Format-tuning of in vivo-launched bispecific T cell engager enhances efficacy against renal cell carcinoma

Sep 21, 2026

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Advanced clear cell renal cell carcinoma (ccRCC) shows poor long-term survival, motivating improved immunotherapies. This study develops carbonic anhydrase 9 (CA9)-targeting bispecific T cell engagers (BTEs) and compares a persistent BTE (PBTE) format with a persistent multivalent T cell engager (PMTE) format, including recombinant and synthetic DNA (synDNA) delivery. Binding kinetics, synaptic strength, T-cell activation, cytotoxicity, pharmacokinetics, and in vivo antitumor activity were evaluated in mouse xenograft models using ccRCC cell lines and human donor T cells.

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Open access Original research Format-tuning of in vivo-launched bispecific T cell engager enhances efficacy against renal cell carcinoma Ryan P O’Connell ,1,2 Kevin Liaw,2 Nils Wellhausen,3 Christopher A Chuckran ,4 Pratik S Bhojnagarwala ,2 Devivasha Bordoloi,2 Daniel Park ,1,2 Nicholas Shupin,2 Daniel Kulp,2 Carl H June,3,5 David Weiner2 To cite: O’Connell RP, Liaw K, Wellhausen N, et al. Formattuning of in vivo-launched bispecific T cell engager enhances efficacy against renal cell carcinoma. Journal for ImmunoTherapy of Cancer 2024;12:e008733 . doi:10 . 1136/ jitc-2023-008733 ► Additional supplemental material is published online only. To view, please visit the journal online ([https://doi.org/10.1136/](https://doi.org/10.1136/)[ ](https://doi.org/10.1136/)[jitc-2023-008733](jitc-2023-008733)) . Accepted 02 May 2024 © Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ. 1University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA 2Vaccine & Immunotherapy Center, Wistar Institute, Philadelphia, Pennsylvania, USA 3Center for Cellular Immunotherapies, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA 4LUMICKS, Waltham, Massachusetts, USA 5Parker Institute for Cancer Immunotherapy, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA Correspondence to Dr David Weiner; [dweiner@wistar.org](dweiner@wistar.org) ABSTRACT Background Advanced clear cell renal cell carcinoma (ccRCC) is a prevalent kidney cancer for which long-term survival rates are abysmal, though immunotherapies are showing potential. Not yet clinically vetted are bispecific T cell engagers (BTEs) that activate T cellmediated cancer killing through intercellular synapsing. Multiple BTE formats exist, however, with limited crosscharacterizations to help optimize new drug design. Here, we developed BTEs to treat ccRCC by targeting carbonic anhydrase 9 (CA9) while characterizing the persistent BTE (PBTE) format and comparing it to a new format, the persistent multivalent T cell engager (PMTE). These antibody therapies against ccRCC are developed as both recombinant and synthetic DNA (synDNA) medicines. Methods Antibody formatting effects on binding kinetics were assessed by flow cytometry and intercellular synaptic strength assays while potency was tested using T-cell activation and cytotoxicity assays. Mouse models were used to study antibody plasma and tumor pharmacokinetics, as well as antitumor efficacy as both recombinant and synDNA medicines. Specifically, three models using ccRCC cell line xenografts and human donor T cells in immunodeficient mice were used to support this study. Results Compared with a first-generation BTE, we show that the PBTE reduced avidity, intercellular synaptic strength, cytotoxic potency by as much as 33-fold, and ultimately efficacy against ccRCC tumors in vivo. However, compared with the PBTE, we demonstrate that the PMTE improved cell avidity, restored intercellular synapses, augmented cytotoxic potency by 40-fold, improved tumor distribution pharmacokinetics by 2-fold, and recovered synDNA efficacy in mouse tumor models by 20-fold. All the while, the PMTE displayed a desirable half-life of 4 days in mice compared with the conventional BTE’s 2 hours. Conclusions With impressive efficacy, the CA9-targeted PMTE is a promising new therapy for advanced ccRCC, which can be effectively delivered through synDNA. The highly potent PMTE format itself is a promising new tool for future applications in the multispecific antibody space. INTRODUCTION Renal cell carcinoma (RCC) is a rare and orphan disease responsible for 90% of kidney cancers, which stand as the 9th most common | WHAT IS ALREADY KNOWN ON THIS TOPIC | | --- | | ⇒ Advanced and metastatic clear cell renal cell carcinoma (ccRCC) is a challenging cancer from

FAQs

What cancer and target does the study focus on?

The study focuses on advanced clear cell renal cell carcinoma (ccRCC) and targets carbonic anhydrase 9 (CA9) using bispecific T cell engager formats.

How do PBTE and PMTE differ in reported performance?

Compared with a first-generation BTE, PBTE reduced avidity, synaptic strength, and cytotoxic potency, while PMTE improved avidity, restored intercellular synapses, and increased cytotoxic potency and in vivo efficacy.

What experimental approaches were used to evaluate the therapies?

Binding kinetics and synaptic strength were assessed using flow cytometry and intercellular synaptic assays; potency was tested via T-cell activation and cytotoxicity assays, with mouse models used for pharmacokinetics and antitumor efficacy.

Sep 21, 2026

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