Clinical translatability of Ldlr-/-.Leiden mouse model for NASH
Eveline Gart1, José A. Inia1, 2, 3, Martine M.C. Morrison1, Robert Kleemann1, Anita M. van den Hoek1
1 Department of Metabolic Health Research, TNO Leiden, The Netherlands. 2 Department of Cardiology, Leiden University Medical Center, Leiden, The Netherlands. 3 Einthoven Laboratoryfor Experimental Vascular Medicine, Leiden University medical Center, Leiden, The Netherlands. 4 Human and Animal Physiology, Wageningen University, Wageningen, The Netherlands

Contact information: a.vandenhoek@tno.nl

The present study aimed to further investigate thetranslatability of Ldlr-/-.Leiden mouse as translational
NASH model.

MethodLdlr-/-.Leiden mice fed a translational energy dense highfat diet without cholesterol supplementation for 20-24
weeks were compared with human NASH patients on (a)obesity and insulin resistance; (b) lipoprotein profiles,endogenous cholesterol synthesis and de novo
lipogenesis (DNL); (c) liver transcriptome andmetabolome profile; (d) histological NASH endpoints and(e) atherosclerosis. In addition, the response totreatments for 10 weeks with 10 mg/kg/d obeticholic acid(OCA) or 20 mg/kg/d cenicriviroc via diet admix or 12weeks with 0.12 mg/kg/d semaglutide via sc injections
was analyzed.

Diet-induced obesity, hyperinsulinemia
and dyslipidemia in Ldlr-/-.Leiden miceAt t=24 weeks:

ChowHFD

Body weight (g)36.7 ± 1.6

51.8 ± 1.0***

Blood glucose (mM)

6.7 ± 0.3

6.9 ± 0.3

Plasma insulin (ng/mL)

2.0 ± 0.3

20.7 ± 4.3***

HOMA-IR

0.6 ± 0.1

6.3 ± 1.4***

Plasma cholesterol (mM)

7.6 ± 0.6

33.2 ± 2.7***

Plasma TG (mM)

1.3 ± 0.1

4.8 ± 0.8****** P<0.001 vs chow

- Ldlr-/-.Leiden mice have an active endogenous cholesterol synthesis asin humans (measured via balance analysis).

A- DNL is upregulated (14C fromacetate is incorporated into
newly synthesized fatty acids.
Fatty acid profiling (A) via
LC/MS showed that 14C signal
was predominantly found in
palmitate (C16:0, the primary
end-point of DNL).

1.5A*** P<0.001 vs controln≥10Control group set at 11.0***
0.5******Healthy Control

BLdlr-/-.Leiden mice recapitulate human NASH
transcriptome and metabolome profiles

AMacrovesicular steatosis

0.0A) A high percentage of human NASH pathways isrecapitulated in Ldlr-/-.Leiden mice on a high fat diet
(HFD, 73%) or fast food diet (FFD, 80%).
B) Furthermore, Ldlr-/-.Leiden serum metabolome isreflected in a substantial proportion of NASH
patients.

B

Ldlr-/-.Leiden mice develop NASH with bridging fibrosis
and atherosclerosis
AB
After 24 wks on HFD Ldlr-/.Leiden mice develop (A)NASH with macrovesicularsteatosis (arrows) andfibrosis (Sirius red staining)and (B) atherosclerotic
plaques (arrows).

Ceni

Sema

p=0.054

1.5

***

1.0
0.50.0******
Healthy Control

COCA

Hepatic inflammation

2.0

Collagen/protein (g/mg)

Aim

Ldlr-/-.Leiden mice respond to NASH
treatments in line with clinical trials
Macrovesicular steatosis(% of hepatocytes)

NASH is one of the most prevalent chronic liverdiseases, which is closely associated with obesity,
insulin resistance, dyslipidemia and cardiovasculardisease. Preclinical validation of novel drug candidates
for the treatment of NASH requires a translational animalmodel that should recapitulate these hallmarks of thehuman disease.

Ldlr-/-.Leiden mice have an active endogenous cholesterolsynthesis and DNL (as in NASH patients)

Hepatic inflammatory
aggregates/mm2

Introduction

1.5

OCA

Ceni

Sema

Hepatic collagen

1.0

p=0.08

0.5

***
0.0
Healthy Control

OCA

Ceni

Sema

Ldlr-/-.Leiden mice respond to treatments with NASH
resolution (OCA & semaglutide, not cenicriviroc) andimprovement with fibrosis (OCA tendency only), in line withclinical trials.

Conclusions
Overall, the Ldlr-/-.Leiden mouse model shows goodclinical translatability and accurately mimics the etiology
and pathology of NASH and fibrosis in humans.