Latent Markov and growth mixture models for ordinal individual responses with covariates: a comparison

We propose a short review between two alternative ways of modeling stability and change of longitudinal data when time-fixed and time-varying covariates referred to the observed individuals are available. They both build on the foundation of the finite mixture models and are commonly applied in many fields. They look at the data by a different perspective and in the literature they have not been compared when the ordinal nature of the response variable is of interest. The latent Markov model is based on time-varying latent variables to explain the observable behavior of the individuals. The model is proposed in a semi-parametric formulation as the latent Markov process has a discrete distribution and it is characterized by a Markov structure. The growth mixture model is based on a latent categorical variable that accounts for the unobserved heterogeneity in the observed trajectories and on a mixture of normally distributed random variable to account for the variability of growth rates. To illustrate the main differences among them we refer to a real data example on the self reported health status

Antimicrobial Efficacy of XF-73 and Photo-Activated XF-73 Against Clinically Relevant Microorganisms in Planktonic and Biofilm Mode of Growth

Novel antimicrobials are urgently needed to combat the global increase in antimicrobial resistance. Exeporfinium chloride (XF-73) is a synthetic dicationic porphyrin antimicrobial, acting through membrane disruption; with a secondary mechanism of action, activated via blue light exposure. The aim of this study was to assess the antimicrobial efficacy of XF-73 and photo-activated XF-73 against a panel of clinically relevant Gram-positive and Gram-negative microorganisms in planktonic and biofilm modes of growth. The antimicrobial efficacy of XF-73 was assessed through minimum inhibitory and bactericidal concentration (MIC/MBC) assays. The biofilm Calgary device generated twenty-four hour biofilms to assess the anti-biofilm effect of XF-73 on biofilm viability through viable cell counts and through minimum biofilm inhibitory and eradication concentrations (MBIC/MBEC) assays. The antimicrobial activity of photo-activated XF-73 was also assessed following fifteen minutes blue light exposure at 420 nm (light dose = 13.8 J/cm2). XF-73 expressed a greater bactericidal activity against Gram-positive planktonic bacteria in comparison to Gram-negative microorganisms as MBCs ranged from ≤ 0.125 – 4 μg/ mL and 128 – >256 μg/ mL respectively. Photo-activation of XF-73 enhanced the bactericidal properties against both Gram-positive and Gram-negative bacteria as MBCs decreased to ≤ 0.03 – 1 μg/ mL and 32 – 128 μg/ mL. Gram-positive biofilms were highly susceptible to XF-73 with MBEC’s ranging from 1 – 2 μg/ mL. Photo-activation increased the bactericidal effect of XF-73 against Gram-positive biofilms with MBEC’s ranging between ≤ 0.125 – 0.5 μg/ mL. XF-73 was unable to eradicate Gram-negative biofilms, but concentrations 128 – 512 μg/ mL significantly reduced biofilm viability (p= <0.0001). XF-73 is a potent antimicrobial against Gram-positive microorganisms in both planktonic and biofilm modes of growth. Photo-activation further enhances the bactericidal effect of XF-73 and potentially offers an adjunct form of treatment in comparison to current antimicrobial treatment strategies. Further research is warranted in this area

In Silico and In Vitro Study of Antioxidant Potential of Urolithins

In this work, quantum chemical calculations based on density functional theory (DFT) were performed to predict the antioxidant potential of four bioactive gut microbiota metabolites of the natural polyphenols ellagitannins (ETs) and ellagic acid (EA), also known as urolithins (UROs). In order to evaluate their ability to counter the effect of oxidative stress caused by reactive oxygen species (ROS), such as the hydroperoxyl radical (•OOH), different reaction mechanisms were investigated, considering water and lipid-like environments. Through our in silico results, it emerged that at physiological pH, the scavenging activity of all urolithins, except urolithin B, are higher than that of trolox and other potent antioxidants existing in nature, such as EA, α-mangostin, allicin, caffeine and melatonin. These findings were confirmed by experimental assays

Synergistic inhibition of the Hedgehog pathway by newly designed Smo and Gli antagonists bearing the isoflavone scaffold

Aberrant activation of the Hedgehog (Hh) pathway is responsible for the onset and progression of several malignancies. Small molecules able to block the pathway at the upstream receptor Smoothened (Smo) or the downstream effector Gli1 have thus emerged recently as valuable anticancer agents. Here, we have designed, synthesized, and tested new Hh inhibitors taking advantage by the highly versatile and privileged isoflavone scaffold. The introduction of specific substitutions on the isoflavone's ring B allowed the identification of molecules targeting preferentially Smo or Gli1. Biological assays coupled with molecular modeling corroborated the design strategy, and provided new insights into the mechanism of action of these molecules. The combined administration of two different isoflavones behaving as Smo and Gli antagonists, respectively, in primary medulloblastoma (MB) cells highlighted the synergistic effects of these agents, thus paving the way to further and innovative strategies for the pharmacological inhibition of Hh signaling

