Cerevel Therapeutics : Evaluation of M4 Muscarinic Receptor Occupancy by CVL-231 in Nonhuman Primates

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Evaluation of M4 Muscarinic Receptor Occupancy by CVL-231 Using [11C]MK-6884 PET in Nonhuman Primates

Sridhar Duvvuri,1 Philip Iredale,1 Matthew Leoni,1

John M. Kane,2 Vasily Belov,3 Nicolas J. Guehl,3 Sung-Hyun Moon,3 Maeva Dhaynaut,3 Peter A. Rice,3 Daniel L. Yokell,3 Georges El Fakhri,3 Marc D. Normandin,3 John Renger1

1Cerevel Therapeutics, Cambridge, MA, USA; 2Zucker Hillside Hospital, Hempstead, NY, USA; 3Gordon Center for Medical Imaging, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA

INTRODUCTION

• CVL-231is a novel, brain-penetrant, positive allosteric modulator selective for M4 muscarinic acetylcholine receptors (mAChRs) in development for the treatment of schizophrenia
• Preclinical characterization of CVL-231 in rodents showed favorable brain penetration, direct target engagement, and robust in vivo activity in animal models of psychosis (eg, reversal of amphetamine-stimulated locomotor activity, prepulse inhibition)1
• Verification of in vivo target engagement in primate brains and quantification of the exposure-occupancy relationship is useful to facilitate clinical dose selection and translation of preclinical data to humans

OBJECTIVE

• Using [11C]MK-6884, an M4 positive allosteric modulator radioligand, this study evaluated M4 receptor occupancy (RO) of CVL-231 in the striatum of nonhuman primates as a function of CVL-231 dose and plasma concentration
• • The first objective was to determine the RO at M4 mAChRs in the striatum using arterial input function-based pharmacokinetic (PK) modeling methods following different intravenous doses of CVL-231
  • The second objective was to assess the accuracy of reference tissue-based PK modeling methods for quantifying the RO of CVL-231

METHODS (CONTINUED)

• Three-dimensional,T-1 weighted, magnetization-prepared rapid gradient-echo (MPRAGE) magnetic resonance images were also acquired for each monkey using a 3T Biograph mMR scanner (Siemens Medical Solutions USA, Inc, Malvern, PA) for anatomical reference
• Methods for arterial blood sampling are presented in the Table

Figure 1. Diagram of a typical testing design for a single dose of CVL-231.

Tracer washout

[11C]MK-6884		[11C]MK-6884
bolus 1	CVL-231 bolus		bolus 2
		and infusion				Post-treatment recovery

-2 h	0 h	1.5 h	2.8 h 3 h	4.5 h	7 d	>30 d
	NHP sedation			Baseline		Blocking		Postprocedural
	and preparation		PET/CT scan		PET/CT scan	observation

Table. Arterial Blood Sampling Methods for PET/CT Scanning Sessions

• • VT was calculated as K1/k2 for a one-tissue compartment model and modeled as (K1/k2) × (1 + k3/k4) for a two-tissue model
  • K1 and k2 are the rate constants for tracer influx and efflux, respectively, in the tissue with respect to plasma; k3 and k4 are the rate constants for specific binding and dissociation from the receptors in the target tissue, respectively
  • VT was also calculated using graphical methods with arterial input functions, such as Logan plot and multilinear analysis 1 (MA1)
• Nondisplaceable binding potential (BPND) was defined as the equilibrium ratio of the concentration of specifically bound radioligand to its nondisplaceable concentration (nonspecifically bound and free in tissue)
• • BPND was a direct outcome of the simplified reference tissue model (SRTM)
  • Logan DVR and MRTM2 methods were also tested and provided distribution volume ratio (DVR) as a direct outcome: DVR = VTtarget region/VTreference tissue. BPND was then calculated as BPND = DVR - 1
• RO was calculated either as the slope of the Lassen plot VTbaseline - VTblocking = RO ×
  (VTbaseline - VND) or as %RO = 100% × (1 - BPNDblocking/BPNDbaseline), where VTbaseline and VTblocking
  are the regional Vt at baseline and drug challenge (blocking) conditions, BPNDbaseline and
  BPNDblocking are the BPND in the target region at the same conditions, and VND is the volume of distribution of nondisplaceable uptake

