Methotrexate-DHFR Inhibitor

With our advanced “DrugLnk” platform and extensive experience in payload chemistry and full synthesis chemistry for antibody-drug conjugates (ADCs) development, Creative Biolabs provides highly customized services for the preparation of ADCs and other targeted antibody therapeutics using methotrexate (MTX) and its analogues as payloads.

Methotrexate, a folate analogue, was initially synthesized in the 1940s to serve as a dihydrofolate reductase (DHFR) inhibitor. It has a core structure comprised of an amino group (NH2) associated with a totally oxidized pteridine ring. MTX has been extensively utilized for the treatment of severe psoriasis and autoimmune diseases such as rheumatoid arthritis (RA). Although limited researches show any specific effects of MTX on the number or function of T cells in RA patients, it does exhibit strong inhibitory effects on neutrophils, particularly on monocytes/macrophages (immune effector cells that play an essential role in RA pathophysiology and inflammatory synovitis) in both in vitro and in vivo analyses. Besides the application in immunological disorders, MTX is one of the most effective therapeutic agents available to treat various malignancies, including lymphocytic leukemia, osteosarcoma, non-Hodgkin’s lymphoma, etc.

Chemical structure of the MTX. MTX is a folate analogue commonly used to treat RA, but it also exhibits potent antitumor activity (Expert Opin Drug Del, 2012).

Methotrexate Mode of Action (MOA)

MTX exerts multiple mechanisms of actions. Up till now, a variety of pharmacological mechanisms of MTX action have been suggested, including DHFR inhibition, suppression of purine and pyrimidine synthesis, inhibition of transmethylation reactions, and improvement of adenosine release with adenosine-mediated inhibition of inflammation. It is speculated that a combination of these mechanisms results in the anti inflammatory effects of methotrexate. In terms of anti-tumor toxicity, studies have shown that the break of the cell cycle induced by high dose MTX treatment may be the first step in the apoptotic sequence of dying cells that might account for the anti-proliferative effect of MTX. What’s more, MTX has been observed to suppress the spontaneous proliferation of U937 monoblastic leukaemia cells in vitro and lead to the rapid expression of the apoptosis receptor CD95.

Schematic representation of the MTX mode of action. After entering the cell, MTX suppresses dihydrofolate reductase (1), thereby interferes with purine and pyrimidine synthesis (2 and 3). MTX also suppresses the enzyme amino-imidazole-carboxamide-ribonucleotide transformylase (4), resulting in an accumulation of adenylate and adenosine, which induces anti-inflammatory effects by binding to adenosine receptors (5) (Int J Clin Rheumatol, 2012).

Methotrexate-based ADCs

ADCs are a novel therapeutic approach for cancer patients. By conjugating the cytotoxic drugs with the antigen-targeting monoclonal antibodies (mAbs), ADCs provide accurate targeting and drug delivery. The carbonyl group and amino group at terminal position of MTX provide excellent functionality for modification with various linkers for subsequent antibody conjugations. MTX conjugates comprising of polyglutamic acid (PGA) or Polyethylene glycol (PEG) spacers have been developed to deliver high dosage of MTX into cancer cells. Meanwhile, other MTX-antibody conjugates have also been developed for the treatment of non-small cell lung cancer and other tumors. With years of experience in ADC development, Creative Biolabs can offer the most suitable linkers and conjugation strategies for innovative MTX-based ADCs production.

With our well-established “DrugLnk” organic synthesis platform, the experienced scientists here at Creative Biolabs are dedicated to help you develop methotrexate-linker complexes using readily available or customized linkers for antibody conjugation in a timely and cost-effective manner. Our customarily tailored services and high quality products will contribute greatly to the success of your projects.

Creative Biolabs also provides other various services regarding ADC development. Please feel free to contact us for more information and a detailed quote.

References：

1. Khan, Z.A.; et al. Methotrexate: a detailed review on drug delivery and clinical aspects. Expert Opin Drug Del. 2012, 9(2): 151-169.
2. Kaltsonoudis, E.; et al. Current and future role of methotrexate in the therapeutic armamentarium for rheumatoid arthritis. Int J Clin Rheumatol. 2012, 7(2): 179-189.