A009: A Translational Platform Quantitative Systems Pharmacology Model for Preclinical to Clinical Translation of In Vivo CRISPR-Cas9 Gene Therapy

Statement of Purpose, Innovation or Hypothesis: In-vivo CRISPR Cas genome editing is a complex therapy including three components: 1) Lipid Nano Particle (LNP); 2) Messenger RNA (mRNA); and 3) Single guide RNA (sgRNA). This next generation of gene editing comes into a class of novel modalities where little is known about their dose-exposure relationship and biodistribution. Within this work we have built a QSP model to characterize, predict and translate the Pharmacokinetics/Pharmacodynamics (PK/PD) of CRISPR therapies from preclinical species (mouse, NHP) to clinic (humans) using two case studies.

Description of Methods & Materials: PK/PD data of LNP, mRNA and sgRNA were obtained from the literature for both case studies for transthyretin amyloidosis and reduction of LDL-cholestrol^1,3. The QSP model includes the following mechanisms upon IV injection: 1) Binding of LNP to opsions in the liver vasculature; 2) Internalization of LNP into Mononuclear Phagocytotic System (MPS); 3) Internalization of LNP in Interstitial layer of liver via endocytosis and binding to LDL receptors via receptor mediated endocytosis as well as via macropinocytosis; 3) Internalization of the LNP into cellular layer of liver and release of transgene product i.e. mRNA and sgRNA; 4) Disposition of mRNA and sgRNA into the systems by including exocytosis as well as clathrin-mediated endocytosis and macropinocytosis; 5) Elimination of LNP and sgRNA via renal elimination; 6) Exonuclease degradation of sgRNA and mRNA; 7) Translation of mRNA into Cas9 and binding with sgRNA to form RNP complex; 8) Gene Editing on site of target protein. Monte-Carlo Simulations were performed for 1000 subjects to account for reduction in TTR.

Data & Results: The model was able to adequately capture the reported pharmacokinetics of LNP across species. The rate of internalization into the interstitial layer (kin) was 0.0379 1/h (30.8% RSE) in NHP and 0.009 1/h (0.252% RSE) in humans. The rate of exocytosis (kout) was found to be 5.01 1/h (3.91% RSE) in mouse, 2610 1/h (57.9% RSE) in NHP, and 780 1/h (0.129 %RSE). All the physiological flows and volumes were fixed from literature⁴. Pharmacodynamics was characterized using an indirect response model – I in which first order rate of degradation (kout) was estimated 0.482 1/d (13.7% RSE); Imax was estimated at 0.961 (0.574 %RSE) and IC50 was estimated to 4.9 ug/mL (5.42% RSE) for reduction in TTR for NHP. The model was able to estimate reduction of TTR in humans as well as reduction in PCSK9 and LDL Cholesterol using a feedback loop model for NHP with adequate precision.

Interpretation, Conclusion or Significance: The developed QSP model is able to characterize the biodistribution and help understand mechanism of action as well as dose-exposure relationship across species. The utility of platform QSP model can be a paramount in facilitating the discovery and development of these novel agents. As lipid nanoparticle has different structural asymmetry’s which can impact the endocytosis a quantitative structural activity pharmacokinetic model should be built in future to account for the differences in unique PK-PD properties.

Disclosures: Authors declare no conflict of interest

Citations/References: 1. Abdelhady, A. M. et al. Clin. Pharmacol. Ther. 114, 591–603 (2023). 2. Gillmore, J. D. et al. N. Engl. J. Med. 385, 493–502 (2021). 3. Finn, J. D. et al. Cell Rep. 22, 2227–2235 (2018). 4. Shah, D. K.et al. J. Pharmacokinet. Pharmacodyn. 39, 67–86 (2012).

Additional Information/Authors: Devam A. Desai1, Stephan Schmidt1, Rodrigo Cristofoletti1
1 Center of Pharmacometrics and Systems Pharmacology, University of Florida, Orlando, FL