Presenting Author: Sridhar Duvvuri; [e-mail]

CONCLUSIONS

Robust quantification of CVL-231 RO in the striatum was obtained via [11C]MK-6884 PET imaging and by using

METHODS

STUDY DESIGN

● Two male adult rhesus macaques aged 9 and 13 years were used in this study; their mean

body weights on day of imaging were 12.9 kg and 15.1 kg, respectively

● Both animals had 3 imaging sessions, each consisting of a 90-min baseline scan followed

by a 90-min positron emission tomography (PET)/computed tomography (CT) blocking

scan with CVL-231 administration; each session was separated by >1 month to allow for

sufficient recovery (Figure 1)

- Before each imaging session, animals were sedated with ketamine/xylazine (10/0.5 mg/kg

Baseline PET/CT

• Arterial blood sampling was performed during each dynamic PET acquisition
• • Samples­ of 1-3 mL were initially drawn every 30 s after the radiotracer injection and decreased in frequency to every 15 min toward the end of the scan
• [11C]MK-6884 metabolism was characterized from blood samples collected at 5, 8, 10, 15, 30, 60, and 90 min

Blocking PET/CT

• CVL-231was administered intravenously as a loading dose (~48% of the total dose by bolus, 10 minutes before radiotracer) followed by a maintenance dose (~52% of the total dose continuously infused until the end of scan); doses ranged from 0.25 mg/kg to 1.7 mg/kg
• • Arterial­ blood samples for determination of CVL-231 concentration were collected at 60 min after radiotracer injection to determine plasma levels of CVL-231 for doses <1.7 mg/kg
  • For­ doses of 1.7 mg/kg, arterial blood samples were drawn at 0, 30, 60, and 90 min after radiotracer injection

STATISTICAL ANALYSES

● Regional brain PET data were analyzed by various PK modeling techniques using blood-

based and reference tissue (cerebellar gray matter) input functions to quantify radiotracer

binding and to calculate RO at different doses of CVL-231

● Goodness of model fits, time stability of measured outcomes, and agreement between

models were evaluated to assess the performance of the models

● A brain tissue suitable for use as a reference region was assessed based on the analysis of

VND derived by Lassen plot method

● Estimates of RO were analyzed in a dose-response fashion against injected CVL-231 dose

and plasma concentration

noninvasive pharmacokinetic modeling techniques

These data confirm the dose-dependent target binding of CVL-231 to M4 receptors in the striatum of nonhuman primates

Evaluation of M4 RO by CVL-231 in humans using [11C]MK-6884 is being explored

intramuscularly) and were intubated for maintenance anesthesia with isoflurane

- Imaging sessions were performed on a Discovery MI (GE Healthcare, Chicago, IL)

PET/CT scanner. A CT scan was acquired prior to each PET acquisition for attenuation

correction. Emission PET data were acquired in three-dimensional list mode for 90 min

following injection of [11C]MK-6884

RESULTS

CT, computed tomography; PET, positron emission tomography.

PRIMARY OUTCOMES

• The total volume of distribution (VT) was assessed, representing the equilibrium ratio of tracer concentration in tissue relative to its plasma concentration, which is linearly related to the tracer binding to the target

- Data were fitted by the function RO = ROMAX × D / (D + ID50), where D is the drug dose,

ID50 is the estimated drug dose needed to achieve half-maximal occupancy, and ROMAX is

the estimated maximum RO that can be asymptotically attained by a high level of drug

- An analogous fitting procedure was performed with the function RO = ROMAX × C / (C +

IC50), where C is the steady-state plasma concentration (ng/mL) at 1 hour, and IC50 is the

estimated plasma concentration needed to achieve half-maximal occupancy

[11C]MK-6884 BRAIN UPTAKE AND TIME-ACTIVITY CURVES (TACS)

