Antibody-Drug Conjugates Targeting the Tumor Microenvironment-202012

Creative BioLabs is a contract research organization based in New York that specializes in antibody discovery, engineering, and biomanufacturing solutions. They discuss the application of antibody drug conjugates (ADC) in tumor killing. ADCs deliver cytotoxic agents to tumor sites, aiming to increase anti-cancer activity while sparing normal tissues. However, ADCs have limitations in their efficacy against solid malignancies, such as breast, lung, and colon cancers. The use of internalizing monoclonal antibodies in ADC therapy has been questioned, leading to the development of non-internalizing ADCs. Non-internalizing ADCs have the potential for strong therapeutic activity against tumors with high mutation rates or antigen loss. They can also induce bystander killing effects and release the payload in the tumor microenvironment. Designing non-internalizing ADCs requires the right payload and a proper cleavable linker. The use of disulfide bonds in non-internalizing ADCs can improve pay Welcome to Creative BioLabs. 100% of the effort, 100% of the service. As a dynamic contract research organization, we are based in New York and serve the whole world. Our seasoned scientists are skilled in antibody discovery, antibody engineering, and biomanufacturing solutions. Today, we are very glad to have you here and share with us some of the new development in the area of ADC. Can you give us a brief introduction of the ADC application that involves with its tumor killing ability? Sure. Antibody drug conjugates have been used in the past 20 years as tools to deliver cytotoxic agents to the tumor site, hoping to increase anti-cancer activity and spare normal tissues from undesired toxicity. ADC, as the name implies, you know, is a conjugate of a cytotoxic agent and a monoclonal antibody, and linking them with a linker. Until recently, most ADC development activities have focused on the use of monoclonal antibodies, which are capable of selective binding and internalization into the target tumor cells. Most of the antibodies that have been used for ADC development display insufficient anti-tumor activity when administered as naked immunoglobulins. On the other hand, the role of the monoclonal antibody moiety in ADC products mainly consists in the selective delivery of a cytotoxic compound at the tumor site, where the latter is released, and acts on cellular targets causing direct damage. According to this mechanism of action, you can think of ADCs as prodrugs for which the cargo release is of fundamental importance for therapeutic activity. We always talk about how great or how innovative the ADC therapies are. But, are there some problems or challenges associated with them today? That's a good question. As far as I know, ADC products specific to internalizing receptors have shown encouraging clinical responses in patients bearing non-solid tumors, but the therapeutic activity against the most frequent solid malignancies, such as breast, lung and colon cancers, is still far from optimal. The clinical efficacy of ADC is not as good as that in animal models. For example, in mice models that bearing tumor, the preclinical data have showed favorable results. There are several internalizing ADC products have led to cancer cures in mice. Whereas in some clinical studies, results have not been as favorable. And for the reason, it could partially be the higher permeability of interstitial tissues in mice than a grafts, compared to solid malignancies in human patients. ADCs that are difficult to penetrate within the tumor mass may not deliver the right payload concentrations. Some researchers have done the immunofluorescence detection studies to test the diffusion properties of monoclonal antibodies and showed the limitations. Studies revealed that M-abs and Ig-formats accumulated on perivascular tumor cells. These monoclonal antibodies are unable, substantially, to penetrate the tumor mass and to reach most neoplastic cells. So what might have been the cause to these problems in ADC therapy? And how can these problems be resolved? Since the beginning of the ADC technology, it has commonly been assumed that the monoclonal antibodies should preferably be directed against tumor-associated antigens expressed on the surface of cancer cells. Ideally, the ADC would internalize upon binding to its cognate target. And ideally, the ADC would facilitate the delivery and release of the cytotoxic cargo inside the malignant cell. Many researchers have showed that this receptor-mediated endocytosis represents the most exploited mechanism for ADC activation. We can imagine that based on internalizing antibodies, ADC products may display at least part of their activity through drug release in the extracellular space. The internalization efficiency is typically variable and rarely reaches 100%. And although we can study the antibody internalization easily in vitro, what we all know that in vivo is a different story. There are many technical limitations related to tumor mass processing and also the specific detection of individual antibody, linker and payload components. So as a result, people begin to question that to develop an ADC therapy, is it actually necessary to use internalizing monoclonal antibodies? There are also additional investigations to find more novel antibodies with tumor targeting properties based on non-internalizing ligands. So to go back to your questions, I think to resolve these problems in