Logic-gated DARPins show targeted T-cell activation in tumours

T-cell engagers (TCEs) have become an important therapeutic approach in oncology, but their development for solid tumours remains difficult. Many TCEs activate T cells systemically or show on-target off-tumour effects, leading to cytokine release and dose-limiting toxicity. Others struggle to achieve sufficient T-cell activity within the tumour itself. Molecular Partners has been developing an alternative approach built around its DARPin technology, designed to activate T cells only where two tumour-associated antigens are present.

At SITC 2025, the company presented new preclinical proof-of-concept data showing that its logic-gated CD3 Switch-DARPin TCE, which also incorporates CD2 co-stimulation, can selectively be activated (“Switched” on) against tumour cells that co-express mesothelin (MSLN) and epithelial cell adhesion molecule (EpCAM). These two proteins are tumour-associated antigens that are frequently found together on the surface of ovarian cancer and other solid tumour types. The data expand on earlier findings and support the potential of this modality to improve the balance between efficacy and safety in solid tumour treatment by restricting T-cell activation to cells that display both targets.

A logic-based approach to T-cell engagement

Traditional TCEs work by binding CD3 on T cells and a single tumour-associated antigen on cancer cells. When that antigen is also present in healthy tissues, these molecules can trigger off-tumour immune activation and systemic toxicity. Molecular Partners developed its Switch-DARPin to address this limitation using an AND-gate design. In this format, the CD3-binding DARPin stays masked and inactive until the molecule binds both mesothelin (MSLN) and epithelial cell adhesion molecule (EpCAM). Only when both antigens are present is the CD3 domain unmasked, enabling T-cell activation specifically in the tumour microenvironment.

MSLN and EpCAM are co-expressed in ovarian cancer and several other solid tumour types, but are generally not co-expressed in healthy tissues. By requiring the presence of both antigens, the Switch-DARPin is intended to function selectively against dual-positive tumour cells while showing reduced activity in tissues where only one antigen is expressed.

Preclinical results presented at SITC 2025

The new data show that the Switch-DARPin produced selective cytotoxicity in vitro against cells expressing both MSLN and EpCAM, with reduced activity against cells expressing only one of the targets. This supports the principle of conditional activation and the intended tumour localisation.

In an in vivo xenograft model expressing both antigens, the Switch-DARPin induced significant tumour regression.

The addition of a CD2-engaging DARPin was also evaluated. CD2 is a co-stimulatory receptor involved in T-cell activation and persistence. When T cells were repeatedly exposed to the CD2/CD3 Switch-DARPin, they displayed improved activation and proliferation compared with CD3 engagement alone. This is particularly relevant in solid tumours, where T-cell exhaustion is a common barrier to effective response.

In an in vivo xenograft model expressing both antigens, the Switch-DARPin induced significant tumour regression. Importantly, there was no evidence of systemic cytokine release in the preclinical model, suggesting a favourable safety profile and reinforcing the conditional, tumour-localised activation mechanism.

Insights from Molecular Partners

Alexander Link, Senior Director of Immuno-Oncology Research at Molecular Partners, commented on the findings:

We show that this particular Switch-DARPin selectively targets and activates T cells only in the presence of both tumour-associated antigens MSLN and EpCAM; otherwise, it remains inactive in circulation.

“The data in our poster presented at SITC 2025 underline the promising preclinical safety and efficacy profile of our logic-gated CD3 Switch-DARPin T-cell engager with CD2 co-stimulation.”

He explained how the AND-gate architecture contributes:

“We show that this particular Switch-DARPin selectively targets and activates T cells only in the presence of both tumour-associated antigens MSLN and EpCAM; otherwise, it remains inactive in circulation.”

On CD2 co-stimulation:

“This allowed us to evaluate the addition of a CD2 DARPin for co-stimulation of T cells, which we found helped combat T-cell exhaustion.”

On the in vivo results:

“In a xenograft solid tumour mouse model, the CD3 Switch-DARPin T-cell engager showed selective tumour cell destruction, enhanced T-cell activation and significant tumour regression, without systemic cytokine release.”

Overcoming limitations of traditional TCEs

The Switch-DARPin design is intended to address two well-recognised challenges in developing TCEs for solid tumours.

1. Systemic toxicity

Traditional TCEs bind CD3 on T cells and a single tumour antigen. When that antigen is also expressed on healthy tissues, T cells can be activated outside the tumour, leading to off-tumour toxicity and systemic cytokine release. The Switch-DARPin remains inactive in circulation and “only activates T cells when both tumour-associated antigens are bound”.

2. T-cell exhaustion and limited durability

Solid tumour TCEs often struggle with insufficient T-cell activity over time. Adding a CD2-engaging DARPin provided co-stimulation that “helped combat T-cell exhaustion”, supporting stronger and more durable activity.

How the DARPin platform enables multispecificity

DARPins are small, engineered ankyrin repeat proteins with high affinity and modularity. Their architecture allows the creation of multispecific and multifunctional molecules, enabling designs such as logic-gated, multispecific TCEs with conditional features. The platform also offers advantages in stability, manufacturability and yield, supporting complex constructs like the CD3 Switch-DARPin.

Implications for solid tumour therapy

Solid tumours present a challenging environment for immune-based therapies due to physical barriers, immunosuppressive signalling and heterogeneous antigen expression. Many TCEs have shown promise in haematological malignancies but have struggled to translate similar benefits to solid tumours.

By integrating antigen specificity with localised activation and co-stimulation, the Switch-DARPin approach aims to bring greater precision and a broader therapeutic window to tumour-directed T-cell engagement. The preclinical data suggest that conditional activation could limit toxicity while preserving or enhancing antitumour activity.

If these findings translate into clinical settings, logic-gated TCEs could represent a meaningful advance in the field, particularly for tumours where pairs of antigens are reliably co-expressed.