#10 Drug design VI: allosteric drugs, chaperones, PPIs, induced proximity-based therapeutics

Introduction and Class Announcements

  • Mentioned the availability of practice questions on the quiz in the practice section.

  • Importance of practice:

    • Students encouraged to sit down and answer questions actively.

    • Reviewing questions and answers passively does not foster problem-solving skills.

  • Monthly review sessions will be held, and students are encouraged to ask questions at any time.

Overview of the Lecture

  • Focus on the different ways that structured molecules can interfere with disease pathways.

  • Classic methods of interference include:

    • Inhibition of enzymes.

    • Allosteric or orthosteric ligands (antagonists or agonists).

  • Importance of understanding disease mechanisms for effective drug development.

  • Emphasis on creativity in research and drug design.

  • Objective: Learn various drug mechanisms and their associated challenges.

  • Discussion of drug examples that have revolutionized treatment.

Types of Drug Mechanisms

  • Allosteric Modulators:

    • Principle: Molecules that bind to a site other than the active site (an allosteric site) and influence protein activity by inducing a conformational change. These allosteric sites are often distinct from the evolutionarily conserved active sites, offering greater opportunities for developing selective drugs with fewer off-target effects and improved drug specificity.

    • Finding selective allosteric sites can be highly beneficial as they often differ significantly between protein classes or even between closely related proteins.

    • Examples:

      • BCR-ABL Kinase Inhibition:

        • Mutations (e.g., in chronic myeloid leukemia) can create a form of BCR-ABL kinase that is constitutively active, driving uncontrolled cell proliferation.

        • Allosteric inhibitors (e.g., imatinib, nilotinib) are designed to fit into altered binding pockets created by these mutations or to bind to a site distinct from the ATP-binding pocket. By binding to these allosteric sites, they stabilize an inactive conformation of the kinase, thereby reducing its activity.

      • GPCR (G Protein-Coupled Receptor) Modulators: GPCRs are a major class of drug targets, and allosteric modulators can bind to sites distinct from the orthosteric ligand-binding site to modify the receptor's response. They can enhance (positive allosteric modulators, PAMs), diminish (negative allosteric modulators, NAMs), or otherwise alter the receptor's response to its natural ligand, offering fine-tuned control over signaling pathways.

      • Non-Nucleoside Reverse Transcriptase Inhibitors (NNRTIs): These are classic examples of allosteric drugs used in HIV treatment. They bind to a non-active site pocket on HIV reverse transcriptase, inducing a conformational change that inhibits its enzymatic activity necessary for viral replication.

  • Chaperones:

    • Principle: Small molecules that assist in protein folding and stabilization, promoting proper protein conformation, preventing misfolding and aggregation, and ensuring correct cellular trafficking to enable functional activity.

    • Importance: Particularly critical in genetic diseases where specific mutations lead to protein misfolding, aggregation, or premature degradation, causing cellular dysfunction.

    • Example:

      • Cystic Fibrosis (CFTR):

        • CFTR Mutation: Mutations in the CFTR (Cystic Fibrosis Transmembrane conductance Regulator) gene, most commonly the F508del mutation, cause improper folding of the CFTR protein. This misfolded protein is then recognized by the cell's quality control mechanisms and targeted for degradation before it can reach the cell surface, resulting in a lack of functional chloride channels and severe respiratory, digestive, and other systemic issues.

        • CFTR Correctors: Drugs specifically designed to stabilize the misfolded CFTR protein, allowing it to bypass cellular quality control, traffic properly to the cell membrane, and integrate as a functional chloride channel.

        • CFTR Potentiators: Drugs that bind to the CFTR channel at the cell surface and increase its opening probability and chloride-ion conductance, thereby maximizing the function of any correctly trafficked CFTR protein.

