Preclinical characterization of LY3484356, a novel, potent


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Preclinical characterization of LY3484356, a novel, potent and orally available selective estrogen receptor degrader (SERD)
Presented at: AACR Annual Meeting 2021 Date: April 10, 2021

Preclinical characterization of LY3484356, a novel, potent and orally bioavailable selective estrogen receptor degrader (SERD)

Shripad V. Bhagwat1, Baohui Zhao1, Weihua Shen1, Cecilia Mur2, Robert Barr2, Lisa J. Kindler1, Almudena Rubio2, Jolie A. Bastian2, Jeffrey D. Cohen2, Brian E. Mattioni2, Eunice Yuen2, Thomas K. Baker2, Mark A. Castanares2, Dongling Fei2, Jason R. Manro2, Maria Jose Lallena2, Sheng-Bin Peng1, Alfonso De Dios2

#1236

1Loxo Oncology at Lilly, 2Eli Lilly and Company, Indianapolis, IN

Background

• Nearly 70% of newly diagnosed breast cancers are
estrogen receptor alpha (ERα) positive, for which endocrine therapy (ET) is the mainstay of treatment.1
• Fulvestrant, the only approved SERD, is administered via
intramuscular injection and as a result, is limited by suboptimal systemic pharmacology, as well as patient administration challenges.2
• Additionally, approximately 40% of patients develop
resistance to ET through mutations in ERα (ESR1) that drive constitutive activation of ERα.3
• Novel degraders and antagonists of ERα have been
developed, to deliver more ERα target coverage, more convenient dosing, and overcome ERα-mediated acquired resistance.4
• Here we describe the preclinical profile of LY3484356, a
novel oral SERD and pure ERα antagonist, with potent activity against the wild type and mutant ERα.
Table 1. Biochemical and cellular potency of LY3484356

Assay

LY3484356

Wild type ERα binding, Ki, nM

0.31

Y537S ERα binding, Ki, nM

2.79

MCF7 cell ERα wild type degradation, IC50, nM

3

MCF7 cell ERα Y537N degradation, IC50, nM

9.6

MCF7 cell ERα wild type antagonism, IC50, nM

41

MCF7 cell ERα Y537N antagonism, IC50, nM

13

MCF7 cell PRα agonism, IC50, nM

>2000

MCF7 cell ERα wild type proliferation, IC50, nM

3

MCF7 cell ERα Y537N proliferation, IC50, nM

17

IC50 values are averages from replicate assays
Assays
• Degradation/Antagonism: Cells were treated with various concentrations of LY3484356 or fulvestrant in phenol red free DMEM containing 10% charcoal stripped FBS and incubated for 24h. ̶ ERα degradation was measured using high content imaging assay ̶ The antagonistic activity was measured by measuring PRα inhibition by high content imaging assay

• Proliferation: Cells were treated with various concentrations of LY3484356 or fulvestrant in phenol red free DMEM or RPMI containing 10% FBS and incubated for 6 or 7 days. Cell number was measured using high content imaging assay.

AACR Annual Meeting, Virtual, April 10-15, 2021

TTuummoorrVVoolluummee (mm33)

TTuummoorr Volummee((mmm3m)3)

Gene Expression Inhibition Gene Expr(e%ssiCoonntIrnolh)ibition
(% Control)

Fig 1. LY3484356 shows potent and sustained ERα signaling modulation in vivo

Fig 2 (cont). LY3484356 demonstrates in vivo efficacy in ERα WT and mutant tumor models

Fig 4. LY3484356 shows enhanced in vivo efficacy in combination with alpelisib or with everolimus

100

50

0

-50
trol Con

PGR DZK1 P

WISP2 SGRP1 GREB1

AREG

RA

24h24ahfteafrtelar slatsdt doosseeooff LY348L4Y3354864, 31506,m10pmk,pQk,DQxD3x, P3 O

100

50

0

-50
3 mpk

10 mpk

30 mpk

LLYY3344884435365,6Q, DQxD3x,3P,OPO

MCF7 xenografts were treated with LY3484356, and plasma and tumor were harvested at indicated timepoints. Inhibition of gene expression was measured by qPCR. GAPDH was used as the control. Data are mean ± SEM.
Control 4h 12h 24h 48h 96h 168h

TTuummoorrVVoolluummee ((mmmm3)3)

TTuummoorrVVoolluummee ((mmmm3)3)

C
600

TT4477DD EESSRR11 WWTT

400 200
0
D
2100

* ****

47

54

61

68

TTrereaattmmeenntt DDaayyss

ST941HI ESR1 Y537S ST941HI ESR1 Y537S

1400 700 0 0

ns ns
****
4 8 12 16 20 24 28
TTrreeaattmment DDaayyss

Vehicle Control, QD x 28, PO LY3484356, 3 mpk, QD x 28, PO LY3484356, 10 mpk, QD x 28, PO
Fulvestrant, 5 mg/dose, Q7D x 4, SC
T47D ESR1 WT cells were implanted in NSG mice with estrogen pellets and ST941HI ESR1 Y537S mutant PDX tumor fragments were implanted in athymic nude mice with no estrogen pellets. All treatments were tolerated. Data are mean ± SEM. *p<0.01, **p<0.001, ns – not significant, treatment vs vehicle control.
Vehicle Control, QD x 28, PO LY3484356, 3 mpk, QD x 28, PO LY3484356, 10 mpk, QD x 28, PO LY3484356, 30 mpk, QD x 28, PO Fulvestrant, 5 mg/dose, Q7D x 4, SC

