Chitosan-Based Polymeric Nanoparticles as an Efficient Gene Delivery System to Cross Blood Brain Barrier: In Vitro and In Vivo Evaluations.

Significant progress has been made in the field of gene therapy, but effective treatments for brain tumors remain challenging due to their complex nature. Current treatment options have limitations, especially due to their inability to cross the blood-brain barrier (BBB) and precisely target cancer cells. Therefore options that are safer, more effective, and capable of specifically targeting cancer cells are urgently required as alternatives. This current study aimed to develop highly biocompatible natural biopolymeric chitosan nanoparticles (CNPs) as potential gene delivery vehicles that can cross the BBB and serve as gene or drug delivery vehicles for brain disease therapeutics. The efficiency of the CNPs was evaluated via in vitro transfection of Green Fluorescent Protein (GFP)-tagged plasmid in HEK293-293 and brain cancer MG-U87 cell lines, as well as within in vivo mouse models. The CNPs were prepared via a complex coacervation method, resulting in nanoparticles of approximately 260 nm in size. In vitro cytotoxicity analysis revealed that the CNPs had better cell viability (85%) in U87 cells compared to the chemical transfection reagent (CTR) (72%). Moreover, the transfection efficiency of the CNPs was also higher, as indicated by fluorescent emission microscopy (20.56% vs. 17.79%) and fluorescent-activated cell sorting (53% vs. 27%). In vivo assays using Balb/c mice revealed that the CNPs could efficiently cross the BBB, suggesting their potential as efficient gene delivery vehicles for targeted therapies against brain cancers as well as other brain diseases for which the efficient targeting of a therapeutic load to the brain cells has proven to be a real challenge.

Platelets' RNA as biomarker trove for differentiation of early-stage hepatocellular carcinoma from underlying cirrhotic nodules.

BACKGROUND & AIMS: Among the multiplicity of factors involved in rising incidence of hepatocellular carcinoma (HCC)-the second deadliest cancer, late diagnosis of early-stage HCC nodules originating from late-stage cirrhotic nodules is the most crucial. In recent years, Tumor-educated platelets (TEPs) have emerged as a strong multimodal tool to be used in liquid-biopsy of cancers because of changes in their mRNA content. This study assessed the reliability of selected mRNA repertoire of platelets as biomarkers to differentiate early HCC from late-stage cirrhotic nodules. METHODS: Quantitative real time PCR (qRT-PCR) was used to evaluate expression levels of selected platelets-specific mRNA between HCC patients compared to cirrhosis patients. ROC curve analysis assessed the sensitivity and specificity of the biomarkers. RESULTS: RhoA, CTNNB1 and SPINK1 showed a significant 3.3-, 3.2- and 3.18-folds upregulation, respectively, in HCC patients compared to cirrhosis patients while IFITM3 and SERPIND1 presented a 2.24-fold change. Strikingly, CD41+ platelets also demonstrated a marked difference of expression in HCC and cirrhosis groups. CONCLUSIONS: Our study reports liquid biopsy-based platelets mRNA signature for early diagnosis of HCC from underlying cirrhotic nodules. Moreover, differential expression of CD41+ platelets in two groups provides new insights into a probable link between CD41 expression on platelets with the progression of cirrhosis to HCC.

Anti-COVID-19 multi-epitope vaccine designs employing global viral genome sequences.

BACKGROUND: The coronavirus SARS-CoV-2 is a member of the Coronaviridae family that has caused a global public health emergency. Currently, there is no approved treatment or vaccine available against it. The current study aimed to cover the diversity of SARS-CoV-2 strains reported from all over the world and to design a broad-spectrum multi-epitope vaccine using an immunoinformatics approach. METHODS: For this purpose, all available complete genomes were retrieved from GISAID and NGDC followed by genome multiple alignments to develop a global consensus sequence to compare with the reference genome. Fortunately, comparative genomics and phylogeny revealed a significantly high level of conservation between the viral strains. All the Open Reading Frames (ORFs) of the reference sequence NC_045512.2 were subjected to epitope mapping using CTLpred and HLApred, respectively. The predicted CTL epitopes were then screened for antigenicity, immunogenicity and strong binding affinity with HLA superfamily alleles. HTL predicted epitopes were screened for antigenicity, interferon induction potential, overlapping B cell epitopes and strong HLA DR binding potential. The shortlisted epitopes were arranged into two multi-epitope sequences, Cov-I-Vac and Cov-II-Vac, and molecular docking was performed with Toll-Like Receptor 8 (TLR8). RESULTS: The designed multi-epitopes were found to be antigenic and non-allergenic. Both multi-epitopes were stable and predicted to be soluble in an Escherichia coli expression system. The molecular docking with TLR8 also demonstrated that they have a strong binding affinity and immunogenic potential. These in silico analyses suggest that the proposed multi-epitope vaccine can effectively evoke an immune response.

