MGL-3196,aβ-Selective Thyroid HormoneReceptor (THR)Agonist,DemonstratesMetabolic,Anti-inflammatory andAnti-fibrotic Benefits inaLong-term HighFatDiet(HFD)MouseNASHModelRebeccaTaub,JohnFranc,MarthaKelly;MadrigalPharmaceuticals,Villanova,PA

INTRODUCTION RESULTSThe hepatic THR-β receptormediates the beneficial effects ofthyroid hormone on LDL-cholesteroland triglycerides, fatty liver andinsulin sensitivity. Studies show thathepatic hypothyroidism is present inhuman NASH. MGL-3196 is a liver-directed, oral, selective THR-βagonist in Phase 2 for treatment ofNASH. MGL-3196 is highly proteinbound (>99%) with high liver uptakeand low extrahepatic permeability.In C57Bl6 mice treated with 60%HFD for 38 weeks, MGL-3196reduced cholesterol and ALT withoutthe adverse effects of T3, whichshowed THRα mediated bone loss inthe 3 week treatment.

• These results confirm thatprolonged treatment of mice withHFD generates a liver gene arrayprofile consistent with activationof NASH and fibrosis pathways

• MGL-3196 potentially normalizeshepatic function in HFD animals,including restoration of normalhepatic metabolic regulationwithout impacting tissues outsidethe liver

• At human equivalent exposuresMGL-3196 appears to reverse andprevent progression of lipid,inflammatory and fibrotic markersof NASH

Mice on a HFD for 17 or 36 weekswere treated by daily oral gavage for24 h or 24 days with 0.3-3 mg/kg/day of MGL-3196. (Figure 2, 7 (panelR)) Mice on a HFD or normal chow(lean) or HFD plus oral admixture of0.1, 0.3, 1.0 or 3 mg/kg/ day MGL-3196 or 3 mg/kg/day rosiglitazonewere treated for 25 weeks (Table 1,Figures 3-6, 7 Panel L). Mice weresacrificed and liver samples wereprocessed and total RNA wasextracted. Gene array studies wereconducted using Illumina chips. Atsacrifice blood samples werecollected for clinical chemistry andlipid levels.

MGL-3196 is specifically taken up into the liver by hepatic transporters (Figure 1). Metabolic effects of MGL-3196 treatment for 24 days (Figure 2; 7R) or 25 weeks (Table 1, Figures 3-7) included elimination of steatosis,normalization of liver size with no effect on overall body weight, improved insulin sensitivity, reduction in ALT (46%), free fatty acids (30%), and cholesterol (67%). Gene expression arrays confirmed regulation of known THR-βtarget genes such as Deiodinase-1, malic enzyme and FGF-21 (Figures 3). Metabolic genes associated with lipotoxicity such as acetyl CoA-carboxylase (ACC1) and 11-β Hydroxysteroid Dehydrogenase (HSD) were alsonormalized. In 25 week studies (Fig. 7, panel L) inflammatory (chemokine ligand 2 (CCL2), other chemokines) and major fibrosis associated genes (tissue inhibitor metalloproteinase (TIMP1), smooth muscle actin (SMA),connective tissue growth factor (CTGF) and collagen genes), were significantly increased in HFD as compared with lean controls and were normalized to the level of the lean control mice by treatment with human-relevantdoses (0.3-1 mg/kg) MGL-3196 but not rosiglitazone which had a modest effect (Figure 7). In 24 day treatment of HFD mice (Fig. 7, panel R), a similar reduction of NASH inflammatory and fibrosis transcripts was observed.

• J. Med Chem. 2014;57(10):3912-3923• Thyroid hormone mediated

autophagy and mitochondrialturnover in NAFLD, Sinha & Yen, CellBiosci (2016) 6:46

CONCLUSIONS

MATERIAL&METHODS

DISCLOSURES

Rebecca Taub, M.D., Chief Medical Officer, Executive Vice PresidentR&D. Madrigal Pharmaceuticals. becky@madrigalpharma.comContactinformation

RebeccaTaub- ManagementPosition:MadrigalPharmaceuticals

JohnFranc– MadrigalPharmaceuticals

MarthaKelly- MadrigalPharmaceuticals

REFERENCES

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To demonstrate the liver-directedpharmacologic effects of MGL-3196in a mouse NASH model.

AIM

Table 1 MGL-3196 Liver Specific, Durable Fat Reduction, Metabolic Effects

CFB, change from baseline; EE, energy expenditure; HFD, high fat diet; ITT, insulin tolerance test, ND: not done, value for 30 min shown, mice became hypoglycemic at 120 min (seizures); Rosi, rosiglitazone. Food intake and energy expenditure were percent difference relative to HFD control. No statistics were performed for food intake, because food intake was measured by group, not individual animal. #, p≤.001; ς, p≤.01; ¶, p≤.05

