Methotrexate exerts its cytotoxic effect primarily by inhibiting which enzyme in the folate metabolism pathway?

Explanation

## Methotrexate: Mechanism of Action **Key Point:** Methotrexate is a folate analog that competitively inhibits dihydrofolate reductase (DHFR), blocking the conversion of dihydrofolate (DHF) to tetrahydrofolate (THF), thereby disrupting one-carbon transfer reactions essential for nucleotide synthesis. ### Mechanism Flowchart ```mermaid flowchart TD A[Methotrexate enters cell]:::action --> B[Binds DHFR competitively]:::action B --> C[Blocks DHF → THF conversion]:::urgent C --> D[Depletes THF pools]:::outcome D --> E[Inhibits purine synthesis]:::outcome D --> F[Inhibits thymidylate synthase]:::outcome E --> G[Reduced dNTP availability]:::outcome F --> G G --> H[Cell cycle arrest & apoptosis]:::outcome ``` ### Polyglutamation & Selectivity 1. MTX enters cells via reduced folate carrier (RFC) 2. Intracellular MTX is polyglutamated by folylpolyglutamyl synthetase 3. Polyglutamated MTX has higher affinity for DHFR and is retained intracellularly 4. This creates selectivity for rapidly dividing cells with active folate metabolism **High-Yield:** MTX is a folate antimetabolite, not a purine or pyrimidine analog. It is used in high doses for cancer (with leucovorin rescue) and low doses for autoimmune diseases (rheumatoid arthritis, psoriasis). ### Downstream Effects - **Purine synthesis:** THF is required for AICAR transformylase and FGAM synthetase - **Pyrimidine synthesis:** THF is required for thymidylate synthase (dUMP → dTMP) - **Amino acid metabolism:** THF is required for serine, glycine, and methionine metabolism **Clinical Pearl:** Leucovorin (folinic acid) rescue is used after high-dose MTX to replenish THF pools and prevent toxicity to normal tissues, particularly bone marrow and GI epithelium. **Mnemonic:** **DHFR** = **D**ihydrofolate **R**eductase is the target of **M**etho**T**rexate. ### Comparison: Folate Antagonists vs Other Antimetabolites | Drug | Target Enzyme | Nucleotide Affected | Cell Cycle Phase | |------|---------------|-------------------|------------------| | Methotrexate | DHFR | Both purines & pyrimidines | S-phase | | 5-Fluorouracil | Thymidylate synthase | Pyrimidines (dTMP) | S-phase | | 6-Mercaptopurine | HGPRT, IMPDH | Purines | S-phase | | Pemetrexed | DHFR, GARFT, TS | Multiple (multi-target) | S-phase |