CHEMICAL APPROACHES TOWARD NEW PERSPECTIVES IN DRUG DISCOVERY: NATURAL PRODUCTS AND BIOACTIVE HITS AS AN INSPIRATION

The quest for novel therapeutics in drug discovery requires a multifaced approach, and innovative strategies that bridge the gap between nature’s diverse chemical landscape and the evolving demands of modern medicine. By employing a synergy between total synthesis, innovative molecule design and targeted modifications, this Ph.D. thesis aims to provide different approaches to the conceptualization and development of hypothetical therapeutic agents. Part I of this work delves into the invaluable role of natural products as intrinsically interesting compounds and as a foundation for drug discovery. Natural products have been a cornerstone in the development of therapeutics for millennia, with their roots deeply embedded in traditional medicine. Even in today’s pharmaceutical landscape, the influence of natural products is profound; they serve as a rich source of chemical diversity and biological potential that has yielded a significant fraction of modern drugs. A striking example is the World Health Organization's recognition that approximately 80% of the global population relies on traditional medicine, exploiting natural products in various forms. Within this context, it is critical to analyze the structural complexity and unique pharmacological properties that render these compounds essential in therapeutic applications, particularly in the realms of oncology and infectious diseases. At the forefront of this exploration is the use of natural products in bivalent compound assemblies, particularly the emerging proteolysis-targeting chimeras (PROTACs), to inspire the design of innovative molecular architectures that can selectively degrade target proteins. The strategic incorporation of bioactive natural compounds into these bivalent frameworks not only enhances target specificity, but also broadens the scope of druggable targets in the proteome. Chapter 1 investigates the potential of four novel putative PROTAC compounds designed to target the E3 ubiquitin ligase cereblon and neuroserpin (NS) via a recruiting motif derived from naturally occurring embelin. Indeed, the accumulation of mutant NS polymers within the endoplasmic reticulum (ER) is responsible of the neurodegenerative and incurable Familial Encephalopathy with Neuroserpin Inclusion Bodies (FENIB). The goal is to facilitate the ER-associated degradation (ERAD) of mutant NS and promote its degradation via the proteasome, in an urgent need for effective therapeutic strategies. Chapter 2 details the rational design and development of PROTACs targeting tubulin, a crucial dimeric protein involved in various cellular functions, particularly in cancer and neurodegenerative therapies. Maytansinol, a known tubulin binder, was chosen as a ligand for tubulin, which was then linked to either cereblon or Von Hippel-Lindau ligands as E3 ligase recruiters. Comprehensive biophysical, biological, and structural characterization of four synthesized PROTAC compounds demonstrated their ability to form the desired ternary complexes with tubulin and E3 ligases. This chapter emphasizes a structured and rational approach to PROTAC development, integrating structural insights and cellular studies to enhance drug discovery efforts. The exploration then transitions to challenges associated with the chemical synthesis of natural products. While the natural world provides a treasure trove of bioactive compounds, access to these resources is often hampered by availability, extraction difficulties, and scalability issues. Overcoming these barriers has given rise to a thriving field of synthetic organic chemistry dedicated to the total synthesis of complex natural products. These synthetic endeavors not only aim to replicate the structures of natural compounds, but also seek to enhance their properties, paving the way for novel, nature-inspired therapeutic agents. Chapter 3 deals with the biomedical potential of Cannabis, particularly its lesser-studied cannabinoids. The project focuses on the synthesis of minor cannabinoids with three or four carbon atom lateral chains, specifically targeting cannabigerol (CBG) and cannabichromene (CBC) analogues. The anti-inflammatory properties of these compounds were tested after their synthesis, identifying five among them as effective inhibitors of skin inflammation mediators; notably, the length of their side chain was critical for activity. This work highlights the potential of Cannabis components in addressing inflammatory skin conditions. Chapter 4 focuses on the efforts towards the total synthesis of glycybridin B, a natural compound whose synthesis has never been reported, and that has shown promise as a potential tubulin binder. This fits to the search for new microtubule targeting agents, particularly looking for simpler, more easily modifiable compounds compared to the complex, currently known natural products, and paves the way for further exploration of its putative binding properties and potential therapeutic applications. Part II is oriented towards the rational chemical modification of well-characterized bioactive hits, offering additional pathways to drug refinement and optimization. This part investigates the innovative design of compounds that enhance brain bioavailability while maintaining therapeutic efficacy, facing the challenges of blood-brain barrier (BBB) permeation in central nervous system (CNS) drug development. By employing chemical modifications, such as incorporating guanyl hydrazone functionalities into bioactive molecules, this research aims to unlock new avenues for CNS drug delivery. Chapter 5 is about small molecule targeting RNA, in particular expanded G4C2 repeats, resulting from mutations in the C9orf72 gene, as contributors to the pathologies of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). The chapter highlights the modification of a hit bisthiophene-based compound, which preferentially stabilizes G-quadruplex RNA structures associated with the G4C2 repeats, by substituting its amidine groups with BBB-compliant guanyl hydrazones, and creating a series of new compounds based on diverse scaffold variations. An eight-membered guanyl hydrazone array was biologically tested, focusing on their potential as binders to the C9orf72 RNA. Chapter 6 delves into the role of Acid-Sensing Ion Channels (ASICs), specifically the ASIC3 isoform, which has been found in Glioblastoma Multiforme (GBM) cancer stem cells (CSCs). The chapter proposes a novel therapeutic approach targeting GBM CSCs through sustained activation of ASIC3, with small organic molecules. Starting from known 2-guanidine-4-methylquinazoline (GMQ), identified as an ASIC3 activator, the enhancement of GMQ's bioavailability by synthesizing novel analogues is described, specifically replacing its 2-guanidino group with a guanyl hydrazone and expanding chemical diversity using computational guidance. In conclusion, this Ph.D. thesis endeavors to present a comprehensive narrative that emphasizes a synergy between natural products, chemical synthesis and drug design. By elucidating the potential of these strategies, this work aims to inspire future research directions that harness the wealth of natural compounds while overcoming the barriers inherent in drug development, ultimately enhancing our ability to address unmet medical needs.