response in molluscs: cross-taxa and evolutionary considerations.

Inflammation represents the rapid and efficient elimination of damaged tissue and microbes and eventually the restoration of tissue functionality. Inflammatory response is one of the vital reactions to body injury, acting alongside the restoration of homeostasis, wound repair and immune response. In mammals, wound healing is a process that seeks to restore tissue integrity and function, and is characterized by a series of biological processes including inflammatory response. Here, we review pioneering experiments and recent observations in invertebrate models suggesting that in highly divergent and evolutionary distant taxa, such as molluscs, insects and vertebrates, the inflammatory response could be driven by a pool of molecules sharing common evolutionary origin.

Erucylphosphohomocholine, the first intravenously applicable alkylphosphocholine, is cytotoxic to acute myelogenous leukemia cells through JNK- and PP2A-dependent mechanisms.

Alkylphospholipids and alkylphosphocholines (APCs) are promising antitumor agents, which target the plasma membrane and affect multiple signal transduction networks. We investigated the therapeutic potential of erucylphosphohomocholine (ErPC3), the first intravenously applicable APC, in human acute myelogenous leukemia (AML) cells. ErPC3 was tested on AML cell lines, as well as AML primary cells. At short (6-12 h) incubation times, the drug blocked cells in G2/M phase of the cell cycle, whereas, at longer incubation times, it decreased survival and induced cell death by apoptosis. ErPC3 caused JNK 1/2 activation as well as ERK 1/2 dephosphorylation. Pharmacological inhibition of caspase-3 or a JNK 1/2 inhibitor peptide markedly reduced ErPC3 cytotoxicity. Protein phosphatase 2A downregulation by siRNA opposed ERK 1/2 dephosphorylation and blunted the cytotoxic effect of ErPC3. ErPC3 was cytotoxic to AML primary cells and reduced the clonogenic activity of CD34(+) leukemic cells. ErPC3 induced a significant apoptosis in the compartment (CD34(+) CD38(Low/Neg) CD123(+)) enriched in putative leukemia-initiating cells. This conclusion was supported by ErPC3 cytotoxicity on AML blasts showing high aldehyde dehydrogenase activity and on the side population of AML cell lines and blasts. These findings indicate that ErPC3 might be a promising therapeutic agent for the treatment of AML patients.

Different isoforms of the B-cell mutator activation-induced cytidine deaminase are aberrantly expressed in BCR-ABL1-positive acute lymphoblastic leukemia patients.

The main reason for the unfavorable clinical outcome of BCR-ABL1-positive acute lymphoblastic leukemia (ALL) is genetic instability. However, how normal B-cell precursors acquire the genetic changes that lead to transformation has not yet been completely defined. We investigated the expression of the activation-induced cytidine deaminase (AID) and its role in clinical outcome in 61 adult BCR-ABL1-positive ALL patients. AID expression was detected in 36 patients (59%); it correlated with the BCR-ABL1 transcript levels and disappeared after treatment with tyrosine kinase inhibitors. Different AID splice variants were identified: full-length isoform; AIDDeltaE4a, with a 30-bp deletion of exon 4; AIDDeltaE4, with the exon 4 deletion; AIDins3, with the retention of intron 3; AIDDeltaE3-E4 isoform without deaminase activity. AID-FL predominantly showed cytoplasmic localization, as did the AID-DeltaE4a and AID-DeltaE3E4 variants, whereas the C-terminal-truncated AID-DeltaE4 showed a slightly increased nuclear localization pattern. AID expression correlated with a higher number of copy number alterations identified in genome-wide analysis using a single-nucleotide polymorphism array. However, the expression of AID at diagnosis was not associated with a worse prognosis. In conclusion, BCR-ABL1-positive ALL cells aberrantly express different isoforms of AID that may act as mutators outside the immunoglobulin (Ig) gene loci in promoting genetic instability.

The molecular anatomy of the FIP1L1-PDGFRA fusion gene.

