0 2 4 6 80.01

0.1

1

10

100

-40

-20

0

20

Time (days)

Seru

m c

onc.

(nM

)FB

G (m

g/dL)

Test materialsRelative ratio (% vs. insulin)

IGF-1R/IRIR-A IR-B IGF-1RInsulin 100 100 100 1IGF-1 0.9 0.6 17,984 23,515Glargine 42 73 579 10AspB10 308 248 449 1.6Insulin 115 93 86 84 0.9

American Diabetes Association’s (ADA) 74th Scientific Sessions, San Francisco, CA, USA; June 13-17, 2014 For any questions, please contact Hanmi Pharm. Co., Ltd., Rep. of KoreaPhone: +82-31-371-5141; cjchoi@hanmi.co.kr

A Novel Very Long-Acting Insulin Analog (HM12470) with Potential for Once-Weekly Dosing, Has a Favorable PK, PD and Mitogenic Profile

SY Hwang1, IY Choi1, , JY Kim1, SY Jung1, DJ Kim1, YM Lee1, YH Kim1, M Trautmann2, M Hompesch2, JW Son1, SC Kwon1

1Hanmi Pharm. Co., Ltd., Seoul, South Korea, 2Profil Institute, Chula Vista, CA, USA

89-LB

ABSTRACTThe long-acting basal insulin, LAPSInsulin 115 (HM12470) has been

developed for once-weekly administration to provide for a better basalinsulin treatment option. The objective of this study was to investigate thein vitro properties, PK/PD of LAPSInsulin 115 in normal and diabetic animalmodels to evaluate once-weekly dosing potential, and combination withonce-weekly GLP-1R agonist. Insulin 115 displayed slightly reducedbinding affinity for IR-A, -B, and IGF-1R compared with human insulin. Inaddition, Insulin 115 triggered mitogenic signals in MCF-7, at a levelcomparable with mitogenic signals triggered by human insulin. In a PKstudy, LAPSInsulin 115 when administered subcutaneously exhibited a halflife of ~ 44 hrs in normal rats, and the extended half-life was also confirme din other species such as mice, dogs, pigs and monkeys. The improved PKprofile contributed to a more prolonged glucose lowering efficacy in db/dbmice compared with the native human insulin conjugate ( LAPSInsulin).Based on the results from three different species, the half-life of LAPSInsulin115 in humans is expected to be 132 hrs and the peak-to-trough ratio wascalculated to be 1.6 for QW dosing by the Wajima Css-MRT method. ThisQW potential of LAPSInsulin 115 is expected to harmonize very well with thePK profile of the long-acting GLP-1R agonist (LAPSCA-Exendin-4) showingpotential for a well matched QW combination therapy. LAPSInsulin 115combined with LAPSCA-Exendin-4 showed synergistic effect on HbA1creduction and neutralized body weight in db/db mice. These observationssuggest that LAPSInsulin 115 has a once-weekly dosing potential with asufficiently extended half-life and a low mitogenic risk, as well as itspromise for once-weekly insulin/GLP-1R agonist combination therapy.

BACKGROUND

METHODS

RESULTS

Hanmi Hanmi Pharm. Co., Ltd.

Table 1. Receptor binding affinities in triplicates

Insulin 115 had slightly reduced affinities on IR-A, -B, and IGF-1R, compared withhuman insulin and no preferential binding on IGF-1R or IR-A.

0 7 14 21 28 35 42 49 56 630.1

1

10

100

1000

Time (days)

Seru

m c

onc.

(nM

)

Figure 5. Predicted pharmacokinetics in human

LAPSInsulin 115, t1/2 = 132 hrs

Peak-to-Trough Ratio = 1.6

LAPSInsulin, t1/2 = 55 hrs

Peak-to-Trough Ratio = 4.0

LAPSInsulin 3.5 nmol/kgLAPSInsulin 115 3.5 nmol/kg

Pharmacokinetics in human was predicted based on PK of three different animals.LAPSInsulin 115 was expected extended half-life and lower peak-to-trough ratio thanLAPSInsulin.

REFERENCES

CONCLUSIONS In vitro binding affinity on IR and IGF-1R showed that the

mitogenic potency of Insulin 115 and LAPSInsulin 115 were notincreased compared to that of human-insulin. Extended PK profiles and prolonged glucose lowering

efficacy of LAPSInsulin 115 were demonstrated. Human PK prediction suggests that LAPSInsulin 115 can

achieve a basal insulin profille suitable for once weekly use. The combination of LAPSInsulin 115 and LAPSCA-Exendin-4

showed synergistic effects on glycemic control withsuppressed body weight gain.

1. Diabetic Medicine (2013) 30: 1293–1297.2. Diabetes (2011) 60 (Suppl. 1):LB11 (37-LB)3. PLoS ONE (2010) 5: e9540.4. PLoS ONE (2012) 7: e34274.

Key requirements for once weekly insulin

Overview of LAPSInsulin 115 as a very long-acting insulin

1. Lower peak-to-trough ratio due to long duration2. Patients’ adherence improvement by once-

weekly administration3. Ideal combination partner with weekly GLP-1

agonist

Long half-life Flat profile

- Low variability (intra/inter patients) Low risk of hypoglycemic episodes Flexible timing/dosing Little or no weight gain CV safety

Prolonged PK profile seems to be a key.

