Building evidence on the PKPD relationship with bispecific co-stimulatory antibodies using in vitro data: FAP-4-1BBL example
Javier Sanchez (1, 2), Christina Claus (3), Rosmarie Albrecht (3), Brenda Gaillard (3), Joana Marinho(3), Christine McIntyre(4), Siv Jonsson (2), Lena E. Friberg (2), Nicolas Frances (1)
(1) Roche Pharma Research and Early Development, Roche Innovation Center Basel, F. Hoffmann-La Roche AG., Basel, Switzerland. (2). Department of Pharmacy, Uppsala University, Uppsala, Sweden. (3). Roche Pharma Research and Early Development, Roche Innovation Center Zurich, Schlieren, Switzerland. (4) Roche Pharma Research and Early Development, Roche Innovation Center Welwyn, Roche Products Ltd.
Objectives: FAP-4-1BBL is a bispecific antibody currently in clinical development for solid tumors. FAP-4-1BBL exerts 4-1BB-associated T-cell activation only while concomitantly bound to the Fibroblast Activation Protein (FAP) expressed on the surface of cancer-associated fibroblasts [1]. The molecular entity formed by FAP-4-1BBL bound to both 4-1BB and FAP is called trimeric complex (TC). The TC formation is expected to follow a bell-shaped curve with increasing FAP-4-1BBL concentrations [2], with TC formation dropping at the highest FAP-4-1BBL concentrations. In addition, the number of TCs formed depends on receptor expression [3] of both FAP and 4-1BB. In this work, we combine information from in vitro data with mathematical modeling to characterize the FAP-4-1BBL pharmacological effect versus TC formation when used in combination with cibisatamab, a CD3 T-cell bispecific targeting the carcinoembryonic antigen cell adhesion molecule 5 [4]. We then use this model to inform on the FAP-4-1BBL concentration allowing maximum TC formation, as well as the effect of FAP expression in TC formation and pharmacology.
Methods: In vitro tumor cell killing with 2 nM cibisatamab in monotherapy or combined with a range of FAP-4-1BBL concentrations was evaluated in a co-culture Incucyte system with fibroblasts expressing high and low levels of FAP. Tumor cell killing was calculated as the area under the curve of the ratio between apoptotic and tumor cell count. A semi-mechanistic non-linear mixed effects (NLME) model quantifying the change in 4-1BB expression across time as a function of cibisatamab and FAP-4-1BBL concentrations was developed using Monolix 2021R1. Final model selection was based on GoF plots, AIC and precision of parameter estimates. Trimeric complex formation was explored with simulations performed in R version 4.2.1 using the RxODE package. The model explaining TC formation assumed independent and sequential FAP-4-1BBL binding to both FAP and 4-1BB [2]. Integration of this TC model together with the NLME quantifying the change in 4-1BB expression across time allowed the calculation of the number of trimeric complexes per T-cell, which was used as an independent variable to explain the observed in vitro tumor cell killing.
Results: The TC model predicts that a FAP-4-1BBL concentration of 0.37 nM would allow maximum TC formation. In line with this prediction, observed tumor cell killing as a function of FAP-4-1BBL concentration followed a bell shaped-like curve, with tumor cell killing being maximum at concentrations between 0.1 and 1 nM. In agreement with model predictions, the FAP-4-1BBL concentration at which tumor cell killing is maximum was experimentally found to be independent of FAP expression. The model describing the change in 4-1BB expression over time consisted of seven transit compartments to model the delay between cibisatamab exposure inducing signaling for 4-1BB expression and actual 4-1BB expression, with a mean transit time (MTT) of 28.6 hours. The relationship between cibisatamab exposure and 4-1BB expression signaling was accounted for with an Emax model, with an estimated EC50 at 16 nM of cibisatamab. A time after which the expression signaling stops was estimated to account for the observed decrease in 4-1BB expression after 48 hours, and was found to be independent from cibisatamab or FAP-4-1BBL exposure. The tumor cell killing followed an Emax-like relationship versus FAP-4-1BBL TC formation, with a maximum estimated 4.4 fold increase compared to cibisatamab monotherapy. 3.91x10-3 TCs formed on 4-1BB receptors per T-cell are needed to achieve 50% of this maximum tumor cell killing increase. The joint model identified that trimeric complex formation, and not FAP expression or FAP-4-1BBL exposure, is the driver of in vitro tumor cell killing when combining cibisatamab with FAP-4-1BBL.
Conclusions: Combining an in vitro system with mathematical modeling allowed establishing the relationship between tumor cell killing and TC formation with the immune co-stimulator FAP-4-1-BBL. Our model predicts that a certain increase in in vitro tumor cell killing with this type of molecule can be expected only if enough TC is formed, with this TC not only depending on FAP-4-1BBL concentration, but also on target expression. This methodology can support dose selection for bispecific clinical development, as well as informing on patient characteristics (such as receptor expression) which may be predictive of clinical benefit.
References:
[1] Claus C, Ferrara C, Xu W, Sam J, Lang S, Uhlenbrock F, et al. Tumor-targeted 4-1BB agonists for combination with T cell bispecific antibodies as off-the-shelf therapy. Sci Transl Med. 2019;11(496).
[2] Betts A, van der Graaf PH. Mechanistic Quantitative Pharmacology Strategies for the Early Clinical Development of Bispecific Antibodies in Oncology. Clin Pharmacol Ther. 2020;108(3):528-41.
[3] Van De Vyver AJ, Weinzierl T, Eigenmann MJ, Frances N, Herter S, Buser RB, et al. Predicting Tumor Killing and T-Cell Activation by T-Cell Bispecific Antibodies as a Function of Target Expression: Combining In Vitro Experiments with Systems Modeling. Mol Cancer Ther. 2021;20(2):357-66.
[4] Bacac M, Fauti T, Sam J, Colombetti S, Weinzierl T, Ouaret D, et al. A Novel Carcinoembryonic Antigen T-Cell Bispecific Antibody (CEA TCB) for the Treatment of Solid Tumors. Clin Cancer Res. 2016;22(13):3286-97.