Shallow whole genome sequencing approach to detect Homologous Recombination Deficiency in the PAOLA-1/ENGOT-OV25 phase-III trial.

The bevacizumab (bev)/olaparib (ola) maintenance regimen was approved for BRCA1/2-mutated (BRCAmut) and Homologous Recombination Deficient (HRD) high-grade Advanced Ovarian Cancer (AOC) first line setting, based on a significantly improved progression-free survival (PFS) compared to bev alone in the PAOLA-1/ENGOT-ov25 trial (NCT02477644), where HRD was detected by MyChoice CDx PLUS test. The academic shallowHRDv2 test was developed based on shallow whole-genome sequencing as an alternative to MyChoice. Analytical and clinical validities of shallowHRDv2 as compared to MyChoice on 449 PAOLA-1 tumor samples are presented. The overall agreement between shallowHRDv2 and MyChoice was 94% (369/394). Less non-contributive tests were observed with shallowHRDv2 (15/449; 3%) than with MyChoice (51/449; 11%). Patients with HRD tumors according to shallowHRDv2 (including BRCAmut) showed a significantly prolonged PFS with bev+ola versus bev (median PFS: 65.7 versus 20.3 months, hazard ratio (HR): 0.36 [95% CI: 0.24-0.53]). This benefit was significant also for BRCA1/2 wild-type tumors (40.8 versus 19.5 months, HR: 0.45 [95% CI: 0.26-0.76]). ShallowHRDv2 is a performant, clinically validated, and cost-effective test for HRD detection.

Glycolytic shift during West Nile virus infection provides new therapeutic opportunities.

BACKGROUND: Viral rewiring of host bioenergetics and immunometabolism may provide novel targets for therapeutic interventions against viral infections. Here, we have explored the effect on bioenergetics during the infection with the mosquito-borne flavivirus West Nile virus (WNV), a medically relevant neurotropic pathogen causing outbreaks of meningitis and encephalitis worldwide. RESULTS: A systematic literature search and meta-analysis pointed to a misbalance of glucose homeostasis in the central nervous system of WNV patients. Real-time bioenergetic analyses confirmed upregulation of aerobic glycolysis and a reduction of mitochondrial oxidative phosphorylation during viral replication in cultured cells. Transcriptomics analyses in neural tissues from experimentally infected mice unveiled a glycolytic shift including the upregulation of hexokinases 2 and 3 (Hk2 and Hk3) and pyruvate dehydrogenase kinase 4 (Pdk4). Treatment of infected mice with the Hk inhibitor, 2-deoxy-D-glucose, or the Pdk4 inhibitor, dichloroacetate, alleviated WNV-induced neuroinflammation. CONCLUSIONS: These results highlight the importance of host energetic metabolism and specifically glycolysis in WNV infection in vivo. This study provides proof of concept for the druggability of the glycolytic pathway for the future development of therapies to combat WNV pathology.

Submucosal fat accumulation in Crohn's disease: evaluation with sonography.

BACKGROUND/AIMS: The study objective is to investigate the ultrasound features that allow suspecting the presence of submucosal fat deposition, called the fat halo sign (FHS), in the intestinal wall of patients with Crohn's disease. METHODS: Computed tomography (CT) examinations over a period of 10 years were reviewed for the presence of the FHS in the bowel wall. A measurement of less than -10 Hounsfield units was regarded as indicative of fat. We included only patients who had undergone ultrasound examinations 3 months before or after CT. The study cohort group comprised 68 patients. Wall and submucosal thickness were measured on longitudinal ultrasound sections. A receiver operating characteristic curve was constructed to determine the best cutoff of ultrasound submucosal wall thickness value for predicting FHS in the bowel wall determined on CT. RESULTS: The FHS was present in 22 patients (31%) on CT. There were significant differences between submucosal thickness of patients with FHS and patients without FHS (4.19 mm vs. 2.41 mm). From the receiver operating characteristic curve, a threshold value of 3.1 mm of submucosal thickness had the best sensitivity and specificity to suspect FHS (95.5% and 89.1%, respectively; area under the curve, 0.962), with an odds ratio of 172. All of 16 patients with a submucosal thickness >3.9 mm had FHS. CONCLUSIONS: FHS in patients with Crohn's disease can be suspected on ultrasound in cases with marked thickening of the submucosa layer. In these cases, the activity of the disease should be measured by other parameters such as the color Doppler.