The FIP1L1-PDGFRA fusion gene is a recurrent molecular abnormality in patients with eosinophilia-associated myeloproliferative neoplasms. We characterized FIP1L1-PDGFRA junction sequences from 113 patients at the mRNA (n=113) and genomic DNA (n=85) levels. Transcript types could be assigned in 109 patients as type A (n=50, 46%) or B (n=47, 43%), which were created by cryptic acceptor splice sites in different introns of FIP1L1 (type A) or within PDGFRA exon 12 (type B). We also characterized a new transcript type C (n=12, 11%) in which both genomic breakpoints fell within coding sequences creating a hybrid exon without use of a cryptic acceptor splice site. The location of genomic breakpoints within PDGFRA and the availability of AG splice sites determine the transcript type and restrict the FIP1L1 exons used for the creation of the fusion. Stretches of overlapping sequences were identified at the genomic junction site, suggesting that the FIP1L1-PDGFRA fusion is created by illegitimate non-homologous end-joining. Statistical analyses provided evidence for clustering of breakpoints within FIP1L1 that may be related to DNA- or chromatin-related structural features. The variability in the anatomy of the FIP1L1-PDGFRA fusion has important implications for strategies to detect the fusion at diagnosis or for monitoring response to treatment.

Age-related toxic effects and recovery from okadaic acid treatment in Enchytraeus crypticus (Annelida: Oligochaeta).

The effects of 48 h okadaic acid (OA) treatment and the ability to recover the induced toxicological injuries were examined in Enchytraeus crypticus of different ages (25 days and 3 months). The results demonstrated that the older worms are more sensitive to the toxin and show less capacity to recover. After 48 h OA treatment, the structural organization of the chloragogenous tissue appeared modified and associated with an immune response involving a higher number of circulating coelomocytes immunoreactive to anti-IL-6 antibody. The toxin effects were more evident in 3-month-old animals compared to specimens aged 25 days. Regarding the morpho-functional recovery from the induced modifications, first signs of recuperation were observed in younger worms at 48 h, and recovery was almost complete within 1 week. In older animals, the morphology of the chloragogenous tissue had not been restored, while a reduction in coelomocyte number was found after 1 week.

Proapoptotic activity and chemosensitizing effect of the novel Akt inhibitor perifosine in acute myelogenous leukemia cells.

The serine/threonine kinase Akt, a downstream effector of phosphatidylinositol 3-kinase (PI3K), is known to play an important role in antiapoptotic signaling and has been implicated in the aggressiveness of a number of different human cancers including acute myelogenous leukemia (AML). We have investigated the therapeutic potential of the novel Akt inhibitor, perifosine, on human AML cells. Perifosine is a synthetic alkylphospholipid, a new class of antitumor agents, which target plasma membrane and inhibit signal transduction networks. Perifosine was tested on THP-1 and MV 4-11 cell lines, as well as primary leukemia cells. Perifosine treatment induced cell death by apoptosis in AML cell lines. Perifosine caused Akt and ERK 1/2 dephosphorylation as well as caspase activation. In THP-1 cells, the proapoptotic effect of perifosine was partly dependent on the Fas/FasL system and c-jun-N-kinase activation. In MV 4-11 cells, perifosine downregulated phosphorylated Akt, but not phosphorylated FLT3. Moreover, perifosine reduced the clonogenic activity of AML, but not normal, CD34(+) cells, and markedly increased blast cell sensitivity to etoposide. Our findings indicate that perifosine, either alone or in combination with existing drugs, might be a promising therapeutic agent for the treatment of those AML cases characterized by upregulation of the PI3K-Akt survival pathway.

WT1 transcript amount discriminates secondary or reactive eosinophilia from idiopathic hypereosinophilic syndrome or chronic eosinophilic leukemia.

Idiopathic hypereosinophilic syndromes (HES) comprise a spectrum of indolent to aggressive diseases characterized by persistent hypereosinophilia. Hypereosinophilia can result from the presence of a defect in the hematopoietic stem cell giving rise to eosinophilia, it can be present in many myeloproliferative disorders or alternatively it may be a reactive form, secondary to many clinical conditions. The hybrid gene FIP1L1-PDGRFalpha was identified in a subset of patients presenting with HES or chronic eosinophilic leukemia (CEL). In spite of this, the majority of HES patients do not present detectable molecular lesions and for many of them the diagnosis is based on exclusion criteria and sometimes it remains doubt. In this study we explored the possibility to distinguish between HES/CEL and reactive hypereosinophilia based on WT1 transcript amount. For this purpose, 312 patients with hypereosinophilia were characterized at the molecular and cytogenetic level and analyzed for WT1 expression at diagnosis and during follow-up. This study clearly demonstrates that WT1 quantitative assessment allows to discriminate between HES/CEL and reactive eosinophilia and represents a useful tool for disease monitoring especially in the patients lacking a marker of clonality.

