In this section:
- Highly potent cytotoxic anticancer agents
- Mechanism of Toxicity
- Use in Antibody-drug Conjugates
|Tubulin polymerase inhibitor
|DNA minor groove alkylating agent
|DNA minor groove cross-linker
|RNA polymerase II inhibitor
Fig. 1.0. Cytotoxic Mechanism of Action
Today, two main categories of highly potent cytotoxic anticancer agents are used in antibody-drug conjugates: microtubule disrupting agents and DNA modifying agents. Microtubules play important roles in the cell cycle. If they malfunction cells are unable to divide. Most of the known microtubule disrupting agents are derived from natural product sources and are extremely cytotoxic. Unlike drugs that affect the microtubules and only work on cells in certain stages of the cell cycle, DNA modifying agents will kill cells at any point.
Several microtubule disrupting agents are being used and tested in clinical trials. Seattle Genetics, for example, has made numerous derivatives of dolastatin 10, a natural cytotoxic pseudopeptide, which was originally isolated from the Indian Ocean sea hare Dolabella auricularia. Dolastatin 10 is a very potent inhibitor of tubulin polymerization. However, while the dolastatin family has demonstrated antineoplastic, bactericidal and fungicidal properties, in clinical trials they did, like the majority of (very) potent ‘natural’ cytotoxins, not show sufficient activity at a a tolerable dose.
Monomethyl Auristatin E (MMAE), with the generic name vedotin, was designed to be released from the antibody by cathepsin B enzymatic cleavage. A second derivative, Monomethyl Auristatin F (MMAF), is attached via a non-cleavable linker, with a charged group to limit cell egress.
Maytansine and Maytansine derivates
Immunogen has also developed a family of microtubule-disrupting payloads. Their agent, an ansamycin antibiotic, is derived from the natural product maytansine, originally isolated from the Ethiopian shrub of the genus Maytenus serrata. Maytansine binds to tubulin at the rhizoxin binding site, thereby inhibiting microtubule assembly, inducing microtubule disassembly, and disrupting mitosis. Maytansine exhibits cytotoxicity against many tumor cell lines and may inhibit tumor growth in vivo.
These derivatives have been engineered to incorporate thiol groups, which are used to conjugate the molecules to the linker. DM1 gives a non-hindered disulfide once coupled to the linker, making it readily cleavable in the reducing environment inside the cells. DM4 has a dimethyl substitution alpha to the disulfide linkage, making it a little less easy to reduce.
Because rapidly dividing cells, such as cancer cells, are much more susceptible to the toxic effects of cytotoxic agents or drugs than normal cells that are dividing slowly, it can be beneficial to use DNA modifying agents to kill off the rapid dividers. DNA damage is the basis for many of the most commonly used chemotherapeutic agents, but those that are used in the clinic have relatively low potency, because the therapeutic index drops as the potency increases.
Different types of DNA modifying agents
The DNA modifying agents also tend to be derived from natural products. Bristol-Myers Squibb and Medarex have developed various natural product-derived APIs which bind in the minor groove of DNA, positioning the toxin to alkylate the DNA via nucleophilic attack to cause mutagenesis. A family of synthetic toxins called duocarmycin analogues from The Netherlands based Synthon (formerly Syntarga) also binds this way. Although none of these are yet in the clinic, a good deal of work has been done to dial in different linker properties and activities.
The second group of DNA damaging agents that are being used in ADCs are DNA strand scission inducing agents. Pfizer (formerly Wyeth) uses derivatives of calicheamicin, an enediyne antibiotic originally isolated from the bacterium Micromonospora echinospora. Again, they bind to the minor groove of DNA. In this case, instead of alkylating the DNA, once attached they undergo a chemical reaction which generates a diradical species and in turn breaks one of the DNA strands.
Calicheamicin is the active agent in gemtuzumab ozogamicin (Mylotarg; Pfizer), which was approved in 2000. However, confirmatory trials beginning in 2004 were stopped early when researchers did not observe a clinical benefit, and after a greater number of deaths were observed in the group of patients receiving gemtuzumab ozogamicin compared to those receiving chemotherapy alone. The results of a randomized study by the Southwest Oncology Group (SWOG), one of the largest National Cancer Institute-supported cancer clinical trials cooperative groups, led to the voluntary withdrawal of gemtuzumab ozogamicin in 2010, when improved efficacy could not be demonstrated while toxicity appeared to be excessive
The same agent is used as the payload for Pfizer’s second generation antibody-drug conjugate CMC-544 (inotuzumab ozogamicin), which is currently in clinical trials.
Other agents used in the developments of ADCs includes:
- pyrrolobenzodiazepines (PBDs);
- centanamycin (ML-970; indolecarboxamide);
- SN38 is the active metabolite of irinotecan (an analog of camptothecin, a topoisomerase I inhibitor);
- doxorubicin, a drug dating back to the 1950s and still used today for the treatment of a number of cancers