C-2 Thiophenyl Tryptophan Trimers Inhibit Cellular Entry of SARS-CoV-2 through Interaction with the Viral Spike (S) Protein.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes COVID-19, by infecting cells via the interaction of its spike protein (S) with the primary cell receptor angiotensin-converting enzyme (ACE2). To search for inhibitors of this key step in viral infection, we screened an in-house library of multivalent tryptophan derivatives. Using VSV-S pseudoparticles, we identified compound 2 as a potent entry inhibitor lacking cellular toxicity. Chemical optimization of 2 rendered compounds 63 and 65, which also potently inhibited genuine SARS-CoV-2 cell entry. Thermofluor and microscale thermophoresis studies revealed their binding to S and to its isolated receptor binding domain (RBD), interfering with the interaction with ACE2. High-resolution cryoelectron microscopy structure of S, free or bound to 2, shed light on cell entry inhibition mechanisms by these compounds. Overall, this work identifies and characterizes a new class of SARS-CoV-2 entry inhibitors with clear potential for preventing and/or fighting COVID-19.

Direct reductive amination of functionalized aldehydes with aniline derivatives of purines and 7-deazapurines.

Reductive amination plays a key role in the medicinal chemistry toolbox since it allows the mono alkylation of an amine or aniline. In this work, reductive amination of functionalized aldehydes with aniline derivatives of adenine and closely related 7-deazapurines has been successfully performed using H-cube technology so that imine formation and its reduction are performed "in situ". The set-up procedure surmounts some of the drawbacks of "in batch" protocols by avoiding the handling of reductant reagents, long reaction times and tedious work-ups. The here described procedure allows a high conversion into the reductive amination products together with an easy work-up by just evaporation. More interestingly, this set-up does not require the presence of acids so that acid-sensitive protecting groups can be present both at the aldehyde and at the heterocycle.

Pharmacological Elevation of Cellular Dihydrosphingomyelin Provides a Novel Antiviral Strategy against West Nile Virus Infection.

The flavivirus life cycle is strictly dependent on cellular lipid metabolism. Polyphenols like gallic acid and its derivatives are promising lead compounds for new therapeutic agents as they can exert multiple pharmacological activities, including the alteration of lipid metabolism. The evaluation of our collection of polyphenols against West Nile virus (WNV), a representative medically relevant flavivirus, led to the identification of N,N'-(dodecane-1,12-diyl)bis(3,4,5-trihydroxybenzamide) and its 2,3,4-trihydroxybenzamide regioisomer as selective antivirals with low cytotoxicity and high antiviral activity (half-maximal effective concentrations [EC50s] of 2.2 and 0.24 µM, respectively, in Vero cells; EC50s of 2.2 and 1.9 µM, respectively, in SH-SY5Y cells). These polyphenols also inhibited the multiplication of other flaviviruses, namely, Usutu, dengue, and Zika viruses, exhibiting lower antiviral or negligible antiviral activity against other RNA viruses. The mechanism underlying their antiviral activity against WNV involved the alteration of sphingolipid metabolism. These compounds inhibited ceramide desaturase (Des1), promoting the accumulation of dihydrosphingomyelin (dhSM), a minor component of cellular sphingolipids with important roles in membrane properties. The addition of exogenous dhSM or Des1 blockage by using the reference inhibitor GT-11 {N-[(1R,2S)-2-hydroxy-1-hydroxymethyl-2-(2-tridecyl-1-cyclopropenyl)ethyl]octanamide} confirmed the involvement of this pathway in WNV infection. These results unveil the potential of novel antiviral strategies based on the modulation of the cellular levels of dhSM and Des1 activity for the control of flavivirus infection.

Identification of West Nile virus RNA-dependent RNA polymerase non-nucleoside inhibitors by real-time high throughput fluorescence screening.

West Nile virus (WNV) is a re-emergent mosquito-borne RNA virus that causes major outbreaks of encephalitis around the world. However, there is no therapeutic treatment to struggle against WNV, and the current treatment relies on alleviating symptoms. Therefore, due to the threat virus poses to animal and human health, there is an urgent need to come up with fast strategies to identify and assess effective antiviral compounds. A relevant target when developing drugs against RNA viruses is the viral RNA-dependent RNA polymerase (RdRp), responsible for the replication of the viral genome within a host cell. RdRps are key therapeutic targets based on their specificity for RNA and their essential role in the propagation of the infection. We have developed a fluorescence-based method to measure WNV RdRp activity in a fast and reliable real-time way. Interestingly, rilpivirine has shown in our assay inhibition of the WNV RdRp activity with an IC50 value of 3.3 µM and its antiviral activity was confirmed in cell cultures. Furthermore, this method has been extended to build up a high-throughput screening platform to identify WNV polymerase inhibitors. By screening a small chemical library, novel RdRp inhibitors 1-4 have been identified. When their antiviral activity was tested against WNV in cell culture, 4 exhibited an EC50 value of 2.5 µM and a selective index of 12.3. Thus, rilpivirine shows up as an interesting candidate for repurposing against flavivirus. Moreover, the here reported method allows the rapid identification of new WNV RdRp inhibitors.