A putative helical cytokine functioning in innate immune signalling in Drosophila melanogaster.

In invertebrates and vertebrates, innate immunity is considered the first line of defense mechanism against non-self material. In vertebrates, cytokines play a critical role in innate immune signalling. To date, however, the existence of genes encoding for invertebrate helical cytokines has been anticipated, but never demonstrated. Here, we report the first structural and functional evidence of a gene encoding for a putative helical cytokine in Drosophila melanogaster. Functional experiments demonstrate that its expression, as well as that of the antimicrobial factors defensin and cecropin A1, is significantly increased after immune stimulation. These observations suggest the involvement of helical cytokines in the innate immune response of invertebrates.

IL-6 immunoreactivity changes during aging in the polychaete Ophryotrocha labronica (Polychaeta: Dorvilleidae).

IL-6 is a pleiotropic pro-inflammatory cytokine thought to play a role in age physiology, even if its possible modulation by aging mechanisms has not been fully defined. In this paper, the morpho-functional modifications and IL-6 immunoreactivity during aging in a simple invertebrate model, Ophryotrocha labronica, are reported. The comparison between newly-hatched, juveniles, young adult and 3-month-old females showed significant differences in the nervous and genital systems. There is evidence of nerve cell loss and a decline in oocyte growth and maturation at the gonad level. Immunohistochemistry reveals a different distribution of IL-6-like molecules, and a decreased number of reactive nerve cells in the central nervous system of aged O. labronica associated to the induced morphological modifications.

Growth factors and chemokines: a comparative functional approach between invertebrates and vertebrates.

Growth factors and cytokines control and coordinate a broad spectrum of fundamental cellular functions, and are evolutionarily conserved both in vertebrates and invertebrates. In this review, we focus our attention on the functional phylogenetic aspects of growth factors/cytokines like the Transforming Growth Factor-beta (TGF-beta), the Connective Tissue Growth Factor (CTGF), and the Vascular Endothelial Growth Factor (VEGF). We will also delve into the activites of two chemokine families, interleukin (IL)-8 (or CXCL8) and CC chemokine ligand 2/monocyte chemoattractant protein-1 (CCL2). These molecules have been selected for their involvement in immune responses and wound healing processes, where they mediate and finely regulate various regeneration processes like angiogenesis or fibroplasia, not only in vertebrates, but also in invertebrates.

Cytokines and invertebrates: TGF-beta and PDGF.

The review outlines the presence and function of TGF-beta and PDGF family members in invertebrates. TGF-beta and PDGF play an important role in development, in immune and neuroendocrine responses and in the wound repair by activating the classical transduction pathways. Generally speaking, these cytokines appear very early in evolution and conserve their functions.

Inflamm-aging, cytokines and aging: state of the art, new hypotheses on the role of mitochondria and new perspectives from systems biology.

In this article we summarise present knowledge on the role of pro-inflammatory cytokines on chronic inflammation leading to organismal aging, a phenomenon we proposed to call "inflamm-aging". In particular, we review genetic data regarding polymorphisms of genes encoding for cytokines and proteins involved in natural immunity (such as Toll-like Receptors and Heat Shock Proteins) obtained from large population studies including young, old and very old people in good health status or affected by age-related diseases such as Alzheimer's Disease and Type II Diabetes. On the whole, despite some controversial results, the available data are in favour of the hypothesis that pro-inflammatory cytokines play an important role in aging and longevity. Further, we present a possible hypothesis to reconcile energetic dysfunction, including mitochondria, and inflamm-aging. New perspectives for future studies, including phylogenetic studies in animal models and in silico studies on mathematical and bioinformatic models inspired by the systems biology approach, are also proposed.