The Therapy and Management of Heart Failure with Preserved Ejection Fraction: New Insights on Treatment.

Heart failure with preserved ejection fraction (HFpEF) is a clinical syndrome characterised by the presence of diastolic dysfunction and elevated left ventricular filling pressure, in the setting of a left ventricular ejection fraction of at least 50%. Despite the epidemiological prevalence of HFpEF, a prompt diagnosis is challenging and many uncertainties exist. HFpEF is characterised by different phenotypes driven by various cardiac and non-cardiac comorbidities. This is probably the reason why several HFpEF clinical trials in the past did not reach strong outcomes to recommend a single therapy for this syndrome; however, this paradigm has recently changed, and the unmet clinical need for HFpEF treatment found a proper response as a result of a new class of drug, the sodium-glucose cotransporter 2 inhibitors, which beneficially act through the whole spectrum of left ventricular ejection fraction. The aim of this review was to focus on the therapeutic target of HFpEF, the role of new drugs and the potential role of new devices to manage the syndrome.

A rescue approach in refractory diffuse large B-cell lymphoma with obinutuzumab-redirected cytokine-induced killer cells: a first-in-human case report.

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Right Ventricular Dysfunction before and after Cardiac Surgery: Prognostic Implications.

Right ventricular dysfunction is a prognostic factor for morbidity and mortality across a broad spectrum of cardiovascular diseases. While the role of the right ventricle in surgical patients has emerged, the prognostic impact of right ventricular dysfunction remains unclear in a large cardiac surgery population. We reviewed the existing literature about the role of right ventricular dysfunction in adults undergoing different kinds of cardiac surgery either present before or developed after surgery itself. Pre- and post-operative right ventricular dysfunction has demonstrated substantial prognostic implications. However, there remains a lack of consensus regarding its definition and diagnostic criteria. The available literature is limited to small-sized studies, underscoring the need for studies with larger populations.

Prognostic Value of sST2 in Heart Failure.

In recent years, there has been growing interest in the risk stratification for heart failure, and the use of multiple biomarkers to identify different pathophysiological processes associated with this condition. One such biomarker is soluble suppression of tumorigenicity-2 (sST2), which has shown some potential for integration into clinical practice. sST2 is produced by both cardiac fibroblasts and cardiomyocytes in response to myocardial stress. Other sources of sST2 are endothelial cells of the aorta and coronary arteries and immune cells such as T cells. Indeed, ST2 is also associated with inflammatory and immune processes. We aimed at reviewing the prognostic value of sST2 in both chronic and acute heart failure. In this setting, we also provide a flowchart about its potential use in clinical practice.

Enalaprilat and losartan decrease erythroid precursors frequency in cells from patients with polycythemia vera.

OBJECTIVE: Polycythemia Vera (PV) is a myeloproliferative neoplasm characterized by the overproduction of red blood cells. First-line therapies are directed at lowering hematocrit levels. After the discovery of a mutation in the Janus kinase 2 (JAK2V617F), JAK2 inhibitors have been tested as second-line therapies. Despite these approaches, there is still the need for a major comprehension of the mechanisms involved in PV erythrocytosis and of more effective therapies. Angiotensin-converting enzyme (ACE) stimulates hematopoietic precursors proliferation and erythroid differentiation. We thus hypothesized that ACE inhibition could help in controlling erythrocytosis in PV. METHODS: We assessed the clonogenic potential by colony-forming unit (CFU) assay of mononuclear cells isolated from PV JAK2 positive or JAK2 negative patients with erythrocytosis treated with enalaprilat or losartan. RESULTS: Treatment with drugs led to a decrease of erythroid precursor frequency both in the presence and absence of JAK2 mutation, with a high extent in JAK2 positive cells and without affecting other types of precursors. No dose-dependent effect was observed. CONCLUSIONS: Our results demonstrate that ACE inhibition reduces erythroid precursor frequency, confirming the involvement of ACE in erythrocytosis despite the presence of JAK2 mutation and encouraging the hypothesis that ACE inhibitors and AT1R antagonists could help in directly managing erythrocytosis in PV.

Selective cell cycle arrest in glioblastoma cell lines by quantum molecular resonance alone or in combination with temozolomide.