Salicylanilides and Their Anticancer Properties.

Salicylanilides are pharmacologically active compounds with a wide spectrum of biological effects. Halogenated salicylanilides, which have been used for decades in human and veterinary medicine as anthelmintics, have recently emerged as candidates for drug repurposing in oncology. The most prominent example of salicylanilide anthelmintic, that is intensively studied for its potential anticancer properties, is niclosamide. Nevertheless, recent studies have discovered extensive anticancer potential in a number of other salicylanilides. This potential of their anticancer action is mediated most likely by diverse mechanisms of action such as uncoupling of oxidative phosphorylation, inhibition of protein tyrosine kinase epidermal growth factor receptor, modulation of different signaling pathways as Wnt/ß-catenin, mTORC1, STAT3, NF-κB and Notch signaling pathways or induction of B-Raf V600E inhibition. Here we provide a comprehensive overview of the current knowledge about the proposed mechanisms of action of anticancer activity of salicylanilides based on preclinical in vitro and in vivo studies, or structural requirements for such an activity.

Development of SARS-CoV-2 specific IgG and IgA antibodies in serum and milk with different SARS-COV-2 vaccines in lactating women.

BACKGROUND: Our main objective was to determine the evolution of IgG and IgA antibodies directed against SARS-CoV-2 protein S in the blood of lactating women and in breast milk. METHODS: A cohort of 110 uninfected and vaccinated breastfeeding women was followed-up for 6 months at the Marqués de Valdecilla University Hospital, Spain, in 2020. An additional group of 23 breastfeeding mothers who had no previously documented infection and had not been vaccinated against SARS-CoV-2 were included as a control group. The antibodies in blood and breast milk and their evolution at 6 months post-vaccination were analysed. RESULTS: One hundred ten breastfeeding mothers were included; 70 women (63.6%) were vaccinated with two doses of BNT162b2, 20 women (18.2%) received two doses of mRNA-1273, and 20 women (18.2%) received a single dose of ChAdOx1-S. No evidence of differences between concentrations of antibodies was found according to the type of vaccine, with the exception of serum IgA antibodies, which was higher in women vaccinated with mRNA-1273: mean [95%CI]: 0.05 AU/mL [0.03,0.06] with mRNA-1273, 0.02 AU/mL [0.01,0.03] with BNT162b2 and 0.01 AU/mL [0.00,0.03] with ChAdOx1-S, ANOVA p value = 0.03. The lack of difference between vaccines was also found when anti-S1 specific IgG in serum and breast milk were measured. CONCLUSIONS: In lactating women vaccinated against COVID-19, anti-SARS-CoV-2 antibodies can be detected in both serum and breastmilk 6 months after receiving the second dose, although their concentrations decreased when compared with concentrations reached immediately after vaccination.

Allosteric Inhibition of Neutral Sphingomyelinase 2 (nSMase2) by DPTIP: From Antiflaviviral Activity to Deciphering Its Binding Site through In Silico Studies and Experimental Validation.

Flavivirus comprises globally emerging and re-emerging pathogens such as Zika virus (ZIKV), Dengue virus (DENV), and West Nile virus (WNV), among others. Although some vaccines are available, there is an unmet medical need as no effective antiviral treatment has been approved for flaviviral infections. The development of host-directed antivirals (HDAs) targeting host factors that are essential for viral replication cycle offers the opportunity for the development of broad-spectrum antivirals. In the case of flaviviruses, recent studies have revealed that neutral sphingomyelinase 2, (nSMase2), involved in lipid metabolism, plays a key role in WNV and ZIKV infection. As a proof of concept, we have determined the antiviral activity of the non-competitive nSMase2 inhibitor DPTIP against WNV and ZIKV virus. DPTIP showed potent antiviral activity with EC50 values of 0.26 µM and 1.56 µM for WNV and ZIKV, respectively. In order to unravel the allosteric binding site of DPTIP in nSMase2 and the details of the interaction, computational studies have been carried out. These studies have revealed that DPTIP could block the DK switch in nSMase2. Moreover, the analysis of the residues contributing to the binding identified His463 as a crucial residue. Interestingly, the inhibitory activity of DPTIP on the H463A mutant protein supported our hypothesis. Thus, an allosteric cavity in nSMase2 has been identified that can be exploited for the development of new inhibitors with anti-flaviviral activity.

