Debajyoti Chakraborty

Lipid nanoparticles (LNPs) have emerged as a revolutionary nonviral platform for administering a wide range of therapeutics, especially nucleic acids. Expanding the diversity of lipid structures, which includes a variety of headgroups, linkers, and tail structures, offers exciting possibilities for developing novel lipid-based drug delivery systems. Consequently, the scientific community laid a greater emphasis on combinatorial methodologies for designing different libraries of ionizable lipid molecules. Herein, we are proposing the synthesis of a library of triazole-based ionizable lipids by using alkyne-azide click chemistry. Systematic variation of the terminal headgroup produced distinct mRNA expression profiles despite broadly comparable encapsulation efficiencies and physicochemical properties. LNP formulated using CL-3, containing primary amine headgroup, exhibited the highest in vitro mRNA expression and outperformed the benchmark ionizable lipids ALC-0315 and SM-102 under the conditions tested. CL-3-based LNPs elicited the highest binding-antibody titers among the evaluated formulations, while pseudovirus-neutralizing activity was comparable to that of the ALC-0315 benchmark. The immunized animals were protected in the mouse viral-challenge model under the conditions investigated. These findings identify CL-3 as a promising candidate within the evaluated lipid series and demonstrate the utility of modular synthesis for investigating structure–activity trends in mRNA delivery.

While inflammasomes detect and defend against pathogens, the precise mechanisms by which enteric viruses trigger these complexes, and the specific viral components involved remain incompletely understood. In this study, we demonstrate that rotavirus infection robustly induced human NLRP1 inflammasome activation via p38 mitogen-activated protein kinase (MAPK)-dependent and -independent pathways. Utilizing cell lines reconstituted with NLRP1 signaling and reverse genetics-rescued recombinant rotaviruses, we identified non-structural protein 1 (NSP1), a well-characterized viral antagonist of host interferon signaling, as an activator of the NLRP1 inflammasome in the absence of double-stranded RNA. Domain mapping analysis revealed that the C-terminus of NSP1 but not the C-terminal IRF3-binding site was essential for triggering inflammasome activation. Mechanistically, we show that NSP1 induced the proteasomal degradation of NLRP1 to elicit its activation, a process dependent on host neddylation pathways and the Cullin-3/Elongin B/C ubiquitin ligase complex. Together, these findings suggest that RV engages parallel NLRP1 surveillance pathways, including a p38-dependent response to infection, and a novel NSP1-dependent pathway involving the ubiquitin-proteosome system, where a viral strategy aimed at host protein degradation inadvertently trips the human NLRP1 inflammasome wire. SIGNIFICANCE STATEMENT: When rotavirus infects the gut, it uses a viral protein called NSP1 to shut down key immune defenses of host cells. However, human cells have an immune sensor called NLRP1 that acts as a tripwire. When NSP1 attempts to force the destruction of NLRP1, the disposal process itself accidentally triggers the NLRP1 alarm. This causes an explosive inflammatory cell death that alerts the immune system. This discovery reveals how a viral strategy to suppress host immunity backfires, offering new insights into viral gut inflammation and future vaccine design.

Alpha-hemolysin (α-HL) is a small pore-forming toxin secreted by pathogenic Staphylococcus aureus, inducing cell death process by forming pores in membrane. So far, detergents or artificial lipid environments have been utilized to characterize the toxin structure. The toxin-induced changes in the membrane, membrane remodeling after toxin treatment, and the role of the toxin during pore formation process remain ambiguous. Thus, understanding pore formation in the cellular environment, including the roles of the plasma membrane and lipid composition, is crucial for drug development. In this study, we captured step-by-step oligomerization of α-HL and membrane rupture of erythrocyte cells using confocal microscopy, cryo-EM imaging, and single-particle analysis. We resolved 3.1–3.8 Å resolution structures of pore, prepore, and immature pore conformations in cellular environment. Furthermore, mass spectrometry analysis demonstrated key erythrocyte lipid components interacting with α-HL. Our findings indicate that shorter or unsaturated lipid chains facilitate pore formation and the role of phosphatidylcholine with varying physical properties in modulating the toxin’s activity.

With the rapid emergence of variants of concern (VOC), the efficacy of currently licensed vaccines has reduced drastically. VOC mutations largely occur in the S1 subunit of Spike. The S2 subunit of SARS-CoV-2 is conserved and thus more likely to elicit broadly reactive immune responses that could improve protection. However, the contribution of the S2 subunit in improving the overall efficacy of vaccines remains unclear. Therefore, we designed, and evaluated the immunogenicity and protective potential of a stabilized SARS-CoV-2 Receptor Binding Domain (RBD) fused to a stabilized S2. Immunogens were expressed as soluble proteins with approximately fivefold higher purified yield than the Spike ectodomain and formulated along with Squalene-in-water emulsion (SWE) adjuvant. Immunization with S2 alone failed to elicit a neutralizing immune response, but significantly reduced lung viral titers in mice challenged with the heterologous Beta variant. In hamsters, SWE-formulated RS2 (a genetic fusion of stabilized RBD with S2) showed enhanced immunogenicity and efficacy relative to corresponding RBD and Spike formulations. Despite being based on the ancestral Wuhan strain of SARS-CoV-2, RS2 elicited broad neutralization, including against Omicron variants (BA.1, BA.5 and BF.7), and the clade 1a WIV-1 and SARS-CoV-1 strains. RS2 elicited sera showed enhanced competition with both S2 directed and RBD Class 4 directed broadly neutralizing antibodies, relative to RBD and Spike elicited sera. When lyophilized, RS2 retained antigenicity and immunogenicity even after incubation at 37 °C for a month. The data collectively suggest that the RS2 immunogen is a promising modality to combat SARS-CoV-2 variants.

Breast cancer is a leading cause of cancer-related deaths, but current programs are expensive and prone to false positives, leading to unnecessary follow-up and patient anxiety. This paper proposes a solution to automated breast cancer detection, to improve the efficiency and accuracy of screening programs. Different methodologies were tested against the RSNA dataset of radiographic breast images of roughly 20,000 female patients and yielded an average validation case pF1 score of 0.56 across methods.

We leverage the fast physics simulator, MuJoCo to run tasks in a continuous control environment and reveal details like the observation space, action space, rewards, etc. for each task. We benchmark value-based methods for continuous control by comparing Q-learning and SARSA through a discretization approach, and using them as baselines, progressively moving into one of the state-of-the-art deep policy gradient method DDPG. Over a large number of episodes, Qlearning outscored SARSA, but DDPG outperformed both in a small number of episodes. Lastly, we also fine-tuned the model hyper-parameters expecting to squeeze more performance but using lesser time and resources. We anticipated that the new design for DDPG would vastly improve performance, yet after only a few episodes, we were able to achieve decent average rewards. We expect to improve the performance provided adequate time and computational resources.