BACKGROUND: Glioblastoma is the most aggressive form of brain cancer, characterised by high proliferation rates and cell invasiveness. Despite advances in surgery and radio-chemotherapy, patients continue to have poor prognoses, with a survival rate of 14-15 months. Thus, new therapeutic strategies are needed. Non-ionising electromagnetic fields represent an emerging option given the potential advantages of safety, low toxicity and the possibility to be combined with other therapies. METHODS: Here, the anticancer activity of quantum molecular resonance (QMR) was investigated. For this purpose, three glioblastoma cell lines were tested, and the QMR effect was evaluated on cancer cell proliferation rate and aggressiveness. To clarify the QMR mechanism of action, the proteomic asset after stimulation was delineated. Mesenchymal stromal cells and astrocytes were used as healthy controls. RESULTS: QMR affected cancer cell proliferation, inducing a significant arrest of cell cycle progression and reducing cancer tumorigenicity. These parameters were not altered in healthy control cells. Proteomic analysis suggested that QMR acts not only on DNA replication but also on the machinery involved in the mitotic spindle assembly and chromosome segregation. Moreover, in a combined therapy assessment, QMR significantly enhanced temozolomide efficacy. CONCLUSIONS: QMR technology appears to be a promising tool for glioblastoma treatment.

Logistics of an advanced therapy medicinal product during COVID-19 pandemic in Italy: successful delivery of mesenchymal stromal cells in dry ice.

BACKGROUND: During the coronavirus disease-2019 (COVID-19) pandemic, Italian hospitals faced the most daunting challenges of their recent history, and only essential therapeutic interventions were feasible. From March to April 2020, the Laboratory of Advanced Cellular Therapies (Vicenza, Italy) received requests to treat a patient with severe COVID-19 and a patient with acute graft-versus-host disease with umbilical cord-derived mesenchymal stromal cells (UC-MSCs). Access to clinics was restricted due to the risk of contagion. Transport of UC-MSCs in liquid nitrogen was unmanageable, leaving shipment in dry ice as the only option. METHODS: We assessed effects of the transition from liquid nitrogen to dry ice on cell viability; apoptosis; phenotype; proliferation; immunomodulation; and clonogenesis; and validated dry ice-based transport of UC-MSCs to clinics. RESULTS: Our results showed no differences in cell functionality related to the two storage conditions, and demonstrated the preservation of immunomodulatory and clonogenic potentials in dry ice. UC-MSCs were successfully delivered to points-of-care, enabling favourable clinical outcomes. CONCLUSIONS: This experience underscores the flexibility of a public cell factory in its adaptation of the logistics of an advanced therapy medicinal product during a public health crisis. Alternative supply chains should be evaluated for other cell products to guarantee delivery during catastrophes.

Regulation of DNA repair pathway choice in S and G2 phases by the NHEJ inhibitor CYREN.

Classical non-homologous end joining (cNHEJ) and homologous recombination compete for the repair of double-stranded DNA breaks during the cell cycle. Homologous recombination is inhibited during the G1 phase of the cell cycle, but both pathways are active in the S and G2 phases. However, it is unclear why cNHEJ does not always outcompete homologous recombination during the S and G2 phases. Here we show that CYREN (cell cycle regulator of NHEJ) is a cell-cycle-specific inhibitor of cNHEJ. Suppression of CYREN allows cNHEJ to occur at telomeres and intrachromosomal breaks during the S and G2 phases, and cells lacking CYREN accumulate chromosomal aberrations upon damage induction, specifically outside the G1 phase. CYREN acts by binding to the Ku70/80 heterodimer and preferentially inhibits cNHEJ at breaks with overhangs by protecting them. We therefore propose that CYREN is a direct cell-cycle-dependent inhibitor of cNHEJ that promotes error-free repair by homologous recombination during cell cycle phases when sister chromatids are present.

A BARF1-specific mAb as a new immunotherapeutic tool for the management of EBV-related tumors.