Meeting report: 34th international conference on antiviral research.

As a result of the multiple gathering and travels restrictions during the SARS-CoV-2 pandemic, the annual meeting of the International Society for Antiviral Research (ISAR), the International Conference on Antiviral Research (ICAR), could not be held in person in 2021. Nonetheless, ISAR successfully organized a remote conference, retaining the most critical aspects of all ICARs, a collegiate gathering of researchers in academia, industry, government and non-governmental institutions working to develop, identify, and evaluate effective antiviral therapy for the benefit of all human beings. This article highlights the 2021 remote meeting, which presented the advances and objectives of antiviral and vaccine discovery, research, and development. The meeting resulted in a dynamic and effective exchange of ideas and information, positively impacting the prompt progress towards new and effective prophylaxis and therapeutics.

Hybridization Approach to Identify Salicylanilides as Inhibitors of Tubulin Polymerization and Signal Transducers and Activators of Transcription 3 (STAT3).

The superimposition of the X-ray complexes of cyclohexanediones (i.e., TUB015), described by our research group, and nocodazole, within the colchicine binding site of tubulin provided an almost perfect overlap of both ligands. This structural information led us to propose hybrids of TUB015 and nocodazole using a salicylanilide core structure. Interestingly, salicylanilides, such as niclosamide, are well-established signal transducers and activators of transcription (STAT3) inhibitors with anticancer properties. Thus, different compounds with this new scaffold have been synthesized with the aim to identify compounds inhibiting tubulin polymerization and/or STAT3 signaling. As a result, we have identified new salicylanilides (6 and 16) that showed significant antiproliferative activity against a panel of cancer cells. Both compounds were able to reduce the levels of p-STAT3Tyr705 without affecting the total expression of STAT3. While compound 6 inhibited tubulin polymerization and arrested the cell cycle of DU145 cells at G2/M, similar to TUB015, compound 16 showed a more potent effect on inhibiting STAT3 phosphorylation and arrested the cell cycle at G1/G0, similar to niclosamide. In both cases, no toxicity towards PBMC cells was detected. Thus, the salicylanilides described here represent a new class of antiproliferative agents affecting tubulin polymerization and/or STAT3 phosphorylation.

SEOM-GEICO clinical guidelines on endometrial cancer (2021)

Endometrial cancer (EC) is the second most common gynecological malignancy worldwide, the first in developed countries [Sung et al. in CA Cancer J Clin 71:209–249, 2021]. Although a majority is diagnosed at an early stage with a low risk of relapse, an important proportion of patients will relapse. Better knowledge of molecular abnormalities is crucial to identify high-risk groups in early stages as well as for recurrent or metastatic disease for whom adjuvant treatment must be personalized. The objective of this guide is to summarize the current evidence for the diagnosis, treatment, and follow-up of EC, and to provide evidence-based recommendations for clinical practice.

SEOM-GEICO clinical guidelines on endometrial cancer (2021).

Endometrial cancer (EC) is the second most common gynecological malignancy worldwide, the first in developed countries [Sung et al. in CA Cancer J Clin 71:209-249, 2021]. Although a majority is diagnosed at an early stage with a low risk of relapse, an important proportion of patients will relapse. Better knowledge of molecular abnormalities is crucial to identify high-risk groups in early stages as well as for recurrent or metastatic disease for whom adjuvant treatment must be personalized. The objective of this guide is to summarize the current evidence for the diagnosis, treatment, and follow-up of EC, and to provide evidence-based recommendations for clinical practice.

How to strengthen the management of acute diverticulitis: the utility of the WSES classification-a prospective single-center observational study.