        • Vertex Pharmaceuticals and TRIKAFTA (ivacaftor, tezacaftor, elexacaftor): Vertex Pharmaceuticals developed combinatory therapies that have revolutionized CF treatment. TRIKAFTA is a triple combination therapy specifically developed to target different defects of the CFTR protein, particularly for patients with the F508del mutation. It combines two correctors (tezacaftor and elexacaftor), which work synergistically to help the misfolded CFTR protein move to the cell surface, and a potentiator (ivacaftor), which helps open the channel once it's expressed on the cell surface. This multi-pronged approach has made significant improvements in patient quality of life by restoring substantial CFTR function.

  • Protein-Protein Interaction (PPI) Inhibitors:

    • The Interactions: PPIs are fundamental to almost all biological processes, and their dysregulation is implicated in numerous diseases, making them crucial drug targets. However, effective inhibition of the large, often flat, and dynamic interaction surfaces of proteins involved in PPIs is challenging for traditional small molecules, which typically bind to small, well-defined pockets.

    • Examples discussed include:

      • Inhibitors that prevent protein interactions (e.g., targeting the interaction between MDMX and p53): The p53 protein is a critical tumor suppressor, often referred to as the "guardian of the genome." MDMX (also known as MDM4) is one of its negative regulators, binding to p53 and promoting its degradation or inhibiting its transcriptional activity. Inhibiting the MDMX-p53 interaction can restore p53's tumor-suppressing activity, making it an attractive strategy in cancer therapy.

      • The PCSK9 (Proprotein Convertase Subtilisin/Kexin type 9) Inhibitor:

        • Mechanism: PCSK9 binds to LDL receptors (LDLRs) on the cell surface and targets them for lysosomal degradation, thereby reducing the number of LDLRs available to clear LDL cholesterol from the blood. This leads to elevated LDL-C levels, a major risk factor for cardiovascular disease.

        • Inhibitors: Drugs designed to inhibit PCSK9 (e.g., monoclonal antibodies like evolocumab, alirocumab, or experimental small molecules) block the interaction between PCSK9 and the LDLR. This prevention leads to more LDLRs on the cell surface, increased LDL-C clearance from the bloodstream, and consequently, lower blood cholesterol levels, providing a significant therapeutic advance for hypercholesterolemia.

  • Induced Proximity-Based Therapeutics:

    • Definition: Innovative strategies that bring two proteins together to facilitate a new interaction or modify an existing one, leading to a desired biological outcome that naturally does not occur or is inefficient. This often involves a small molecule acting as a 'bridge' or 'molecular glue'.

    • Historical Example: Thalidomide:

      • Initially used for morning sickness, but its severe teratogenic effects led to its withdrawal. It was later re-purposed for oncology and immunology due to its immunomodulatory properties.

      • Thalidomide as a Molecular Glue: It was discovered that thalidomide acts as a "molecular glue" by binding to the E3 ubiquitin ligase cereblon (CRBN) and simultaneously presenting certain target proteins (like Ikaros and Aiolos, and importantly, SALL4) to CRBN. This induced proximity leads to the ubiquitination of these target proteins, marking them for degradation by the proteasome. Its initial teratogenic effects were later understood to be due to its ability to induce the degradation of SALL4, a transcription factor crucial for limb development.

    • Protein Ubiquitination: Ubiquitination is a post-translational modification where a ubiquitin protein is attached to a substrate protein. This process, facilitated by E1, E2, and E3 enzymes, often marks proteins for degradation by the 26S proteasome. Induced proximity therapeutics leverage this natural cellular degradation pathway.

    • Molecular Glues:

      • Definition: These are small molecules that bind to two different proteins (or protein domains) that do not ordinarily interact, bringing them into close proximity to facilitate a new functional interaction, often leading to protein degradation.

      • Mechanism: They typically bind to both an E3 ubiquitin ligase (e.g., CRBN) and a target protein, inducing ubiquitination of the target, which marks it for degradation by the proteasome.

      • Advantages over traditional inhibitors: While a traditional inhibitor only blocks one active site, a single molecular glue molecule can recruit an E3 ligase to ubiquitinate and thus clear multiple target protein molecules, potentially leading to more sustained and complete target suppression.

