Nitin Aniruddha Patil

Alzheimer's disease (AD) is the most prevalent neurodegenerative disease characterized by memory loss and other cognitive functions. The key hallmarks of AD include extracellular beta-amyloid clumps and intracellular neurofibrillary tau tangles in the neurons. Cholinesterase inhibitors and NMDA-receptor antagonists and their combination are already approved treatments; however, these only give short-term symptom relief. Therefore, new therapeutic techniques and novel drugs are required to combat the century-old AD. This study includes the screening of nine novel small compounds (spiro-indenoquinoxaline-pyrrolidines) via in silico approaches; these compounds have been scrutinized to explore their potential as antiamyloidogenic drugs. Computational tools, including ADMET analysis, molecular docking, and molecular dynamics (MD) simulations, have been used for screening the selected compounds against monomeric peptides of Aβ (Aβ_1-40 and Aβ_1-42) and their oligomeric counterparts, i.e., 6Aβ_9-40 and 6Aβ_1-42. Among the nine molecules screened for this study, ADPR-d reflected the best drug-likeness and negligible toxicity. Further, ADPR-d has the highest binding affinity for all the peptides selected for this study. Additionally, MD simulations of Aβ peptide-ADPR-d complexes confirmed a stable complex formation. In vitro aggregation assay and cell culture studies for Aβ_1-42 also support our in silico findings. The positive findings of the presented study highlight that the ADPR-d molecule may prove to be a potential therapeutic molecule against AD. However, these results would require further in vitro and in vivo analysis before proceeding to clinical settings with these compounds against AD.

Multidrug-resistant Gram-negative bacteria have caused a serious threat to global health, and polymyxins are an important last-line therapy. As resistance to polymyxins is emerging, understanding the structure–activity relationship (SAR) of the polymyxins can facilitate the discovery of novel antimicrobial lipopeptides with improved antibacterial activity. However, l-2,4-diaminobutyric acid (Dab) is a key amino acid for the antibacterial activity of polymyxins, and the SAR of five Dab residues has not been well studied. Here, we employed an all-atom molecular dynamics simulation approach by integrating a lipidomics-informed outer membrane (OM)-based model and systematically investigated the SAR of the five Dab residues, specifically the length of their side chains. The impact of the length of the Dab side chain on three activity-related aspects, namely, the conformation of polymyxins, OM penetration ability, and membrane deformation, was systematically examined at the atomic level and compared with in vitro antimicrobial activity results. We uncovered that altering the side chain length of the Dab residues at different positions significantly affected the antibacterial activity via distinct mechanisms. Longer side chains of Dab residues in the linear tripeptide segment (Dab¹ and Dab³) and shorter side chains in the heptapeptide ring (Dab⁵, Dab⁸, and Dab⁹) resulted in marked changes in OM deformation. Importantly, polymyxin activity is governed by the interplay of multiple structural and functional factors. Our mechanism-based SAR model predicted how these position-specific modifications in Dab side chains modulate polymyxin activity, as supported by experimental validation. Specifically, elongation of the Dab¹ and Dab⁵ side chains led to significantly reduced antibacterial activity, while shortening of the Dab³ side chain enhanced activity. Collectively, the atomic-level SAR of polymyxins centered on the Dab residues will help expedite the rational design of new-generation antimicrobial lipopeptides, and our transferable framework provides broad implications for advancing membrane-active therapeutic agents.

A significant increase in life-threatening infections caused by Gram-negative "superbugs" is a serious threat to global health. With a dearth of new antibiotics in the developmental pipeline, antibiotics with novel mechanisms of action are urgently required to prevent a return to the preantibiotic era. A key strategy to develop novel anti-infective treatments is to discover new natural scaffolds with distinct mechanisms of action. Laterocidine is a unique cyclic lipodepsipeptide with activity against multiple problematic multidrug-resistant Gram-negative pathogens, including Pseudomonas aeruginosa, Acinetobacter baumannii, and Enterobacterales. Here, we developed a total chemical synthesis methodology for laterocidine and undertook systematic structure-activity relationship studies with chemical biology and NMR. We discovered important structural features that drive the antimicrobial activity of laterocidine, leading to the discovery of an engineered peptide surpassing the efficacy of the original peptide. This engineered peptide demonstrated complete inhibition of the growth of a polymyxin-resistant strain of Pseudomonas aeruginosa in static time-kill experiments.

Nitrile‐aminothiol conjugation (NATC) stands out as a promising biocompatible ligation technique due to its high chemo‐selectivity. Herein we investigated the reactivity and substrate scope of NAT conjugation chemistry, thus developing a novel pH dependent orthogonal NATC as a valuable tool for chemical biology. The study of reaction kinetics elucidated that the combination of heteroaromatic nitrile and aminothiol groups led to the formation of an optimal bioorthogonal pairing, which is pH dependent. This pairing system was effectively utilized for sequential and dual conjugation. Subsequently, these rapid (≈1 h) and high yield (>90 %) conjugation strategies were successfully applied to a broad range of complex biomolecules, including oligonucleotides, chelates, small molecules and peptides. The effectiveness of this conjugation chemistry was demonstrated by synthesizing a fluorescently labelled antimicrobial peptide‐oligonucleotide complex as a dual conjugate to imaging in live cells. This first‐of‐its‐kind sequential NATC approach unveils unprecedented opportunities in modern chemical biology, showcasing exceptional adaptability in rapidly creating structurally complex bioconjugates. Furthermore, the results highlight its potential for versatile applications across fundamental and translational biomedical research.

Multidrug-resistant Gram-negative bacteria present an urgent and formidable threat to the global public health. Polymyxins have emerged as a last-resort therapy against these 'superbugs'; however, their efficacy against pulmonary infection is poor. In this study, we integrated chemical biology and molecular dynamics simulations to examine how the alveolar lung surfactant significantly reduces polymyxin antibacterial activity. We discovered that lung surfactant is a phospholipid-based permeability barrier against polymyxins, compromising their efficacy against target bacteria. Next, we unraveled the structure-interaction relationship between polymyxins and lung surfactant, elucidating the thermodynamics that govern the penetration of polymyxins through this critical surfactant layer. Moreover, we developed a novel analog, FADDI-235, which exhibited potent activity against Gram-negative bacteria, both in the presence and absence of lung surfactant. These findings shed new light on the sequestration mechanism of lung surfactant on polymyxins and importantly pave the way for the rational design of new-generation lipopeptide antibiotics to effectively treat Gram-negative bacterial pneumonia.

Polymyxins (polymyxin B and colistin) are lipopeptide antibiotics used as a last-line treatment for life-threatening multidrug-resistant (MDR) Gram-negative bacterial infections. Unfortunately, their clinical use has been affected by dose-limiting toxicity and increasing resistance. Structure-activity (SAR) and structure-toxicity (STR) relationships are paramount for the development of safer polymyxins, albeit very little is known about the role of the conserved position 10 threonine (Thr) residue in the polymyxin core scaffold. Here, we synthesized 30 novel analogues of polymyxin B₁ modified explicitly at position 10 and examined the antimicrobial activity against Gram-negative bacteria and in vivo toxicity and performed molecular dynamics simulations with bacterial outer membranes. For the first time, this study revealed the stereochemical requirements and role of the β-hydroxy side chain in promoting the correctly folded conformation of the polymyxin that drives outer membrane penetration and antibacterial activity. These findings provide essential information for developing safer and more efficacious new-generation polymyxin antibiotics.