Ayaan Agarwal, Inquirer, Creator, Advocate

Multiple sclerosis (MS) is an autoimmune, neurodegenerative disease where the immune system damages myelin sheaths in the central nervous system. MS pathogenesis was attributed to auto-reactive T cells. However, recent studies suggest that B cells also play a vital role, including antibody-dependent and independent effects in compartmentalized inflammation. Clonotypes are groups of B cells with the same receptor.

To explore the role of B cell clonotypes in MS pathogenesis, we conducted a meta-analysis of raw high-throughput sequencing data of the B cell repertoire of 82 subjects (48 MS patients and 34 controls). A novel combination of density-based clustering (DBSCAN) and supervised ML (random forest) was applied to cluster clonotypes into unique clonotype groups (UCG), and their disease relevance was assessed. Random forest feature importances, Mann-Whitney tests, and empirical power estimation revealed six significant, potentially pathogenic UCGs.

Additionally, tertiary analyses revealed genetic variability and clonal expansion as hallmarks of MS-associated B cells. Interestingly, new genomic motifs among our potentially pathogenic UCGs aligned with known IGHV4 family biases. Overall, the study highlights the utility of machine learning analysis in decoding immune repertoires and contributes to our understanding of B-cell involvement in MS pathogenesis, paving the way for future therapeutic exploration.

Alternative lengthening of telomeres (ALT) is a telomerase-independent telomere maintenance pathway used by approximately 5-10% of all cancers (Mori et al., 2024). ALT-positive cancers rely on recombination-based telomere extension. Therefore, recombination-associated proteins could act as therapeutic vulnerabilities. The PSMC3IP-MND1 protein heterodimer is involved in homologous pairing during ALT recombination (Cho et al., 2014). However, whether PSMC3IP represents a targetable functional dependency in ALT-positive cancers is unknown.

Here, we developed a CRISPR interference-based genetic engineering tool for future studies investigating the effect of PSMC3IP silencing on ALT-positive cancer viability under ionizing radiation. A PSMC3IP-targeting guide RNA sequence was assessed in silico, cloned into an expression vector, transformed into bacterial cells, purified, and confirmed by Sanger sequencing to match the expected sequence with no discrepancies in the sequenced region. This verified construct was used for lentiviral preparation in HEK293T cells. HEK293T cells exhibited morphological changes that provided supportive evidence that the cells responded to the transfection process.

Overall, this PSMC3IP-targeting CRISPRi sgRNA construct provides a verified genetic tool for future experiments designed to test whether PSMC3IP can be silenced and whether such silencing affects ALT-positive cancer cell viability or sensitivity to standard-of-care ionizing radiation.

RNA-based therapeutics, including small interfering RNA (siRNA), microRNA (miRNA), and antisense oligonucleotides (ASOs), have rapidly emerged as promising approaches for treating hereditary diseases, cancers, and other conditions that have historically been difficult to manage.

This research paper explores the potential of RNA therapeutics in the treatment of neurodegenerative diseases, including Parkinson's disease, Batten disease, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), Huntington's disease, spinal muscular atrophy (SMA), and Creutzfeldt–Jakob disease. We examine existing RNA-based therapeutics and their mechanisms of action, discuss the biological mechanisms underlying these neurodegenerative diseases, and focus on how RNA therapeutic tools could be applied to Parkinson's disease, Alzheimer's disease, and SMA by matching their underlying molecular mechanisms with one or more appropriate therapeutic strategies.