SAMHD1 expression is a surrogate marker of immune infiltration and determines prognosis after neoadjuvant chemotherapy in early breast cancer.

PURPOSE: The lack of validated surrogate biomarkers is still an unmet clinical need in the management of early breast cancer cases that do not achieve complete pathological response after neoadjuvant chemotherapy (NACT). Here, we describe and validate the use of SAMHD1 expression as a prognostic biomarker in residual disease in vivo and in vitro. METHODS: SAMHD1 expression was evaluated in a clinical cohort of early breast cancer patients with stage II-III treated with NACT. Heterotypic 3D cultures including tumor and immune cells were used to investigate the molecular mechanisms responsible of SAMHD1 depletion through whole transcriptomic profiling, immune infiltration capacity and subsequent delineation of dysregulated immune signaling pathways. RESULTS: SAMHD1 expression was associated to increased risk of recurrence and higher Ki67 levels in post-NACT tumor biopsies of breast cancer patients with residual disease. Survival analysis showed that SAMHD1-expressing tumors presented shorter time-to-progression and overall survival than SAMHD1 negative cases, suggesting that SAMHD1 expression is a relevant prognostic factor in breast cancer. Whole-transcriptomic profiling of SAMHD1-depleted tumors identified downregulation of IL-12 signaling pathway as the molecular mechanism determining breast cancer prognosis. The reduced interleukin signaling upon SAMHD1 depletion induced changes in immune cell infiltration capacity in 3D heterotypic in vitro culture models, confirming the role of the SAMHD1 as a regulator of breast cancer prognosis through the induction of changes in immune response and tumor microenvironment. CONCLUSION: SAMHD1 expression is a novel prognostic biomarker in early breast cancer that impacts immune-mediated signaling and differentially regulates inflammatory intra-tumoral response.

SAMHD1 expression modulates innate immune activation and correlates with ovarian cancer prognosis.

Purpose: SAMHD1 is a deoxynucleotide triphosphate (dNTP) triphosphohydrolase which has been proposed as a putative prognostic factor in haematological cancers and certain solid tumours, although with controversial data. Here, we evaluate SAMHD1 function in ovarian cancer, both in vitro and in ovarian cancer patients. Methods: SAMHD1 expression was downregulated in ovarian cancer cell lines OVCAR3 and SKOV3 by RNA interference. Gene and protein expression changes in immune signalling pathways were assessed. SAMHD1 expression in ovarian cancer patients was evaluated by immunohistochemistry and survival analysis was performed according to SAMHD1 expression. Results: SAMHD1 knockdown induced a significant upregulation of proinflammatory cytokines concomitant to increased expression of the main RNA-sensors, MDA5 and RIG-I, and interferon-stimulated genes, supporting the idea that the absence of SAMHD1 promotes innate immune activation in vitro. To assess the contribution of SAMHD1 in ovarian cancer patients, tumours were stratified in SAMHD1-low and SAMHD1-high expressing tumours, resulting in significantly shorter progression free survival (PFS) and overall survival (OS) in SAMHD1-high expression subgroup (p=0.01 and 0.04, respectively). Conclusions: SAMHD1 depletion correlates with increased innate immune cell signalling in ovarian cancer cells. In clinical samples, SAMHD1-low expressing tumors showed increased progression free survival and overall survival irrespective of BRCA mutation status. These results point towards SAMHD1 modulation as a new therapeutic strategy, able to enhance innate immune activation directly in tumour cells, leading to improved prognosis in ovarian cancer.

Pharmacological Inhibition of IKK to Tackle Latency and Hyperinflammation in Chronic HIV-1 Infection.

HIV latent infection may be associated with disrupted viral RNA sensing, interferon (IFN) signaling, and/or IFN stimulating genes (ISG) activation. Here, we evaluated the use of compounds selectively targeting at the inhibitor of nuclear factor-κB (IκB) kinase (IKK) complex subunits and related kinases (TBK1) as a novel pathway to reverse HIV-1 latency in latently infected non-clonal lymphoid and myeloid cell in vitro models. IKK inhibitors (IKKis) triggered up to a 1.8-fold increase in HIV reactivation in both, myeloid and lymphoid cell models. The best-in-class IKKis, targeting TBK-1 (MRT67307) and IKKß (TCPA-1) respectively, were also able to significantly induce viral reactivation in CD4+ T cells from people living with HIV (PLWH) ex vivo. More importantly, although none of the compounds tested showed antiviral activity, the combination of the distinct IKKis with ART did not affect the latency reactivation nor blockade of HIV infection by ART. Finally, as expected, IKKis did not upregulate cell activation markers in primary lymphocytes and innate immune signaling was blocked, resulting in downregulation of inflammatory cytokines. Overall, our results support a dual role of IKKis as immune modulators being able to tackle the HIV latent reservoir in lymphoid and myeloid cellular models and putatively control the hyperinflammatory responses in chronic HIV-1 infection.