Assessment Chow Control

HFD Control

MGL-3196 Rosi3mg/kg0.1 mg/kg 0.3 mg/kg 1 mg/kg 3 mg/kg

Insulin (ug/L) .27±.09ς 3.24±3.21 2.46±2.58ς 1.25±.60 1.09±1.5 .39±.26¶ .39±.22¶

Glucose (g/dL) 250±47.3 269±39 270±25.7 299±32.2 266±37.9 212±28.7ς 270±27.3

Hgb A1C (%) 2.8±.23# 3.2±.18 3.1±.22 3.1±.17 3.0±.32 2.8±.27ς 3.2±.24

ITT (glucose CFB) @120 ‘ -14.3±9.9 -8.1±10.8 -17.7±18.0 -32.8±22.3ς -42.4±12.1# -73.2±10.9#

ND -31.0±19.8ς

ALT (U/L) 53±12 85±57 55±37 46±11¶ 54±50 58±22 49±19

Cholesterol (mg/dL) 87±11.8# 219±26.4 170±45.5ς 117±19.8# 96±20.2# 72±23.9# 147±13.5#

Triglycerides (mg/dL) 132±34.8 127±49.2 118±45.4 91±31.8 101±50.2 83±24.5¶ 104±36.9

FFA (mmol/L) 2.15±.60 1.91±.25 1.80±.34 1.33±.36# 1.76±.52 1.62±.41 1.36±.23#

Body Weight (g) 30.1±2.5# 49.7±2.1 46.8±9.8 45.6±5.5ς 41.0±4.9# 37.0±6.1# 51.0±7.8

Food Intake (% Control) 37.5 2.2 19.6 27.7 30.6 -1.4

EE (% control) -10.64ς 6.58 5.67¶ 16.9# 26.9# 4.24

Liver Weight (%BW) 4.7±.7 4.4±.7 3.8±.7 3.1±.2# 3.3±.16# 3.3±.4# 3.4±.3ς

Fat Mass (g) 5.2±.5 20.8±3.9 20.1±3.4 20.0±5.1 20.1±3.8 13.0±1.8ς 28.7±3.7#

Lean Mass (g) 26.3±1.1 27.4±2.0 26.2±2 26.3±1.7 25.7±1.0 24.6±2.2¶ 25.0±.8¶

Figure 6. Liver-specific Fat Reduction

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• 0.3mg/kgexposureequivalenttohuman80mgdose• Liverweightnormalizedat0.3mg/kgwithnoeffecton

bodyweight-animalsremainobese

Figure 5.

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Liver Histology in 36 WeekHFD Mice Treated with MGL-3196

• 24daytreatmentwithMGL-3196normalizedliverhistologyatalldoses(.3-10mg/kg)

• ConsistentwithTHR-βeffectstoincreasefatoxidation• Nodetectablesteatosis,fibrosisorinflammationinMGL-3196treatedlivers

Figure 1. RadiographicTissueDistribution-Rats

Rat14CADMEstudy(Ref1) • MGL-3196uptakeinliver

• Lowtoundetectablelevelinheart,bone,brain

• Primarilyfecaleliminationviabiliaryexcretion

Figure 4. Normalization ofHepatic Gene Expression• 25weektreatmentwithHFDchangesthe

hepaticexpressionofalargenumberofgenetranscripts,includingbothincreased(red)anddecreased(blue)expression

• TreatmentwithMGL-3196normalizeshepaticgeneexpressionaswellashepaticarchitectureandsize

HFD 0.1 0.3 1 3 Rosi MGL-3196 (mg/kg)

Figure 2. Improvement in Liver Health

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• 24dtreatmentMGL-3196andT3reduceALT• MGL-3196alsoreducesliversizeandlivertriglycerides

Figure 7. MGL-3196 Reduces NASH & Fibrosisand Increases Mitophagy Pathway Genes

“HFD”, lane 1 mean HFD gene expression normalized to mean Lean; Lanes (2-7) mean gene expression normalized to mean of HFD; “Rosi” (rosiglitazone, 3 mg/kg, 25 weeks); lanes 8-10, treatment 24 d MGL-3196 or Rosi (10 mg/kg), normalized to HFD Control; Red, higher expression; blue decreased expression

Inflammatory HFDLean 0.1 0.3 1 3 Rosi 0.3 1 3 RosiMCP-1/CCL2MIP-2α/CXCL2MIP-2ß/CXLCL3A20/TNFaip3CRPAnnexin 2SAA1FibrosisTGFbetaOsteopontinHyaluron synCollagen 1Galectin-3TIMP1Collagen 4a2SMACollagen 4a1CTGFKeratin 18Collagen 3Galectin-1Mitophagy-Ulk-1

25weektreatmentMGL-3196(mg/kg) MGL-3196

24daytreatment

• NormalizationofNASH-relatedtranscriptstolevelofleancontrol

• StimulationofmitophagyviaULK1inductionbyTHR-βleadstomitochondrialbiogenesiswhichprotectsagainstNASH-relatedROS,lipotoxicity

Figure 3.

DIO1,deiodinase1;TBG,thyroidbindingglobulin(SerpinA);mGPDH,mitochondrialglycerol-3-phosphatedehydrogenase

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MGL-3196 Regulates Metabolicand THR-β Responsive Genes