Engineered smart materials for RNA based molecular therapy to treat Glioblastoma.

Glioblastoma (GBM) is an aggressive malignancy of the central nervous system (CNS) that remains incurable despite the multitude of improvements in cancer therapeutics. The conventional chemo and radiotherapy post-surgery have only been able to improve the prognosis slightly; however, the development of resistance and/or tumor recurrence is almost inevitable. There is a pressing need for adjuvant molecular therapies that can successfully and efficiently block tumor progression. During the last few decades, non-coding RNAs (ncRNAs) have emerged as key players in regulating various hallmarks of cancer including that of GBM. The levels of many ncRNAs are dysregulated in cancer, and ectopic modulation of their levels by delivering antagonists or overexpression constructs could serve as an attractive option for cancer therapy. The therapeutic potential of several types of ncRNAs, including miRNAs, lncRNAs, and circRNAs, has been validated in both in vitro and in vivo models of GBM. However, the delivery of these RNA-based therapeutics is highly challenging, especially to the tumors of the brain as the blood-brain barrier (BBB) poses as a major obstacle, among others. Also, since RNA is extremely fragile in nature, careful considerations must be met while designing a delivery agent. In this review we have shed light on how ncRNA therapy can overcome the limitations of its predecessor conventional therapy with an emphasis on smart nanomaterials that can aide in the safe and targeted delivery of nucleic acids to treat GBM. Additionally, critical gaps that currently exist for successful transition from viral to non-viral vector delivery systems have been identified. Finally, we have provided a perspective on the future directions, potential pathways, and target areas for achieving rapid clinical translation of, RNA-based macromolecular therapy to advance the effective treatment of GBM and other related diseases.

Recombination drives the emergence of orf virus diversity: evidence from the first complete genome sequence of an Indian orf virus isolate and comparative genomic analysis.

Contagious pustular dermatitis is a disease that primarily infects small ruminants and possesses zoonotic potential. It is caused by orf virus (ORFV), a member of the genus Parapoxvirus. In this study, we evaluated an ORFV outbreak in goats in Madhya Pradesh, a state in central India, during 2017. The transboundary potential of this virus was evaluated by constructing phylogenetic trees. The complete genome sequence of an ORFV isolate named Ind/MP/17 was found to be 139,807 bp in length with 63.7% GC content and 132 open reading frames (ORFs) flanked by 3,910-bp inverted terminal repeats (ITRs). An investigation into evolutionary parameters such as selection pressure (θ = dN/dS) and nucleotide diversity (π) demonstrated that ORFV has undergone purifying selection. A total of 40 recombination events were identified, 21 of which were evident in the Ind/MP/17 genome, indicating its ability to generate new variants.

Genome-wide identification and characterization of microsatellite markers within the Avipoxviruses.

Microsatellite markers or Simple Sequence Repeats (SSRs) are gaining importance for molecular characterization of the virus as well as estimation of evolution patterns due to its high-polymorphic nature. The Avipoxvirus is the causative agent of pox-like lesions in more than 300 birds and one of the major diseases for the extinction of endangered avian species. Therefore, we conducted a genome-wide analysis to decipher the type, distribution pattern of 14 complete genomes derived from the Avipoxvirus genus. The in-silico screening deciphered the existence of 917-2632 SSRs per strain. In the case of compound SSRs (cSSRs), the value was obtained 44-255 per genome. Our analysis indicates that the di-nucleotide repeats (52.74%) are the most abundant, followed by the mononucleotides (34.79), trinucleotides (11.57%), tetranucleotides (0.64%), pentanucleotides (0.12%) and hexanucleotides (0.15%) repeats. The specific parameters like Relative Abundance (RA) and Relative Density (RD) of microsatellites ranged within 5.5-8.12 and 33.08-53.58 bp/kb. The analysis of RA and RD value of compound microsatellites resulted between 0.25-0.82 and 4.64-15.12 bp/kb. The analysis of motif composition of cSSR revealed that most of the compound microsatellites were made up of two microsatellites, with some unique duplicated pattern of the motif like, (TA)-x-(TA), (TCA)-x-(TCA), etc. and self-complementary motifs, such as (TA)-x-(AT). Finally, we validated forty sets of compound microsatellite markers through an in-vitro approach utilizing clinical specimens and mapping the sequencing products with the database through comparative genomics approaches. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03169-4.