PPGGRR IInnhhiibbiittiioonn ((%%CCoontnrtrol)ol)

Fig 2. LY3484356 demonstrates in vivo efficacy in ERα WT and mutant tumor models

Fig 3. LY3484356 shows enhanced in vivo efficacy in combination with abemaciclib

A
750

MCMFC7FE7SERS1RW1TWT

500

250

****

0

50

60

70

80

90

TrTeraeatmtmeennttDDaayyss

B
1000

ZZRR-7-755-1-1EESSRR1 1WWT T

750

500

250

****

0 21 28 35 42 49 56 63 70
TTrreeaattmmeenntt DDaayyss

Vehicle Control, QD x 42, PO LY3484356, 10 mpk, QD x 42, PO Fulvestrant, 5 mg/dose, Q7D x 6, SC
MCF7 and ZR-75-1 ESR1 WT cells were implanted in NOD SCID mice with estrogen pellets or estradiol valerate injections. All treatments were tolerated. Data are mean ± SEM. **p<0.001, treatment vs vehicle control.
Vehicle Control, QD x 42, PO LY3484356, 3 mpk, QD x 42, PO LY3484356, 10 mpk, QD x 42, PO Fulvestrant, 5 mg/dose, Q7D x 6, SC

TTuummoorrVVoolluumee ((mmm3m)3)

TTuummoorrVVoolluummee((mmm3m)3)

A

TT4477DD EESSRR11 WWTT

800

600

400
200
0 51

** ** **

58

65

72

79

TTrereaatmtmeennttDDaayyss

B
1600

SSTT994411PPBBRRESERS1RY153Y75S37S ((ppaalbbooccicicliblibresreisstaisnt)ant)

1200

800

400

0

0

7

14

21

TreTartematemnetnDt Dayayss

Vehicle Control, QD x 28, PO LY3484356, 5 mpk, QD x 28, PO Abemaciclib, 20 mpk, QD x 28, PO Combo (LY3484356, 5 mpk, QD x 28, PO + abemaciclib, 20 mpk, QD x 28, PO)
T47D ESR1 WT cells were implanted in NSG mice with estrogen pellets. ST941PBR ESR1 Y537S PDX tumor fragments were implanted in athymic nude mice. ST941PBR ESR1 Y537S is a palbociclib resistant PDX model rederived in mice. All treatments were tolerated. Data are mean ± SEM, except where n=1. **p<0.001, treatment vs vehicle control.
Vehicle Control, QD x 28, PO (N=8)
Combo (fulvestrant, 5 mg/dose, Q7D x 4, SC + abemaciclib, 50 mpk, QD x 28, PO) (N=1)
LY3484356, 10 mpk, QD x 28, PO (N=1) Combo (LY3484356, 10 mpk, QD x 28, PO + abemaciclib, 50 mpk, QD x 28, PO) (N=1)
Tamoxifen, 0.1 mpk, IP, QD x 5 days/week x 4 weeks (N=1)

Presenter: Shripad Bhagwat, [email protected]

A MMCCF7FE7SERS1RW1T,WPIT3,KPAI3EK54A5KE4545K4
600

Tuummoorr VVoolluummee((mmmm33))

400

200

*

** **
0 32 39 46 53 60 67 74

TrTeraeatmtmeennt DDaayyss

B MMCCFF77ESERS1RW1 TW, TP,I3PKI3AKEA54E55K445K4
600

Vehicle Control, QD x 42, PO
LY3484356, 15 mpk, QD x 42, PO
Alpelisib, 25 mpk (d1-7), 10 mpk (d8-42), QD x 42, PO
Combo (LY3484356, 15 mpk, QD x 42, PO + alpelisib, 25 mpk (d1-7), 10 mpk (d8-42), QD x 42, PO)
MCF7 ESR1 WT cells were implanted in NOD SCID with estrogen pellets. All treatments were tolerated. Data are mean ± SEM. *p<0.01, **p<0.001, treatment vs vehicle control.

TTumor Voolluummee ((mmmm33))

400
200
**** **
0 32 39 46 53 60 67 74
TrTereaattmmennttDDaaysys

Vehicle Control, QD x 42, PO
LY3484356, 15 mpk, QD x 42, PO
Everolimus, 5 mpk, QD x 42, PO
Combo (LY3484356, 15 mpk, QD x 42, PO + everolimus, 5 mpk, QD x 42, PO)

Conclusions
• LY3484356, an oral SERD and a potent degrader of WT
and mutant ERα, is a pure antagonist of ERα with no tissue-specific agonism in the uterus. LY3484356:
‒ Inhibits ERα-mediated gene transcription as shown by
sustained inhibition of a set of ERα-dependent genes
‒ Demonstrates in vivo efficacy in ESR1 WT and mutant
models
‒ Demonstrates enhanced in vivo efficacy in
combination with abemaciclib, alpelisib and everolimus in xenograft models
• The first-in-human Phase 1/2 clinical trial of LY3484356
(EMBER, NCT04188548) is ongoing. A window-ofopportunity clinical trial (EMBER-2, NCT04647487) evaluating the pharmacodynamic effects of LY3484356 in early-stage breast cancer patients will begin in the first half of 2021.
References
1. Jordan VC. Nat Rev Cancer 2007; 7:46–53 2. van Kruchten M et al Cancer Discov 2015;5:72–81 3. Jeselsohn R et al Clin Cancer Res 2014;20:1757–67 4. Liu S, et al Invest New Drugs 2018;36:763-772
Sponsored by Loxo Oncology at Lilly

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Preclinical characterization of LY3484356, a novel, potent