PURPOSE: This study aimed to validate the World Society for Emergent Surgery (WSES) scale for the management of acute left-sided colonic diverticulitis (ALCD). METHODS: An observational study based on a prospective database of patients with ultrasound (US) and computerized tomography (CT) confirmed ALCD was conducted at our center from April 2018 to May 2019. The primary outcome was the success rate of outpatient management. Secondary outcomes were the association between different WSES stages, clinical and analytical parameters, treatments modalities, and outcomes, and the accuracy of US for management decisions. RESULTS: A total of 230 patients were included. Outpatient management was successful in 51/53 (96.23%) cases with ALCD stage 0 and 62/72 (86.11%) patients with stage 1A. There were no differences in age (p = 0.076) or the presence of pericolic air bubbles (p = 0.06) between patients who underwent admission or outpatient management. Clinical and analytical data, treatment decisions, and outcomes showed statistically significant differences between WSES stages. In 7/12 patients with stage 2A, percutaneous drainage or emergency surgery was required. All cases with stage 2B (distant air) underwent conservative management without the need for emergency or elective surgery. The accuracy of US WSES stages for management decisions, when compared with CT, was 96.96%. CONCLUSION: The WSES classification for ALCD seemed to be valid helping clinicians in the decision-making process to select between admission or outpatient management. Differences in clinical and analytical data, elected treatments, and outcomes were found between WSES stages. The US WSES stages showed high accuracy for management decisions.

Antivirals against (Re)emerging Flaviviruses: Should We Target the Virus or the Host?

The COVID pandemic has evidenced how vulnerable we are to emerging infectious diseases and how short our current armamentarium is. Flavivirus, single stranded RNA viruses transmitted by arthropods, are considered a global health challenge. No drugs to treat these infections have been approved. In this Viewpoint, we analyze the advantages and disadvantages of two different, but probably also complementary, therapeutic approaches: virus-targeting antivirals and host-targeting drugs.

Inhibition of XPO-1 Mediated Nuclear Export through the Michael-Acceptor Character of Chalcones.

The nuclear export receptor exportin-1 (XPO1, CRM1) mediates the nuclear export of proteins that contain a leucine-rich nuclear export signal (NES) towards the cytoplasm. XPO1 is considered a relevant target in different human diseases, particularly in hematological malignancies, tumor resistance, inflammation, neurodegeneration and viral infections. Thus, its pharmacological inhibition is of significant therapeutic interest. The best inhibitors described so far (leptomycin B and SINE compounds) interact with XPO1 through a covalent interaction with Cys528 located in the NES-binding cleft of XPO1. Based on the well-established feature of chalcone derivatives to react with thiol groups via hetero-Michael addition reactions, we have synthesized two series of chalcones. Their capacity to react with thiol groups was tested by incubation with GSH to afford the hetero-Michael adducts that evolved backwards to the initial chalcone through a retro-Michael reaction, supporting that the covalent interaction with thiols could be reversible. The chalcone derivatives were evaluated in antiproliferative assays against a panel of cancer cell lines and as XPO1 inhibitors, and a good correlation was observed with the results obtained in both assays. Moreover, no inhibition of the cargo export was observed when the two prototype chalcones 9 and 10 were tested against a XPO1-mutated Jurkat cell line (XPO1C528S), highlighting the importance of the Cys at the NES-binding cleft for inhibition. Finally, their interaction at the molecular level at the NES-binding cleft was studied by applying the computational tool CovDock.

Declared Reasons for Cessation of Breastfeeding during the First Year of Life: An Analysis Based on a Cohort Study in Northern Spain.

BACKGROUND: Breastfeeding is the gold standard of infant feeding due to the many advantages it offers to both the child and the mother. OBJECTIVE: To identity the main reasons for cessation of breastfeeding reported by mothers during the first year of life. DESIGN: A prospective cohort study was conducted, recruiting 970 infants from a university hospital in Spain. The main maternal variables studied were maternal age, parity, educational level, work occupation, smoking habit, weeks of gestation at birth, birth weight, feeding type, and duration of breastfeeding. All participants were followed for one year to determinate the duration of breastfeeding and to gather reasons for abandoning breastfeeding. RESULTS: At six months, the percentage of breastfeeding experienced a decline of 50%, and only 24.5% of these mothers maintained breastfeeding. Up to 15.8% of the mothers decided to give up exclusive breastfeeding by their own choice, whereas 15.4% did so because they suspected low milk production. Work-related causes represent the third reason of abandonment. CONCLUSIONS: Our results highlight the need to improve the health policies for the promotion, protection, and support for the initiation and maintenance of breastfeeding. In particular, our results highlight the importance of researching women's low milk production and work-related factors, with particular emphasis on improving conciliation measures.