The use of monoclonal antibodies (mAb) for the diagnosis and treatment of malignancies is acquiring an increasing clinical importance, thanks to their specificity, efficacy and relative easiness of use. However, in the context of Epstein-Barr virus (EBV)-related malignancies, only cancers of B-cell origin can benefit from therapeutic mAb targeting specific B-cell lineage antigens. To overcome this limitation, we generated a new mAb specific for BARF1, an EBV-encoded protein with transforming and immune-modulating properties. BARF1 is expressed as a latent protein in nasopharyngeal (NPC) and gastric carcinoma (GC), and also in neoplastic B cells mainly upon lytic cycle induction, thus representing a potential target for all EBV-related malignancies. Considering that BARF1 is largely but not exclusively secreted, the BARF1 mAb was selected on the basis of its ability to bind a domain of the protein retained at the cell surface of tumor cells. In vitro, the newly generated mAb recognized the target molecule in its native conformation, and was highly effective in mediating both ADCC and CDC against BARF1-positive tumor cells. In vivo, biodistribution analysis in mice engrafted with BARF1-positive and -negative tumor cells confirmed its high specificity for the target. More importantly, the mAb disclosed a relevant antitumor potential in preclinical models of NPC and lymphoma, as evaluated in terms of both reduction of tumor masses and long-term survival. Taken together, these data not only confirm BARF1 as a promising target for immunotherapeutic interventions, but also pave the way for a successful translation of this new mAb to the clinical use.

Fighting Viral Infections and Virus-Driven Tumors with Cytotoxic CD4+ T Cells.

CD4+ T cells have been and are still largely regarded as the orchestrators of immune responses, being able to differentiate into distinct T helper cell populations based on differentiation signals, transcription factor expression, cytokine secretion, and specific functions. Nonetheless, a growing body of evidence indicates that CD4+ T cells can also exert a direct effector activity, which depends on intrinsic cytotoxic properties acquired and carried out along with the evolution of several pathogenic infections. The relevant role of CD4+ T cell lytic features in the control of such infectious conditions also leads to their exploitation as a new immunotherapeutic approach. This review aims at summarizing currently available data about functional and therapeutic relevance of cytotoxic CD4+ T cells in the context of viral infections and virus-driven tumors.

Role of VHL, HIF1A and SDH on the expression of miR-210: Implications for tumoral pseudo-hypoxic fate.

The hypoxia-inducible factor 1α (HIF-1α) and its microRNA target, miR-210, are candidate tumor-drivers of metabolic reprogramming in cancer. Neuroendocrine neoplasms such as paragangliomas (PGLs) are particularly appealing for understanding the cancer metabolic adjustments because of their associations with deregulations of metabolic enzymes, such as succinate dehydrogenase (SDH), and the von Hippel Lindau (VHL) gene involved in HIF-1α stabilization. However, the role of miR-210 in the pathogenesis of SDH-related tumors remains an unmet challenge. Herein is described an in vivo genetic analysis of the role of VHL, HIF1A and SDH on miR-210 by using knockout murine models, siRNA gene silencing, and analyses of human tumors. HIF-1α knockout abolished hypoxia-induced miR-210 expression in vivo but did not alter its constitutive expression in paraganglia. Normoxic miR-210 levels substantially increased by complete, but not partial, VHL silencing in paraganglia of knockout VHL-mice and by over-expression of p76del-mutated pVHL. Similarly, VHL-mutated PGLs, not those with decreased VHL-gene/mRNA dosage, over-expressed miR-210 and accumulate HIF-1α in most tumor cells. Ablation of SDH activity in SDHD-null cell lines or reduction of the SDHD or SDHB protein levels elicited by siRNA-induced gene silencing did not induce miR-210 whereas the presence of SDH mutations in PGLs and tumor-derived cell lines was associated with mild increase of miR-210 and the presence of a heterogeneous, HIF-1α-positive and HIF-1α-negative, tumor cell population. Thus, activation of HIF-1α is likely an early event in VHL-defective PGLs directly linked to VHL mutations, but it is a late event favored but not directly triggered by SDHx mutations. This combined analysis provides insights into the mechanisms of HIF-1α/miR-210 regulation in normal and tumor tissues potentially useful for understanding the pathogenesis of cancer and other diseases sharing similar underpinnings.

A site-selective hyaluronan-interferonα2a conjugate for the treatment of ovarian cancer.