Phylogeography of hepatitis B virus: The role of Portugal in the early dissemination of HBV worldwide.

In Portugal, the genetic diversity, origin of HBV and the Portuguese role in the dissemination of HBV worldwide were never investigated. In this work, we studied the epidemic history and transmission dynamics of HBV genotypes that are endemic in Portugal. HBV pol gene was sequenced from 130 patients followed in Lisbon. HBV genotype A was the most prevalent (n = 54, 41.5%), followed by D (n = 44, 33.8%), and E (n = 32, 24.6%). Spatio-temporal evolutionary dynamics was reconstructed in BEAST using a Bayesian Markov Chain Monte Carlo method, with a GTR nucleotide substitution model, an uncorrelated lognormal relaxed molecular clock model, a Bayesian skyline plot, and a continuous diffusion model. HBV subgenotype D4 was the first to be introduced in Portugal around 1857 (HPD 95% 1699-1931) followed by D3 and A2 a few decades later. HBV genotype E and subgenotype A1 were introduced in Portugal later, almost simultaneously. Our results indicate a very important role of Portugal in the exportation of subgenotypes D4 and A2 to Brazil and Cape Verde, respectively, in the beginning of the XX century. This work clarifies the epidemiological history of HBV in Portugal and provides new insights in the early and global epidemic history of this virus.

IRF7 expression correlates with HIV latency reversal upon specific blockade of immune activation.

The persistence of latent HIV reservoirs allows for viral rebound upon antiretroviral therapy interruption, hindering effective HIV-1 cure. Emerging evidence suggests that modulation of innate immune stimulation could impact viral latency and contribute to the clearing of HIV reservoir. Here, the latency reactivation capacity of a subclass of selective JAK2 inhibitors was characterized as a potential novel therapeutic strategy for HIV-1 cure. Notably, JAK2 inhibitors reversed HIV-1 latency in non-clonal lymphoid and myeloid in vitro models of HIV-1 latency and also ex vivo in CD4+ T cells from ART+ PWH, albeit its function was not dependent on JAK2 expression. Immunophenotypic characterization and whole transcriptomic profiling supported reactivation data, showing common gene expression signatures between latency reactivating agents (LRA; JAK2i fedratinib and PMA) in contrast to other JAK inhibitors, but with significantly fewer affected gene sets in the pathway analysis. In depth evaluation of differentially expressed genes, identified a significant upregulation of IRF7 expression despite the blockade of the JAK-STAT pathway and downregulation of proinflammatory cytokines and chemokines. Moreover, IRF7 expression levels positively correlated with HIV latency reactivation capacity of JAK2 inhibitors and also other common LRAs. Collectively, these results represent a promising step towards HIV eradication by demonstrating the potential of innate immune modulation for reducing the viral reservoir through a novel pathway driven by IRF7.

Healthcare provider-to-patient perspectives on the uptake of teleconsultation services in the Nigerian healthcare system during the COVID-19 pandemic era.

The urgency to curtail the devastating effects of the ongoing COVID-19 Pandemic has led to the implementation of several measures to limit its spread, including movement restrictions and social distancing. As most developing countries rely solely on hospital visitations for their medical needs, this impediment to assessing healthcare services compounded by low uptake of telehealth services could result in dire consequences. This is a cross-sectional study among Healthcare providers (HCP) and Healthcare consumers (HCC) in Nigeria. We administered a pre-validated self-administered online questionnaire comprising questions to assess the knowledge, use, perceptions, and benefits of telemedicine among study participants. Descriptive statistics were used to examine participants' perceptions on telemedicine use and to summarize participants' characteristics. A total of 158 healthcare providers and 1381 healthcare consumers completed the online survey. Ninety percent of HCP reported that they used some form of telemedicine to deliver health care, and 63% of HCC had received healthcare through telemedicine. A significant proportion of HCP (62%) and HCC (69%) agreed that telemedicine would improve healthcare consultation experience and satisfaction. However, fewer (21%) HCP agreed that they liked that there would be no physical contact with patients using telemedicine. In contrast, 52% of HCC agreed that they liked that there would be no physical contact with healthcare providers while using telemedicine. The majority of the participants believed that benefits of telemedicine would include: being a safe way for healthcare delivery during pandemics (HCP = 62%, HCC = 83%), affordability (HCP = 62%, HCC = 82%), and time-saving (HCP = 54%, HCC = 82%,). Teleconsultation services have been shown to aptly complement face-to-face hospital visits in ensuring effective triaging in hospitals and providing adequate healthcare delivery to patients regardless of geographical and physical barriers. These results support telemedicine use for the provision of healthcare services during the COVID-19 pandemic.