ADC therapy, the solution could lie in the development of new non-internalizing ADC products, liberating their toxic payload in the extracellular environment. Wow, sounds exciting. So you think this is a new direction to develop non-internalizing ADC products? Not entirely, but it is definitely one of the important directions. There have been some research about the non-internalizing ADC products. Why is non-internalizing ADC feasible? So theoretically, non-internalizing ADCs can have very strong therapeutic activities against tumors with high mutation rates or characterized by antigen loss. But when it comes to conventional internalizing ADCs, some cell populations can develop resistance to them. And the bystander killing effect could also impair structures which support tumor growth, such as stromal cells, leukocytes, and tumor blood vessels, thus enhancing the anti-tumor effect of the product. And we know that especially in vivo, the tumor environment can be very complex. Some dying cells inside this environment are constantly releasing reducing agents to the surrounding areas. And disulfide bonds-based non-internalizing ADC products could be cleaved in the tumor extracellular milieu, releasing the payload and promote apoptosis in cancer cells. So when it comes to non-internalizing ADCs, ideally, we can find monoclonal antibodies specific to both tumor-specific extracellular structures and poorly or non-internalizing transmembrane antigens. Okay, so it seems that the mechanisms of non-internalized ADC products is different from internalized ADC. Can you talk about non-internalized ADCs mechanism of action? So different from the traditional receptor mediated endocytic process, when using non-internalized ADCs, drug release from tumor targeting devices could ideally take place in the tumor microenvironment. This allows the subsequent diffusion of the active payload and its internalization into neighboring neoplastic cells. Since passive diffusion is a nonspecific process, the cytotoxic agent has the potential to reach antigen-negative cancerous cells within the tumor mass. We call this mechanism the bystander effect. What are some key points when designing non-internalized ADC? Other than choosing the right payload, when designing non-internalizing ADC, a proper linker is crucial to generate an effective and well-tolerated ADC. There are the cleavable and non-cleavable linkers. Both types of linkers are used in internalizing ADC products. But when it comes to the non-internalized ADC, only the cleavable linker is used so far. Can you elaborate more on the cleavable linker that is used in the design of non-internalized ADCs? Various cleavable linkers have been proposed for the preferential drug release in the tumor interstitium. To prevent premature drug release and the related side effects, one main requirement for the linker is a high linker stability in plasma after ADC administration. Provided that a sufficient amount of the ADC reaches intact the tumor microenvironment, a second key attribute of the linker is the ability to efficiently release the payload at the tumor site. Several ADCs in small molecule-targeted cytotoxicity that incorporate reducible linker systems, such as disulfide bridges, have been considered for clinical development. Disulfides are typically stable in the absence of free thiols at physiological pH, with a serum half-life that can be longer than one week. In vivo, certain disulfide-based ADCs have exhibited stability in blood for two to four days. Moreover, this stability can be dramatically improved by increasing the steric hindrance of substituents at the cleavage site. While disulfide-based linkers have been designed for the intracellular release of anti-cancer drugs, the same chemical structures can be considered for the extracellular drug release as a consequence of tumor cell death and increased glutathione concentration. Glutathione represents the most abundant thiol and reducing agent in the intracellular space, both in normal cells and in tumors, which often contain higher concentrations of this species. So as we mentioned before, we can use disulfide bonds in non-internalizing ADC products to better release the payload and promote apoptosis in cancer cells. What else do you think need to be improved in the development of non-internalizing ADC? I think that improving the potency and selectivity of non-internalizing ADC products is something that we really need to look into right now. Thanks for sharing your wonderful insights. And before we finish today, would you have any additional comments on the ADC? Okay, I'm very glad to have the opportunity to be here with you. So to conclude, the possibility to develop non-internalizing ADCs is, by now, firmly established, at least at the preclinical level. The field of ADC research, both for internalizing and non-internalizing products, will continue to face an important scientific challenge, namely the translation of preclinical data into a prediction of efficacy in patients. One of the most challenging issues for future developments in the ADC field relates to the quantification of product uptake in mouse and man, as well as to the comparative evaluation of drug release kinetics in different species. In particular, a quantitative evaluation of the targeting properties of ADCs in human patients is often missing, which negatively impacts on the clinical development of drug candidates. Although many challenges remain, we hope to see more breakthroughs in the future.