● PET images demonstrated high brain penetration 0-10 min after radiotracer injection, with

● Among evaluated regions of interest (caudate, cerebellum,

cortical gray matter, hippocampus, putamen, central

Figure 3. 2T4K estimates of regional VT (mL/cc) for all studies for (A) monkey 1 and (B) monkey 2.

baseline standard uptake value (SUV) levels in the striatum exceeding 4.2; at the dynamic

equilibrium phase (30-90 min), the highest baseline SUV levels in the striatum reached 1.4,

in contrast with neighboring regions (SUV ~0.5)

● A strong dose-dependent blocking effect of CVL-231 was observed on [11C]M6884 binding

in the striatum (Figure 2)

Figure 2. Individual MRI MEMPRAGE images and [11C]MK-6884 PET SUV images (summed 30-90 min after tracer injection) at baseline and after blocking at different CVL-231 doses in the brains of monkey 1 (A) and monkey 2 (B).

A. Monkey 1	B. Monkey 2
SUV30-90min	SUV30-90min
1.4	1.4

white matter), only the caudate and putamen displayed

significant blockade of [11C]MK-6884 by CVL-231

(Figure 3)

SIMPLIFIED REFERENCE TISSUE MODELS

● All evaluated reference-tissue methods (simplified

reference tissue model [SRTM], multilinear tissue reference

model 2 [MRTM2] and Logan distribution volume

ratio [DVR]) demonstrated a very strong agreement in

quantifying regional BPND using cerebellar grey matter as

a reference region; due to consistently high performance,

the SRTM method was selected for the final quantification

of RO

● Using cerebellar grey matter as a reference tissue, striatal

RO was dose dependent from 18% to 67% over the range

of evaluated CVL-231 doses (0.25 to 1.7 mg/kg, total of

A 5
4
3
2
1
0	nucleus	Cerebellum	Cortex	Hippocampus
Caudate		matter)
	(grey

Baseline 1

2T4K, 2-tissue4-parameter; VT, volume of distribution.

B 5

4

3

2

1

0

Putamen	Striatum	White	matter	nucleus	Cerebellum	Cortex	Hippocampus	Putamen	Striatum	White	matter
		matter)
			Caudate		(grey
0.25 mg/kg		Baseline 2	0.5 mg/kg	Baseline 3	1.7 mg/kg

ACKNOWLEDGMENTS: Editorial/Medical writing support was provided under the direction of the authors by Alexandra Kennedy, PhD and Emilia Raszkiewicz, ELS at MedThink SciCom and was funded by Cerevel Therapeutics.

REFERENCES: 1. Iredale et al. Presented at: Society for Neuroscience 2021, November 3-7, 2021; Virtual.

Baseline					Baseline
0.25 mg/kg					0.5 mg/kg
mg/kg				0	mg/kg				0
0.5				1.0
1.7 mg/kg					1.7 mg/kg

loading and maintenance components)

● The respective plasma concentrations of CVL-231 ranged

from 126 ng/mL to 1040 ng/mL

- The relationship of striatal RO with CVL-231-injected

dose and plasma concentration was described by the

classical Hill dose-response function, with an ID50 of

1.1 ± 0.1 mg/kg and an IC50 of 581 ± 55 ng/mL (Figure 4)

Figure 4. (A) CVL-231mass-dose response and (B) plasma-exposure response.

Presented at the American College of Neuropsychopharmacology Annual Meeting December 5-8, 2021 • San Juan, Puerto Rico

Images are presented in the MRI NIMH macaque template space. Sagittal, coronal, and transverse views are centered on the putamen. MEMPRAGE, multiecho magnetization prepared rapid acquisition gradient echo; MRI, magnetic resonance imaging; NIMH, National Institute of Mental Health; PET, positron emission tomography; SUV, standard uptake value.

ROMAX is set to 1. R2=0.96. IC50, plasma concentration needed to achieve half-maximal occupancy; ID50, drug dose needed to achieve half-maximal occupancy; SRTM, simplified reference tissue model; RO, receptor occupancy; ROMAX, maximum receptor occupancy.