Drug Discovery Process for CML

Drug Discovery Process

• Identify an unmet medical need
• Determine the disease's mechanism of action
• Identify the target protein
• Conduct proof of concept: can the target be effectively targeted; what strategy?
• Ensure drug benefits outweigh potential harm
• Screen various compounds against the target protein
• Develop promising compounds into leads
• Identify optimal drug candidates
• Analyze toxicity and pharmacology in animals
• Conduct clinical trials

Drug Design for Chronic Myelogenous Leukemia (CML)

• Unmet Medical Need: Long-term stabilization (4-6 years) is precarious, often leading to accelerated or blast crisis, with bone marrow transplants having high mortality rates (around 45%). Survival rates plummet as the disease progresses.
• Mechanism of Action & Target: Ph translocation is key. p210bcr-abl, a specific fusion protein, has a clear link to CML. This protein is present in over 95% of initial diagnoses. Animal studies confirm direct causal link. Second Ph chromosome signifies disease progression.
• Proof of Concept: Genotype-specific therapy is possible. Gleevec (STI571), an inhibitor of the Bcr-abl tyrosine kinase, is a potential solution. It targets the abl tyrosine kinase for treatment.

Tyrosine Kinases (TKs)

• Structure: TKs have two binding sites: ATP (phosphate donor) and protein (phosphate acceptor). Tyrosine is positioned near ATP for transfer of phosphate.
• Inhibition Strategies: Competitive inhibition at the ATP binding site is useful. However, this is not very specific. More specific inhibition can occur at the protein binding site, preventing substrate binding.

Enzyme-Substrate Interactions

• Lock and Key Model: Enzymes and substrates have specific interactions. Enzyme modifies the substrate into the product. Inhibitors prevent this modification.
• ATP Inhibitors: ATP is a crucial molecule. Inhibiting ATP binding sites poses toxicity risks due to inhibiting too many other enzymes. Thus, inhibitors of ATP binding sites are not appropriate.
• Specific TK Inhibitors: Early kinase inhibitors were often nonspecific. Tyrosine kinase targeted inhibitors (tyrphostins) demonstrated some specificity initially. Focus shifted to inhibitors targeting the protein binding site, leading to greater selectivity for specific substrates.

Abl Tyrosine Kinases (TKs)

• Specificity of Abl TK: Researchers aimed to distinguish normal c-abl from the oncogenic p210bcr-abl fusion protein during drug development. Affinity(strength of interaction) via Km values (substrate concentration for 50% enzyme activity). Low Km=high affinity.
• Km Differences: Kinase domain sequences are nearly identical, but different substrate affinities and Km values exist between p210bcr-abl and c-abl varieties.
• AG-957: A selective inhibitor targeting the p210bcr-abl ATP binding site was found. IC50 (inhibitor concentration for 50% enzyme inhibition) values reflect potency. Lower IC50 values indicate higher activity of the inhibitor.

In Vitro Studies using K562 Cells

• K562 Cell Line: Established CML cell line useful for in vitro studies, known for expressing p210bcr-abl. Key to observing drug activity in live cells.
• AG1112 Inhibition: AG1112 (inhibitor) reduces p210bcr-abl activity in K562 cells, leading to reduced phosphorylation and increased apoptosis. Differentiation toward red cells was also improved.

Development of Gleevec (STI-571)

• Mechanism: Gleevec is a competitive inhibitor of the ATP binding site on p210bcr-abl. It prevents binding, stops phosphorylation, ultimately inhibiting the tyrosine kinase activity.
• Reversible Inhibition: The nature of the inhibitor-bcr-abl complex is reversible: Allows drug dissociated from complex, the kinase remains inactive enabling apoptosis.
• Clinical Trials (Phase I): Trials focused on safety and efficacy. Results showed positive hematological and cytogenetic responses, with mild to moderate side effects. Maximal tolerated dose not determined.

Gleevec-Treated CML Cell

• Apoptosis Induction: Gleevec induces apoptosis in leukaemic cells, while allowing normal cells to retain their function. The drug functions through inhibition of TK activity which prompts apoptosis process.
• Nuclear Import: Gleevec's ability to induce apoptosis in leukaemic cells relies on the drug's interaction with this kinase.

Side Effects & Success of Gleevec

• Comparison to IFN-α: Gleevec demonstrated significantly better outcomes in CML patients compared to treatments based on interferon alpha (IFN-α).
• High Response Rates and Survival: Gleevec treatment resulted in high rates of cytogenetic and hematologic responses, and a significant improvement in long-term survival (approaching 95% at 5 years).
• Drug Resistance: Though highly effective, some patients develop resistance to Gleevec. Development of Gleevec drug derivatives that exhibit mutations to overcome these mutations.
• Broader Applications: Gleevec's success has expanded its therapeutic use beyond CML to GIST.

Explore the intricate steps involved in the drug discovery process specifically tailored for Chronic Myelogenous Leukemia (CML). This quiz covers unmet medical needs, mechanisms of action, target identification, and clinical trials. Test your knowledge on developing effective treatments for this challenging disease.