Engineering Antibodies & Beyond

Conditionally-active biologics have emerged as a powerful approach to improve the therapeutic index of protein drugs by restricting activity to diseased tissues. In this presentation, we will first review current design trends, successes, and remaining limitations. We then present a de novo computational workflow for minimal cleavable peptide masks that reversibly inactivate miniprotein binders, enabling library-free generation of next-generation conditionally-active biologics.

Immunomodulatory antibodies represent an exciting new modality for immunotherapy. Whereas checkpoint blockers have translated successfully into the clinic, agonistic antibodies directed to immune receptors have lagged behind and represent an untapped opportunity for immunotherapy. Here, we highlight the salient properties of antibodies required to agonise both activating and inhibitory receptors and discuss the role of affinity, hinge rigidity and geometry. Using examples across the TNFR super-family, (CD40, 4-1BB), the immunoglobulin super-family (CD28), and including inhibitory receptors (PD-1, FcgRIIB), the contribution of each of these aspects will be examined: 1) Affinity, 2) Geometry, 3) Hinge rigidity.

A central limitation of antibody therapeutics is the inability of full-length IgG to productively access the cytosol. I will present our second generation cytosol-penetrating antibody platform, engineered with pH-responsive endosomal escape motifs to enable efficient intracellular delivery in an IgG format. In tumour-targeted designs, this platform supports cytosolic delivery of an engineered granzyme B bio-payload and potent anti-tumour activity in solid tumour models. The presentation will discuss antibody-toxin conjugates as a mechanistically differentiated modality that extends antibody drugs beyond extracellular blockade and lysosomal payload release toward programmable intracellular biologic delivery.

Cancer can involve aberrant transcription factor complexes considered undruggable. Intracellular antibody variable region fragments are effective inhibitors of protein interactions. Further, they can be augmented with effector functions, such as E3 ligases or procaspases. However, delivering these proteins into cells as potential drugs is problematic. We will describe nanoparticles to deliver mRNA cargoes expressing intracellular proteins, coupled to enhanced effectiveness with nanoparticle-coated gateway molecules interacting with receptors on target cells.

Stronger tumour growth inhibition was demonstrated with targeted disruption of the hyaluronan (HA) barrier that surrounded solid tumour cells. Systemic delivery of hyaluronidase can break down HA but is not effective when used alone and can cause adverse effects. We demonstrated how an “antibody–enzyme complex” (AbEn) disrupted the HA barrier via homing to the tumour or cancer associated fibroblast cells.

Targeting oncogenic Ras mutants with small molecules remains highly challenging and has yielded mixed results. We explored a powerful and innovative strategy to target Ras-driven pancreatic cancer by leveraging the natural Ras effector and tumor suppressor Nore1A. We engineered Nore1A into a RasG12D-specific “superbinder” and endowed it with cell-penetrating capabilities. When tested in pancreatic cancer cells harboring the RasG12D mutation, the engineered protein exhibited a dual therapeutic effect, simultaneously suppressing pro-tumourigenic processes while activating anti-cancer pathways.

Elucidating the mechanisms driving response to CD3 bispecifics, alongside strategies to mitigate toxicity, is critical for expanding and optimising their efficacy in solid tumour indications.

Engagement of costimulatory receptors is an emerging frontier in solid tumor treatments with the promise to promote sustained T cell activity and tumor cell killing. We will highlight our CD28 platform design and the development of RNDO-564, a precisely tuned CD28 × Nectin-4 bispecific antibody with an optimized therapeutic window for the treatment of advanced bladder cancer. An update of the clinical development of RNDO-564 will be provided.