Conditionally activated immunotherapies alleviate cancer treatment toxicity

Our immune system guards against cancer development by constantly surveilling the body and eliminating unhealthy cells. Immune cells are highly suspicious of other cells. While this leads to a swift response when cancer is developing or a foreign pathogen is invading, sometimes immune cells can overreact and target healthy cells. To prevent this from happening, immune checkpoint proteins interact with immune cells to block their functions. These proteins are crucial for health, but some cancers use these same checkpoint proteins to evade the immune system. This allows the cancer cells to remain in the body, proliferate, and drive disease uninterrupted.

Researchers have developed a type of immunotherapy called immune checkpoint inhibition to combat this cloaking by the cancer cells. Immune checkpoint inhibitors are antibodies that block the checkpoint proteins from turning off the immune response to cancer. Ideally, checkpoint blockade would overcome tumor immunosuppression and allow immune cells to kill cancer cells. While this type of therapy is often effective, it comes with major downsides. As noted above, checkpoint proteins are there for a reason—they’re necessary to ensure immune cells avoid attacking healthy tissue. Checkpoint inhibitors are typically administered intravenously, meaning that they can circulate to block immune checkpoints in the tumor (good!), but also in healthy tissues (bad!). Because of this, checkpoint-blockading therapies come with some potentially serious side effects like rashes, diarrhea, fatigue, and even chronic inflammation. In the past, researchers have tried injecting checkpoint inhibitors directly into the tumor to prevent these off-target effects, but this strategy is ineffective when patients have metastatic cancers.

Blockade of the checkpoint protein CTLA-4 is notorious for causing adverse reactions in patients. Targeting CTLA-4 blockade to the site of the tumor is one way to overcome these adverse reactions and improve the safety and efficacy of immune checkpoint inhibition. Dr. Larry Fong, Scientific Director of the Immunotherapy Integrated Research Center and faculty member at Fred Hutch, and his team tested this approach in a new study published in the Journal for Immunotherapy of Cancer. They leveraged previous discoveries that prostate tumors have high amounts of an enzyme called matriptase that cleaves other proteins at specific sites to create new tumor-targeted checkpoint inhibitors. “The goal here is to think about ways that we can use this tumor-conditional approach to really improve the therapeutic index…of treatments,” says Fong.

Unlike previous iterations of antibodies that block CTLA-4 everywhere, Fong’s new antibodies have an extra cleavable domain that binds to a prostate tumor marker. Once bound, enzymes produced by tumor cleaves the tumor-targeting portion of the antibody to reveal the CTLA-4-binding portion of the checkpoint inhibitor. This design means that these new checkpoint inhibitors could be administered intravenously, but they would not activate until they arrive at the prostate tumors, preventing side effects in healthy tissues. The team tested two different forms of these conditionally active antibodies: one form was incompletely cleaved so that it stayed bound to the tumor marker, and the other was completely cleaved so it could move around the tumor microenvironment.

To start, the group tested these new antibodies with traditional CTLA-4 blocking antibodies. They induced prostate tumors in mice and treated them with three doses of the partially cleaved conditionally activated antibody, the completely cleaved conditionally activated antibody, or the traditional therapy when all the tumors reached the same size. They found that the completely cleaved new antibody inhibited the tumor growth to the same extent as the traditional anti-CTLA-4 antibody. Mice treated with either of these checkpoint inhibitors had similar survival outcomes as well, indicating that the fully cleavable antibody was just as effective as approved therapies. Mice treated with the partially cleaved antibody had more tumor growth and worse survival outcomes compared to mice in the other two groups, indicating that this checkpoint inhibitor is less effective. To assess off-target toxicity, the team monitored the body weight of the mice throughout the study. If the mice lost significant amounts of weight, they inferred that their treatment more toxic. Unsurprisingly, mice receiving the current anti-CTLA-4 inhibitors lost a significant amount of weight throughout the study. Mice treated with either form of the conditionally activated inhibitor lost no weight, suggesting that the conditionally activated therapies are less toxic than current therapies.

Because metastases are common in prostate cancer, the group wanted to see how their antibodies would perform in models of metastatic cancer. To do this, they induced metastatic prostate cancer in mice and treated them with the different checkpoint inhibitors. Again, they found that mice treated with the cleavable antibody had similar tumor control and survival outcomes to the group treated with current CLTA-4 inhibitors. The group analyzed how their new checkpoint inhibitor induced immune responses in mice. They found that mice that received the new checkpoint inhibitor had more activating immune cells and fewer repressive immune cells throughout their bodies. These immune changes at the site of the tumor and throughout the body indicate that the new antibody construct is more effective at inducing immune responses than the current construct.

In the future, the Fong lab hopes to use similar conditional activation approaches to improve other types of cancer immunotherapies. “We’re really looking forward to pushing this whole area forward at the Hutch,” says Fong, “Having the resources to be able to do that will hopefully help accelerate these types of discoveries.”

Fred Hutch/UW/Seattle Children’s Cancer Consortium member Dr. Larry Fong contributed to this work.

This work was supported by funding from AbbVie and the National Institute of Health.

Arias-Badia M, Pai CS, Lwin YM, Chen P, Srinath A, Tanaka M, Musser E, Goodearl A, Gorman JV, Ritacco W, Fong L. 2025. Impact of tumor localization on antitumor immunity with conditionally activated CTLA-4 blockade. J Immunother Cancer. 13(4):e010566. doi: 10.1136/jitc-2024-010566.