JAK-STAT Pathway: A Novel Target to Tackle Viral Infections.

Modulation of the antiviral innate immune response has been proposed as a putative cellular target for the development of novel pan-viral therapeutic strategies. The Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway is especially relevant due to its essential role in the regulation of local and systemic inflammation in response to viral infections, being, therefore, a putative therapeutic target. Here, we review the extraordinary diversity of strategies that viruses have evolved to interfere with JAK-STAT signaling, stressing the relevance of this pathway as a putative antiviral target. Moreover, due to the recent remarkable progress on the development of novel JAK inhibitors (JAKi), the current knowledge on its efficacy against distinct viral infections is also discussed. JAKi have a proven efficacy against a broad spectrum of disorders and exhibit safety profiles comparable to biologics, therefore representing good candidates for drug repurposing strategies, including viral infections.

Cyclin-dependent Kinases as Emerging Targets for Developing Novel Antiviral Therapeutics.

Besides its prominent role in cell proliferation, cyclin-dependent kinases (CDKs) are key players in viral infections as both DNA and RNA viruses modify CDK function to favor viral replication. Recently, a number of specific pharmacological CDK inhibitors have been developed and approved for cancer treatment. The repurposing of these specific CDK inhibitors for the treatment of viral infections may represent a novel effective therapeutic strategy to combat old and emergent viruses. In this review, we describe the role, mechanisms of action, and potential of CDKs as antiviral drug targets. We also discuss the current clinical state of novel specific CDK inhibitors, focusing on their putative use as antivirals, especially against new emerging viruses.

L-alanine supplementation improves blood glucose level and biochemical indices in alloxan-induced diabetic rats.

Diabetes is a metabolic disorder whose complications are among the leading cause of death. In this study, the antidiabetic effect of L-alanine was tested in alloxan-induced diabetic rats. Thirty-five male albino Wistar rats were divided into five groups viz; Group I and II: nondiabetic and diabetic controls respectively; Group III and IV: 150 and 300 mg/kg b.w. L-alanine treated, respectively; Group V: glibenclamide (0.5 mg/kg b.w.) treated. Weight and blood glucose were monitored during the study, while liver and kidney functions, lipid profile, and antioxidant markers were examined at the end of the study. The outcomes indicate that 300 mg/kg L-alanine resulted to a significant decrease (p < .05) in weight and blood glucose. L-alanine restored tissue antioxidants, kidney, and liver functions by improving important parameters. Histopathological studies showed the potential of L-alanine in regeneration of the islets of Langerhans. These findings suggest that L-alanine has an alleviating effect on alloxan-induced diabetes. PRACTICAL APPLICATIONS: Several medicinal plants have been tested for their antidiabetic potentials, however, the isolation of the active compounds from these plants for medicinal use is often challenging. Here, we present data that suggests the potential use of a pure and harmless amino acid compound (L-alanine) for the management of diabetes. L-alanine is readily available, cheap and can also be found in many foods we eat. Therefore, L-alanine may be taken by diabetic patients as a food supplement for the treatment/management of diabetes or taken as part of foods rich in the amino acid such as meat, poultry, fish, eggs, and dairy products.

Epidemic history and baseline resistance to NS5A-specific direct acting drugs of hepatitis C virus in Spain.

