Unlocking IL-12

Interleukin-12 (IL-12) is a highly promising cytokine for the treatment of solid tumors. This potent molecule plays a crucial role in both innate and adaptive immune responses, significantly remodeling harsh, immune-suppressive tumor microenvironments (TME) to make them more conducive to immune control. Despite its potential, there are currently no IL-12 cytokine drugs approved for patient use. The potent anti-cancer responses observed in pre-clinical settings have solidified our commitment to developing IL-12 based therapies for the benefit of a growing number of solid cancer patients.

In this article, we’re examining the function and effects of this powerful immune stimulatory cytokine, why it’s such a strong candidate in cancer immunotherapy, the issues that are currently preventing it from being used as a cytokine therapy, and how we’re overcoming these obstacles at Audax Biosciences.

Why Use the IL-12 Cytokine in Cancer Immunotherapy?

There are many different types of cytokines with different effects on the immune system, but IL-12 really stands out in the context of cancer. Let’s investigate what this cytokine is and how it functions in cell-mediated immunity.

What is IL-12?

IL-12 is a proinflammatory cytokine that displays potent pleiotropic effects on the innate and adaptive immune systems. In terms of structure, it is made up of the p35 and p40 subunits, which are joined by disulfide bonds to make a p70 heterodimeric protein. This cytokine is mainly produced by antigen-presenting cells (APCs) – like macrophages, B cells and dendritic cells – in response to cancer. It binds to its receptors (IL-12RB1/2) expressed on the surface of T cells and natural killer (NK) cells, with signal transduction occurring through the JAK-STAT pathway.

What does IL-12 do?

As we already mentioned, the IL-12 cytokine is highly pleiotropic, meaning that it has a wide range of effects. It acts either directly or indirectly on several different immune cell types, enhancing the function of cytotoxic effector lymphocytes and increasing their tumor-killing capacity.

IL-12 positively regulates the differentiation and activation of T helper 1 (Th1) cells, which support the effector function of cytotoxic CD8+ T cells. It directly activates NK cells and CD8+ T cells. By elevating the production of various chemokines, it promotes the trafficking of cytotoxic lymphocytes into tumors. Once at the site of the tumor, it allows CD8+ T cells and NK cells to proliferate and accumulate within the tumor microenvironment (TME).

This powerful cytokine also causes activated cytotoxic lymphocytes to secrete interferon-gamma (IFN-γ), granzyme, and perforin – molecules that can induce the death of cancer cells and limit the growth of tumors. IFN-γ is another important pro-inflammatory cytokine that recruits more myeloid cells to the TME, which in turn secrete more IL-12, encouraging more CD8+ T cells and NK cells to infiltrate the tumor.

The presentation of antigens on tumor cells is another way in which this cytokine helps the immune system fight cancer: it upregulates the expression of major histocompatibility complex I (MHC-I) & MHC-II the molecules responsible for placing antigens on the surface of cancer cells. It also drives the differentiation of macrophages into the M1 type, which suppress tumor growth!

Importantly, IL-12 is able to prevent negative regulatory receptors on CD8+ T cells from signalling; it can downregulate the expression of PD-1 on CD8+ T cells and can protect them from IFN-γ-induced apoptosis. Additionally, it has interesting effects on immunosuppressive regulatory T cells (Tregs): it may impair Tregs suppressor activity or arrest their cell cycle, preventing them from accumulating within tumors.

IL-12 Cytokine Therapy: Potential, Obstacles, and Solutions

The use of IL-12 as a cytokine therapy has shown incredible promise in preclinical cancer research, but there are many obstacles we need to overcome before we can use it to treat human cancer patients. Let’s look at how it can be used as a cytokine therapy, the issues that are preventing its success in the clinic, and some of the different strategies that are being employed to solve these problems.

The power of IL-12 to treat solid tumors

IL-12 is a powerful immunomodulator that dramatically affects the immune system’s response to cancer, making it an attractive cytokine for cancer treatment. Its pleiotropic effects allow it to remodel the immunosuppressive tumor microenvironment (TME), recruiting cytotoxic lymphocytes, encouraging them to infiltrate immunologically ‘cold’ tumors, and enhancing their tumor-killing capacity.

Another major advantage is that this cytokine is highly potent at much lower doses than other cytokines like IL-2. Its biology is also more dominant and predictable than other cytokines. For example, while IL-2 can benefit effector T cell responses to cancer, it preferentially promotes the activity of suppressive regulatory T cells (Tregs) at low, tolerated doses, meaning that depending on the exposure of IL-2 within the TME, this approach might work against the desired effect of improving tumor control.