Novel association of genetic variants in non-coding regulatory regions with HIV-1 infection.

Host genetic variability interplays with the environment and variegating viral factors to determine the outcome in HIV-1/AIDS. Several GWAS studies have reported that genetic heterogeneity of individuals leads to differential HIV susceptibility. Proxy SNPs that are in Linkage Disequilibrium to the GWAS SNPs could be important targets in HIV pathogenesis and need to be analyzed further for their potential regulatory role. Current study thus aimed to identify novel proxy SNPs that may play a critical role in HIV susceptibility and disease progression. 372 SNPs, associated with HIV-1/AIDS pathogenesis, were retrieved via GWAS catalogue. 1854 proxy SNPs, in Linkage Disequilibrium (r2 = 0.8) to the GWAS reported SNPs, were identified using the SNAP web tool. Regulatory functions of aforementioned 1854 polymorphic sites (GWAS SNPs and their proxy SNPs) were acquired from RegulomeDB. 178 of the proxy SNPs showed evidence of strong regulatory potential returning a score of ≤3. Among these regulatory SNPs, 22 had already been reported for their association with HIV/AIDS while 156 SNPs showed novel association. Three of these novel SNPs (g.rs6457282T>C, g.rs17064977C>T and g.rs3130350G>T) were validated using sequence specific PCR (SSP-PCR) on HIV-infected patients. For g.rs6457282T>C and rs17064977C>T, CT genotype was determined to be significantly associated with increased risk of HIV-1 infection (rs6457282T>C: OR = 9.5, 95% CI = 3.0792-29.3099, p = 0.0001; rs17064977C>T: OR = 8.1077, 95% CI = 3.1125-21.119, p = 0.0001). Moreover, the association of interacting protein partners of affected genes with HIV-1 elucidates the significance of corresponding SNPs in HIV disease outcome that further needs to be functionally deciphered.

Tumor educated platelets, a promising source for early detection of hepatocellular carcinoma: Liquid biopsy an alternative approach to tissue biopsy.

PURPOSE: Liver cancer is considered to be the sixth deadliest cancer worldwide. Hepatocellular carcinoma (HCC) is known to be the most prevalent type of liver cancer. The number of deaths due to HCC reported per year is on a constant rise especially in lesser developed countries. There are several contributing factors to this rise in number. Among other contributing factors is the late diagnosis of HCC. Patients are usually diagnosed when the disease reaches its advance stage. The present study was conducted with total 30 samples. It was designed for investigating the potential of TGF-ß, NF-κß, VEGF, AKT and PI3K as RNA based biomarkers in tumor educated platelets for early detection of HCC. RESULTS: The results obtained from the transcriptional analysis revealed a significant high expression of TGF-ß, NF-κß, VEGF by 2.48, 2.35 and 2.78 folds respectively in comparison to the control. On the other hand, a decrease in expression by 0.6 and 0.65 folds was observed in AKT and PI3K respectively in comparison to controls. Although all selected RNA biomarkers showed promising potential to detect HCC however, AKT and PI3K were better able to detect early stage HCC. CONCLUSIONS: The results obtained clearly indicate the increased expression of TGF-ß, NF-κß, VEGF in HCC patients. All these biomarkers are previously known for cancer initiation, progression and metastasis. The significant decrease in expression of AKT and PI3K in HCC patients needs further investigation. All the selected RNA biomarkers can be used for detection of HCC as they were able to distinguish HCC patients from controls successfully with AKT and PI3K showing better potential to detect early stage HCC. However, translational analysis for all these RNA biomarkers should be performed to gain further evidence for the ability of these biomarkers to be used for early HCC detection.