Hepatitis C virus (HCV) infection remains a global health problem. Previously, the prevalence of NS5A resistance-associated substitutions (RASs) to elbasvir, a new direct-acting antiviral (DAA) against the NS5A viral protein was assessed by our group before its introduction into clinical use in Spain. However, the origin, epidemic history, transmission dynamics, diversity and baseline RASs to NS5A direct-acting agents of HCV-GT1a in Spain remain unknown. A nationwide cross-sectional survey of individuals chronically-infected with HCV-G1a and DAAs-naïve was performed. HCV population sequencing, phylogenetic analysis and Bayesian methods were used. GT1a clade II was more prevalent than clade I (82.3% vs. 17.7%; P < 0.001) and older (estimated origin in 1912 vs. 1952). Clade II epidemic is currently declining whereas clade I epidemic has reached equilibrium. A total of 58 single RASs were identified, which account for the moderate level (10%) of baseline resistance observed. When considering the regional data, marked differences were observed, with thirteen regions showing an intermediate level (5-15%) and one a high level (20%) of resistance. Current HCV-GT1a epidemic in Spain is driven by clade I which seem to have different dissemination routes relative to clade II. A moderate level of baseline RASs to NS5A-DAAs with marked differences among regions was observed. Close surveillance of response to treatment with DAAs will be important.

Erratum: Castellví, M. et al. Pharmacological Modulation of SAMHD1 Activity by CDK4/6 Inhibitors Improves Anticancer Therapy. Cancers 2020, 12, 713.

The authors wish you make the following corrections to this paper[...].

Pharmacological Modulation of SAMHD1 Activity by CDK4/6 Inhibitors Improves Anticancer Therapy.

Sterile alpha motif and histidine-aspartic acid domain-containing protein 1 (SAMHD1) is a dNTP triphosphohydrolase involved in the regulation of the intracellular dNTP pool, linked to viral restriction, cancer development and autoimmune disorders. SAMHD1 function is regulated by phosphorylation through a mechanism controlled by cyclin-dependent kinases and tightly linked to cell cycle progression. Recently, SAMHD1 has been shown to decrease the efficacy of nucleotide analogs used as chemotherapeutic drugs. Here, we demonstrate that SAMHD1 can enhance or decrease the efficacy of various classes of anticancer drug, including nucleotide analogues, but also anti-folate drugs and CDK inhibitors. Importantly, we show that selective CDK4/6 inhibitors are pharmacological activators of SAMHD1 that act by inhibiting its inactivation by phosphorylation. Combinations of a CDK4/6 inhibitor with nucleoside or folate antimetabolites potently enhanced drug efficacy, resulting in highly synergic drug combinations (CI < 0.04). Mechanistic analyses reveal that cell cycle-controlled modulation of SAMHD1 function is the central process explaining changes in anticancer drug efficacy, therefore providing functional proof of the potential of CDK4/6 inhibitors as a new class of adjuvants to boost chemotherapeutic regimens. The evaluation of SAMHD1 expression in cancer tissues allowed for the identification of cancer types that would benefit from the pharmacological modulation of SAMHD1 function. In conclusion, these results indicate that the modulation of SAMHD1 function may represent a promising strategy for the improvement of current antimetabolite-based treatments.

Epidemic history of hepatitis C virus genotypes and subtypes in Portugal.

Any successful strategy to prevent and control HCV infection requires an understanding of the epidemic behaviour among the different genotypes. Here, we performed the first characterization of the epidemic history and transmission dynamics of HCV subtypes in Portugal. Direct sequencing of NS5B was performed on 230 direct-acting antiviral drugs (DAA)-treatment naïve patients in Lisbon. Phylogenetic analysis was used for subtyping and transmission cluster identification. Bayesian methods were used to reconstruct the epidemic history of HCV subtypes. Sequences were analysed for resistance-associated substitutions (RAS). The majority of strains were HCV-GT1 (62.6%), GT3 (18.3%, all subtype 3a) and GT4 (16.1%). Among GT1, the most frequent were subtypes 1a (75.5%) and 1b (24.5%). Polyphyletic patterns were found in all but 12 lineages suggesting multiple introductions of the different subtypes in this population. Five distinct epidemics were identified. The first significant HCV epidemic in Portugal occurred between 1930s and 1960s, was caused almost exclusively by GT1b and was likely associated with blood transfusions. Rapid expansion of GT3a occurred in the 1960s and GT1a in the 1980s, associated with intravenous drug use. The most recent epidemics were caused by GT4a and GT4d and seem to be associated with the resurgence of opioid use. The C316N substitution was found in 31.4% of GT1b-patients. Close surveillance of patients bearing this mutation and undergoing dasabuvir-based regimens will be important to determine its impact on treatment outcome.