Since IL-12 induces IFN-γ production from various immune cells, it can indirectly enhance PD-L1 expression on tumor and antigen-presenting cells. While induction of PD-L1 may reduce some of the immune-activating effects of this cytokine, this could be counteracted by combining with PD-1 or PD-L1 immune checkpoint inhibitor antibodies – likely even improving the overall therapeutic effect. IL-12 could also be used in rational combination with other cancer treatments, such as chemotherapy and radiation therapy. In fact, if it is administered before the surgical removal of tumors, it can limit cancer recurrence and the growth of secondary tumors (metastases).

At this stage, you might be wondering: If this cytokine is so powerful, why isn’t it being used to treat cancer right now?

Limitations of IL-12 cytokine therapy

While IL-12 possesses powerful anti-tumor capacity, there are some key limitations that have hindered its clinical translation. A major issue is how long the molecule lasts – much like other cytokines, it has an extremely short half-life in vivo, meaning that repeat doses are necessary to achieve any therapeutic benefit. The short half-life of cytokines such as IL-12 is in part due to its high affinity for its receptors on peripheral immune cells, such as NK cells and CD8+ T cells circulating in our blood.

As such, this cytokine has a very narrow therapeutic window, meaning the dose required for potent anti-tumor immunity is similar to, or even above, doses that are toxic to patients. This leads to a phenomenon known as dose-limiting toxicity, largely driven by the fact that systemic administration of IL-12 results in the activation of circulating immune cells, rather than just the immune cells in and around the tumor site. It also induces cytokine release syndrome – the release of IFN-γ and other inflammatory cytokines throughout the body, rather than only at the site of the tumor where they are needed.

These toxicity issues, combined with the need for repeat high doses, have largely prevented this amazing cytokine from being used to treat cancer in humans.

Approaches to next-generation IL-12 therapies

To create a successful IL-12 cytokine therapy for cancer, we need to overcome these key obstacles, extending the half-life of the molecule and keeping it within an effective therapeutic window to avoid toxicity. There are several different approaches that are currently being explored in this field, with a focus on localized delivery to avoid the systemic activation of circulating lymphocytes.

These strategies include plasmid-based delivery, the use of viral vectors, the creating of tumor-specific chimeric antigen receptor (CAR) T cells that are ‘armoured’ with IL-12, and mRNA and lipid nanoparticle (LNP)-based approaches.

The Audax Approach to IL-12 Cytokine Therapy

We are determined to harness the power of IL-12 for the treatment of solid tumors. Our goal is to create therapies that will treat a wide range of solid tumors rather than focusing on a single type of cancer, and we believe this cytokine can do just that.

Our DAX-044 candidate has addressed all the issues that have plagued IL-12 therapies in the past to deliver a molecule that truly behaves like a drug. Firstly, we have engineered the molecule to reduce its affinity for IL-12R, meaning it does not trigger the activation of peripheral immune cells and therefore prevents systemic toxicity. By avoiding peripheral immune cells, the half-life of DAX-044 is also increased as its not being bound and used up by those circulating immune cells. Its half-life is further extended by fusion to the Fc region of an antibody, meaning DAX-044 stays within an effective therapeutic window for longer and does not require frequent repeated high doses that typically lead to toxicity with other IL-12-based drugs.

After we tackled the safety issues and enhanced its pharmacokinetic parameters, we were determined to improve the specificity of the molecule, further reducing the activation of non-target immune cells and amplifying its effects within the TME. To do so, we engineered DAX-044 to contain a binding domain (an antibody fragment) that directs it toward an antigen that is specifically expressed on certain immune cells within the TME that are responsible for potent anti-tumor immunity. While DAX-044 has a reduced potency on immune cells that lack the expression of this antigen, immune cells in the TME that express this antigen experience greatly enhanced IL-12R agonism or IL-12 potency due to an avidity phenomenon.

With this innovative approach, we have created a product that retains the powerful anti-tumor capacity of the cytokine while displaying a remarkable safety profile – DAX-044 packs a punch at just a fraction of the doses normally required for this type of therapy. With the potential to treat a variety of solid tumors, we think that DAX-044 is the most promising IL-12 based therapeutic currently in development.

When Will IL-12 Cytokine Therapy Reach the Clinic?

While a few IL-12 based therapies are currently in clinical trials, their therapeutic window as systemic therapies is narrow. DAX-044 has shown a dramatically improved therapeutic window and safety profile in preclinical models compared to previous approaches. This improvement should allow DAX-044 to be administered at highly effective doses without causing the serious adverse side effects associated with the peripheral immune activity of earlier IL-12 based therapies. Consequently, we can finally evaluate the potent immunomodulatory effects of this cytokine in solid cancer patients in future clinical trials.