While interferon alpha (IFNα) is used in several viral and cancer contexts, its efficacy against ovarian cancer (OC) is far from being incontrovertibly demonstrated and, more importantly, is hindered by heavy systemic side effects. To overcome these issues, here we propose a strategy that allows a targeted delivery of the cytokine, by conjugating IFNα2a with an aldehyde-modified form of hyaluronic acid (HA). The resulting HA-IFNα2a bioconjugate was biochemically and biologically characterized. The conjugation with HA did not substantially modified both the antiviral function and the anti-proliferative activity of the cytokine. Moreover, the induction of STAT1 phosphorylation and of a specific gene expression signature in different targets was retained. In vivo optical imaging biodistribution showed that the i.p.-injected HA-IFNα2a persisted into the peritoneal cavity longer than IFNα2a without being toxic for intraperitoneal organs, thus potentially enhancing the loco-regional therapeutic effect. Indeed, in OC xenograft mouse models bioconjugate significantly improved survival as compared to the free cytokine. Overall, HA-IFNα2a bioconjugate disclosed an improved anticancer efficacy, and can be envisaged as a promising loco-regional treatment for OC.

Reverse immunoediting: When immunity is edited by antigen.

Immune selective pressure occurring during cancer immunoediting shapes tumor features revealed at clinical presentation. However, in the "Escape" phase, the tumor itself has the chance to influence the immunological response. Therefore, the capacity of the immune response to sculpt the tumor characteristics is only one side of the coin and even the opposite is likely true, i.e. that an antigen can shape the immune response in a sort of "reverse immunoediting". This reciprocal modeling probably occurs continuously, whenever the immune system encounters a tumor/foreign antigen, and can be operative in the pathogen/immune system interplay, thus possibly permeating the protective immunity as a whole. In line with this view, the characterization of a T cell response as well as the design of both active and passive immunotherapy strategies should also take into account all Ag features (type, load and presentation). Overall, we suggest that the "reverse immunoediting" hypothesis could help to dissect the complex interplay between antigens and the immune repertoire, and to improve the outcome of immunotherapeutic approaches, where T cell responses are manipulated and reprogrammed.

Drug conjugation to hyaluronan widens therapeutic indications for ovarian cancer.

Management of ovarian cancer still requires improvements in therapeutic options. A drug delivery strategy was tested that allows specific targeting of tumor cells in combination with a controlled release of a cytotoxic molecule. To this aim, the efficacy of a loco-regional intraperitoneal treatment with a bioconjugate (ONCOFID-S) derived by chemical linking of SN-38, the active metabolite of irinotecan (CPT-11), to hyaluronan was assessed in a mouse model of ovarian carcinomatosis. In vitro, the bioconjugate selectively interacted with ovarian cancer cells through the CD44 receptor, disclosed a dose-dependent tumor growth inhibition efficacy comparable to that of free SN-38 drug, and inhibited Topoisomerase I function leading to apoptosis by a mechanism involving caspase-3 and -7 activation and PARP cleavage. In vivo, the intraperitoneal administration of ONCOFID-S in tumor-bearing mice did not induce inflammation, and evidenced an improved therapeutic efficacy compared with CPT-11. In conclusion, SN-38 conjugation to hyaluronan significantly improved the profile of in vivo tolerability and widened the field of application of irinotecan. Therefore, this approach can be envisaged as a promising therapeutic strategy for loco-regional treatment of ovarian cancer.

In vitro study of normoxic epidermal growth factor receptor-induced hypoxia-inducible factor-1-alpha, vascular endothelial growth factor, and BNIP3 expression in head and neck squamous cell carcinoma cell lines: Implications for anti-epidermal growth factor receptor therapy.