      • Therapeutics targeting oncogenic proteins and disease pathways are actively being explored in clinical trials.

    • PROTACs (PROteolysis TArgeting Chimeras):

      • Definition: PROTACs are bivalent small molecules engineered to induce the degradation of specific target proteins. Unlike molecular glues, PROTACs are designed with two distinct binding motifs: one that binds to an E3 ubiquitin ligase (e.g., VHL, CRBN) and another that binds to the target protein of interest.

      • Mechanism: By simultaneously binding to both the E3 ligase and the target protein, PROTACs create an artificial ternary complex, leading to the ubiquitination of the target protein. This ubiquitinated protein is then recognized and degraded by the 26S proteasome.

      • Advantages: PROTACs offer potential advantages over traditional inhibitors, including catalytic degradation (one PROTAC molecule can induce the degradation of multiple target proteins), the ability to target scaffolding and non-enzymatic proteins (which are often considered "undruggable" by traditional inhibitors), and potential for overcoming drug resistance mechanisms.

    • KRAS Mutant and Pan-RAS Inhibitors, Darovasertib:

      • KRAS: The KRAS oncogene is a highly prevalent driver of various cancers (e.g., pancreatic, colorectal, lung), and historically, it has been considered "undruggable" due to its smooth surface and lack of obvious binding pockets. Induced proximity strategies (including PROTACs and molecular glues) are actively being developed for degrading mutant KRAS proteins or achieving pan-RAS inhibition.

      • First-in-class KRAS G12C inhibitors (e.g., sotorasib, adagrasib) work by covalently binding to a mutant cysteine residue, locking KRAS in an inactive GDP-bound state. While these are not directly induced proximity degraders, they represent innovative approaches beyond traditional competitive inhibition, paving the way for further advanced modalities.

      • Darovasertib (formerly darasertib): This compound is a protein kinase C (PKC) inhibitor currently in clinical trials for solid tumors like uveal melanoma. Its mechanism does not directly involve induced proximity for degradation but rather competitive inhibition of an enzyme, showcasing another modern approach in targeted therapy.

    • Calcineurin Inhibitors (Induced Proximity Mechanism of Action):

      • Mechanism: Immunosuppressants like tacrolimus and cyclosporine are classic examples of induced proximity. These drugs do not directly inhibit calcineurin. Instead, they first bind to small intracellular proteins known as immunophilins (e.g., tacrolimus binds to FKBP12, cyclosporine binds to cyclophilin).

      • The resulting drug-immunophilin complex (e.g., Tacrolimus-FKBP12) then binds to and inhibits calcineurin. Thus, the drug acts as an "inducer" to bring the immunophilin into proximity with calcineurin, forming a new complex that blocks calcineurin's phosphatase activity, which is essential for T-cell activation and immune response. This prevents organ rejection in transplant patients.

    • Lenacapavir (Induced Proximity-Based Therapeutics):

      • Mechanism: Lenacapavir is a first-in-class HIV-1 capsid inhibitor approved for the treatment of multi-drug resistant HIV-1. It acts via an induced proximity mechanism by binding to the HIV-1 capsid protein and promoting its premature and incorrect assembly and disassembly pathways. This leads to the aggregation of capsid proteins and disruption of key steps in the viral life cycle, including uncoating, reverse transcription, and nuclear import of the viral preintegration complex. By inducing the mis-assembly or aggregation of viral components, it effectively halts viral replication, representing a novel application of induced proximity principles.

Conclusion

  • Summary of the lecture topics:

    • The exploration of various pathways for drug action, including allosteric modulation, chaperone-mediated folding, protein-protein interaction inhibition, and induced proximity-based therapeutics, demonstrates a broad creative potential in scientific research.

  • Anticipation for a recap session in the following week.

  • Encouragement for continued inquiry and creativity in drug development.

  • Closing remarks on the advancements in drug therapies and their potential to impact patient care positively.