A Smo/Gli multitarget hedgehog pathway inhibitor impairs tumor growth

Pharmacological Hedgehog (Hh) pathway inhibition has emerged as a valuable anticancer strategy. A number of small molecules able to block the pathway at the upstream receptor Smoothened (Smo) or the downstream effector glioma-associated oncogene 1 (Gli1) has been designed and developed. In a recent study, we exploited the high versatility of the natural isoflavone scaffold for targeting the Hh signaling pathway at multiple levels showing that the simultaneous targeting of Smo and Gli1 provided synergistic Hh pathway inhibition stronger than single administration. This approach seems to effectively overcome the drug resistance, particularly at the level of Smo. Here, we combined the pharmacophores targeting Smo and Gli1 into a single and individual isoflavone, compound 22, which inhibits the Hh pathway at both upstream and downstream level. We demonstrate that this multitarget agent suppresses medulloblastoma growth in vitro and in vivo through antagonism of Smo and Gli1, which is a novel mechanism of action in Hh inhibition

Close linkage with the RET protooncogene and boundaries of deletion mutations in autosomal dominant Hirschsprung disease

Tight linkage with the RET proto-oncogene (Zmax = 3.41 at θ = 0.00), analysis of recombinants and detection of a familial microdeletion in a large pedigree restrict the mapping of the Hirschsprung (HSCR) gene previously localized on proximal 10q. The molecular characterization of the familial microdeletion and of 3 additional cytogenetically visible de novo deletions, isolated in somatic cell hybrids, identify a smallest region of overlap of 250 Kb. This contains the RET proto-oncogene where missense mutations causing multiple endocrine neoplasia type 2A (MEN 2A) phenotype were recently found. The pentagastrin test (which detects preclinical forms of MEN 2A or B) is negative in adult HSCR patients with deletions of the RET gene. This represents a good candidate for the search of mutations causing HSC

cDNA Sequence and Genomic Structure of the Rat Ret Proto-Oncogene

The RET proto-oncogene, a member of the Receptor Tyrosine Kinase family, plays a crucial role during the development of the excretory system and the enteric nervous system, as demonstrated by in vivo animal studies and by its involvement in the pathogenesis of several human neurocristopathies like Hirschsprung disease and Multiple Endocrine Neoplasia type 2. Using a multistep RT-PCR approach we have isolated and sequenced the cDNA of the whole rat RET proto-oncogene, reporting the deduced amino acid sequence in comparison with the human and mouse counterparts. Moreover, two different isoforms (RET9 and RET51) have been confirmed in the rat, while a third RET isoform demonstrated in human (RET43) has not resulted to be conserved in this species. Finally, we have determined the genomic structure of the rat RET proto-oncogene comparing the exon-intron boundaries and intron sizes with the known structure of the human homologous gene. Our findings will facilitate the molecular study of appropriate rat models of RET related human diseases

Targeting SARS-CoV-2 Main Protease: A Successful Story Guided by an In Silico Drug Repurposing Approach

The SARS-CoV-2 main protease (Mpro) is a crucial enzyme for viral replication and has been considered an attractive drug target for the treatment of COVID-19. In this study, virtual screening techniques and in vitro assays were combined to identify novel Mpro inhibitors starting from around 8000 FDA-approved drugs. The docking analysis highlighted 17 promising best hits, biologically characterized in terms of their Mpro inhibitory activity. Among them, 7 cephalosporins and the oral anticoagulant betrixaban were able to block the enzyme activity in the micromolar range with no cytotoxic effect at the highest concentration tested. After the evaluation of the degree of conservation of Mpro residues involved in the binding with the studied ligands, the ligands’ activity on SARS-CoV-2 replication was assessed. The ability of betrixaban to affect SARS-CoV-2 replication associated to its antithrombotic effect could pave the way for its possible use in the treatment of hospitalized COVID-19 patient

Targeting SARS-CoV-2 nsp13 Helicase and Assessment of Druggability Pockets: Identification of Two Potent Inhibitors by a Multi-Site In Silico Drug Repurposing Approach

The SARS-CoV-2 non-structural protein 13 (nsp13) helicase is an essential enzyme for viral replication and has been identified as an attractive target for the development of new antiviral drugs. In detail, the helicase catalyzes the unwinding of double-stranded DNA or RNA in a 5′ to 3′ direction and acts in concert with the replication–transcription complex (nsp7/nsp8/nsp12). In this work, bioinformatics and computational tools allowed us to perform a detailed conservation analysis of the SARS-CoV-2 helicase genome and to further predict the druggable enzyme’s binding pockets. Thus, a structure-based virtual screening was used to identify valuable compounds that are capable of recognizing multiple nsp13 pockets. Starting from a database of around 4000 drugs already approved by the Food and Drug Administration (FDA), we chose 14 shared compounds capable of recognizing three out of four sites. Finally, by means of visual inspection analysis and based on their commercial availability, five promising compounds were submitted to in vitro assays. Among them, PF-03715455 was able to block both the unwinding and NTPase activities of nsp13 in a micromolar range