Ultrasonic Atomizer-Driven Development of Biocompatible and Biodegradable Poly(d,l-lactide-co-glycolide) Nanocarrier-Encapsulated Suberoylanilide Hydroxamic Acid to Combat Brain Cancer.

The path to the discovery of anticancer drugs and investigating their potential activity has remained a quest for several decades. Suberoylanilide hydroxamic acid (SAHA), also known as "Vorinostat", is a well-known histone deacetylase inhibitor (HDACi) and has the potential to act as a therapeutic agent against tumorigenesis. Herein, we have fabricated SAHA incorporated into biocompatible and biodegradable poly(d,l-lactide-co-glycolide) PLGA nanoparticles (NPs) using a facile method of ultrasonic atomization and evaluated their anticancer property. We have explored their characteristics using dynamic light scattering (DLS), scanning electron microscopy (SEM), high-resolution transmission electron microscopy (HR-TEM), encapsulation efficiency, and in vitro drug release and have investigated their efficacy on U87 glioblastoma (GBM) cells. SAHA-PLGA NPs synthesized were of average mean size of 80 ± 23 and 105 ± 6.0 nm observed through cryo-field-emission gun SEM and HR-TEM with a polydispersity index of 0.068 and a ζ-potential value of -13.26 mV. The encapsulation efficiency was 53%, with a sustained in vitro release up to 48 h. The in vitro assessment of SAHA-PLGA NPs for their anticancer activity on U87 GBM cells showed cellular cytotoxicity with an IC50 of 19.91 µM. SAHA-PLGA NP-treated cells also showed suppression in migration with 8.77 µM concentration, and cell growth inhibition was observed in the wound scratch assay for up to 24 h. The cellular uptake studies have been utilized by time-dependent experiments, revealing their cellular internalization. Taking this into account, our present experimental findings indicate that SAHA-PLGA NPs could play a significant role in enhancing the effectiveness and bioavailability and reducing adverse effects of cancer chemotherapy. It also highlights the inherent potential of these biocompatible entities for chemotherapeutic applications in biomedical and pharmaceutics.

Comparative analysis, distribution, and characterization of microsatellites in Orf virus genome.

Genome-wide in-silico identification of microsatellites or simple sequence repeats (SSRs) in the Orf virus (ORFV), the causative agent of contagious ecthyma has been carried out to investigate the type, distribution and its potential role in the genome evolution. We have investigated eleven ORFV strains, which resulted in the presence of 1,036-1,181 microsatellites per strain. The further screening revealed the presence of 83-107 compound SSRs (cSSRs) per genome. Our analysis indicates the dinucleotide (76.9%) repeats to be the most abundant, followed by trinucleotide (17.7%), mononucleotide (4.9%), tetranucleotide (0.4%) and hexanucleotide (0.2%) repeats. The Relative Abundance (RA) and Relative Density (RD) of these SSRs varied between 7.6-8.4 and 53.0-59.5 bp/kb, respectively. While in the case of cSSRs, the RA and RD ranged from 0.6-0.8 and 12.1-17.0 bp/kb, respectively. Regression analysis of all parameters like the incident of SSRs, RA, and RD significantly correlated with the GC content. But in a case of genome size, except incident SSRs, all other parameters were non-significantly correlated. Nearly all cSSRs were composed of two microsatellites, which showed no biasedness to a particular motif. Motif duplication pattern, such as, (C)-x-(C), (TG)-x-(TG), (AT)-x-(AT), (TC)- x-(TC) and self-complementary motifs, such as (GC)-x-(CG), (TC)-x-(AG), (GT)-x-(CA) and (TC)-x-(AG) were observed in the cSSRs. Finally, in-silico polymorphism was assessed, followed by in-vitro validation using PCR analysis and sequencing. The thirteen polymorphic SSR markers developed in this study were further characterized by mapping with the sequence present in the database. The results of the present study indicate that these SSRs could be a useful tool for identification, analysis of genetic diversity, and understanding the evolutionary status of the virus.