Raltitrexed disodium

Raltitrexed disodium is the disodium salt form of Raltitrexed (RTX, CAS No.: 112887-68-0, Cat. No.: B1476). The core biological function of RTX is as an antifolate antimetabolite, primarily targeting human thymidylate synthase (hTS). By competing with the natural cofactor mTHF for binding and synergistically inhibiting dUMP→dTMP conversion with FdUMP, it depletes dTTP and inhibits DNA synthesis; in Mycobacterium tuberculosis, RTX has also been identified as a potent 4′-phosphopantetheinyl transferase (PptT) inhibitor, blocking the transfer of Ppt from CoA to multiple ACPs, thereby sequentially inhibiting the biosynthesis of mycolic acids and various virulence-related lipids. At the same time, it only weakly inhibits Mtb DHFR, and some analogs can switch from the PptT target to the DHFR target through slight structural changes.

At the cellular level, RTX significantly inhibits cell proliferation at low nanomolar concentrations of 5–20 nM in multiple ovarian cancer cell lines (A2780、A2780/CP、2008、C13*、IGROV‑1, under RPMI‑1640+10%FBS conditions). Single-agent treatment at 5 nM and 20 nM produces approximately 24.5–36% growth inhibition in the 2008、C13*、A2780、A2780/CP cell lines, and can be combined with 5‑FU 5 μM at ratios of 1:1000 or 1:250, showing synergistic TS inhibition with FdUMP; in Mtb H37Rv whole-cell 7H9‑ADN culture, no growth inhibition was observed even when the RTX concentration was increased to 100 μM. Among 800+ analogs, only a few compounds (such as 3e and the DHFR-targeting 4a) showed limited anti-tuberculosis whole-cell activity at high micromolar levels (e.g., MIC90 = 5 μM).

At the clinical level, RTX, under the trade name Tomudex, is used as an antifolate chemotherapeutic agent and has been used for the treatment of advanced colorectal cancer and head and neck squamous cell carcinoma. The typical dose is approximately 2–3 mg/m² by intravenous infusion, often in combination with 5‑FU (clinical exposure approximately 28–39 mg·h/L, corresponding to steady-state plasma levels of approximately 4.7–6.5 μM), with an estimated RTX peak concentration of approximately 1.5 μM; considering that the intracellular TS concentration is several hundred nM and that RTX/FdUMP have nM-level affinity for hTS, together with the enhanced potency and prolonged retention time of RTX after FPGS-mediated polyglutamation, as well as its inhibition of dihydropyrimidine dehydrogenase to slow 5‑FU metabolism, clinical dosing can achieve a high degree of TS occupancy in vivo and pharmacokinetic/pharmacodynamic synergy with 5‑FU.

References:

[1] Singh A, Ottavi S, Krieger I, Planck K, Perkowski A, Kaneko T, Davis AM, Suh C, Zhang D, Goullieux L, Alex A, Roubert C, Gardner M, Preston M, Smith DM, Ling Y, Roberts J, Cautain B, Upton A, Cooper CB, Serbina N, Tanvir Z, Mosior J, Ouerfelli O, Yang G, Gold BS, Rhee KY, Sacchettini JC, Fotouhi N, Aubé J, Nathan C. Redirecting raltitrexed from cancer cell thymidylate synthase to Mycobacterium tuberculosis phosphopantetheinyl transferase. Sci Adv. 2024 Mar 15;10(11):eadj6406. doi: 10.1126/sciadv.adj6406. Epub 2024 Mar 15. PMID: 38489355; PMCID: PMC10942122.

[2] Pozzi C, Santucci M, Marverti G, D'Arca D, Tagliazucchi L, Ferrari S, Gozzi G, Losi L, Tassone G, Mangani S, Ponterini G, Costi MP. Structural Bases for the Synergistic Inhibition of Human Thymidylate Synthase and Ovarian Cancer Cell Growth by Drug Combinations. Cancers (Basel). 2021 Apr 24;13(9):2061. doi: 10.3390/cancers13092061. PMID: 33923290; PMCID: PMC8123127.