BACKGROUND: We previously showed that activation of epidermal growth factor receptor (EGFR) induces hypoxia inducible factor-1α (HIF-1α) in head and neck squamous cell carcinoma (HNSCC) cells. In this study, we have furthered this by investigating the mechanism of HIF-1α activation by epidermal growth factor (EGF) and its association with the sensitivity to gefitinib. METHODS: EGFR/HIF-1α signaling was tested by immunoblot, polymerase chain reaction (PCR), cell proliferation, and apoptosis assays. RESULTS: HIF-1α accumulated in cells overexpressing EGF and phosphorylated epidermal growth factor receptor (pEGFR), phosphatidylinositol-3-kinase (pPI3K), and mitogen-activated protein kinase (pMAPK). EGF-induced expression of HIF-1α and its targets, vascular endothelial growth factor (VEGF) and BNIP3, were blocked by gefitinib and PI3K-inhibitors and MAPK-inhibitors. HIF-1α-siRNAs abrogated EGF-induced BNIP3 but not VEGF expression. Gefitinib inhibited cell proliferation and induced apoptosis more strongly in cells with constitutively active EGFR/HIF-1α signaling than in cells lacking activation of these pathways. HIF-1α-siRNA treatment reduced sensitivity to gefitinib. CONCLUSION: The search for molecular predictors of sensitivity to gefitinib in HNSCC should be extended to the activation status of EGFR-downstream pathways, phosphorylated protein kinase B, pMAPK, and HIF-1α.

Peritoneal tumor carcinomatosis: pharmacological targeting with hyaluronan-based bioconjugates overcomes therapeutic indications of current drugs.

Peritoneal carcinomatosis still lacks reliable therapeutic options. We aimed at testing a drug delivery strategy allowing a controlled release of cytotoxic molecules and selective targeting of tumor cells. We comparatively assessed the efficacy of a loco-regional intraperitoneal treatment in immunocompromised mice with bioconjugates formed by chemical linking of paclitaxel or SN-38 to hyaluronan, against three models of peritoneal carcinomatosis derived from human colorectal, gastric and esophageal tumor cell xenografts. In vitro, bioconjugates were selectively internalized through mechanisms largely dependent on interaction with the CD44 receptor and caveolin-mediated endocytosis, which led to accumulation of compounds into lysosomes of tumor cells. Moreover, they inhibited tumor growth comparably to free drugs. In vivo, efficacy of bioconjugates or free drugs against luciferase-transduced tumor cells was assessed by bioluminescence optical imaging, and by recording mice survival. The intraperitoneal administration of bioconjugates in tumor-bearing mice exerted overlapping or improved therapeutic efficacy compared with unconjugated drugs. Overall, drug conjugation to hyaluronan significantly improved the profiles of in vivo tolerability and widened the field of application of existing drugs, over their formal approval or current use. Therefore, this approach can be envisaged as a promising therapeutic strategy for loco-regional treatment of peritoneal carcinomatosis.

PSMA-specific CAR-engineered T cells eradicate disseminated prostate cancer in preclinical models.

Immunology-based interventions have been proposed as a promising curative chance to effectively attack postoperative minimal residual disease and distant metastatic localizations of prostate tumors. We developed a chimeric antigen receptor (CAR) construct targeting the human prostate-specific membrane antigen (hPSMA), based on a novel and high affinity specific mAb. As a transfer method, we employed last-generation lentiviral vectors (LV) carrying a synthetic bidirectional promoter capable of robust and coordinated expression of the CAR molecule, and a bioluminescent reporter gene to allow the tracking of transgenic T cells after in vivo adoptive transfer. Overall, we demonstrated that CAR-expressing LV efficiently transduced short-term activated PBMC, which in turn were readily stimulated to produce cytokines and to exert a relevant cytotoxic activity by engagement with PSMA+ prostate tumor cells. Upon in vivo transfer in tumor-bearing mice, CAR-transduced T cells were capable to completely eradicate a disseminated neoplasia in the majority of treated animals, thus supporting the translation of such approach in the clinical setting.

Functional avidity-driven activation-induced cell death shapes CTL immunodominance.

Immunodominance is a complex phenomenon that relies on a mere numerical concept, while being potentially influenced at every step of the immune response. We investigated the mechanisms leading to the establishment of CTL immunodominance in a retroviral model and found that the previously defined subdominant Env-specific CD8(+) T cells are endowed with an unexpectedly higher functional avidity than is the immunodominant Gag-recognizing counterpart. This high avidity, along with the Env Ag overload, results in a supraoptimal TCR engagement. The overstimulation makes Env-specific T lymphocytes more susceptible to apoptosis, thus hampering their expansion and leading to an unintentional "immune kamikazing." Therefore, Ag-dependent, hyperactivation-induced cell death can be regarded as a novel mechanism in the establishment of the immunodominance that restrains and opposes the expansion of high-avidity T cells in favor of lower-affinity populations.