“Paradoxical Safety”Longer is safer than shorter.

Incidence of nocturnal hypoglycemia

Day-to-day variability

t1/2 = 25.4 ht1/2 = 12.5 h

PK profile comparison (Degludec vs Glargine)

These images are adapted from Diabetic Medicine (2013) 30: 1293–1297 and Diabetes (2011) 60 (Suppl. 1):LB11 (37-LB)

Figure 1. Mitogenic signaling in MCF-7 cells

IGF-1R

p-ERK1/2

ERK1/2

p-IGF-1R

p-AKT

AKT

Actin

0

5

10

15

Control HumanInsulin

AspB10

Insulin115

IGF-1 IGlar

IGF-

1R p

hosp

hory

latio

n(F

old

of c

ontr

ol)

IGF-1Rphosphorylation

0

5

10

Control HumanInsulin

AspB10

Insulin115

IGF-1 IGlar

Akt

pho

spho

ryla

tion

(Fol

d of

con

trol

)

AKT phosphorylation

0

2

4

6

8

Control HumanInsulin

AspB10

Insulin115

IGF-1 IGlar

Erk1

/2 p

hosp

hory

latio

n(F

old

of c

ontr

ol)

ERK1/2phosphorylation

0 2 4 6 80.1

1

10

100

1000

Time (days)

Seru

m c

onc.

(nM

)

LAPSInsulin 115t1/2 = 44.1 hrs

LAPSInsulint1/2 = 18.5 hrs

20K PEG Insulint1/2 = 6.7 hrsIDeg

t1/2 = 2.9 hrs

LAPSInsulin 115 had extended half-life than LAPSInsulin and daily insulins.

LAPSInsulin 65.1nmol/kg (weekly)LAPSInsulin 115 65.1nmol/kg (weekly)

20K PEG Insulin 65.1 nmol/kg (daily)IDegludec 55.8 nmol/kg (daily)

Figure 2. PK in SD rats (n=3, s.c.)

LAPSInsulin 115 had extended half-life and duration of action than LAPSInsulin innormal pigs.

Figure 3. PK/PD in pigs (n=3, s.c.)

Figure 4. Glucose lowering efficacy in db/db mice (n=7,s.c)

LAPSInsulin 115 showed prolonged glucose lowering effect compared to LAPSInsulin indb/db mice at the same dose and at a even as well as 1/6 dose level (Fig 4a).

LAPSInsulin 115 achieved a comparable HbA1c reduction only with 1/4 the dose ofLAPSInsulin (Fig 4b).

VehicleLAPSInsulin 21.5 nmol/kg, Q2DLAPSInsulin 43.1 nmol/kg, Q2DLAPSInsulin 86.1 nmol/kg, Q2DLAPSInsulin 115 5.5 nmol/kg, Q2DLAPSInsulin 115 11.1 nmol/kg, Q2DLAPSInsulin 115 22.2 nmol/kg, Q2D

VehicleLAPSInsulin 43.1 nmol/kg LAPSInsulin 258.3 nmol/kg LAPSInsulin 115 43.1 nmol/kgLAPSInsulin 115 129.2 nmol/kg

5

6

7

8

9

10

11

12

HbA

1c (%

)

***

10.5

8.0

9.0

7.26.8

††

***

(a) HbA1c

LAPSInsulin 115 and LAPSCA-Exendin-4 showed synergistic effects on HbA1c and bodyweight.

Figure 8. HbA1c and body weight change with combinationtreatment in db/db mice (n=5-6, s.c., Q2D, 5 weeks)

Vehicle

LAPSCA-Exd4 0.36 nmol/kg

LAPSInsulin 115 8.8 nmol/kg LAPSInsulin 115 2.2 nmol/kg + LAPSCA-Exd4 0.36 nmol/kg

LAPSInsulin 115 8.8 nmol/kg + LAPSCA-Exd4 0.36 nmol/kg

*** p<0.001 Oneway ANOVA with Dunnett’s post test vs. Vehicle † p<0.05, †† p<0.01 Oneway ANOVA with Dunnett’s post test

02468

1012141618

� B

ody

wei

ght (

g)

6.4

12.1

4.87.0

9.2

***

†

(b) Body weight change

Neutralized BWG

1/4 reduced dose

LAPSInsulin 115 could be ideal partner to LAPSCA-Exendin-4 for once-weekly insulin/GLP-1RA combination with well harmonized PK profiles. Insulin

IGF-1R

IRS-1/2

PI3K

Apoptosis

SHC

RAS/RAF

Cell proliferation

IGF-1 receptor signalingIGF-1

AKT ERK1/2

Once weekly basal insulinLAPSInsulin 115 (HM12470)

PK_LAPSInsulin 3.5 nmol/kgPK_LAPSInsulin 115 3.5 nmol/kg

6

8

10

12

HbA

1c (%

)

*p<0.05, **p<0.01 vs vehicle by Anova test

*****

*

*** ***

-1.0

-2.0

-3.4

-1.8

-2.6-3.3

1/4 reduced dose

(b) 4-week repeated dose

0 5 10 15100

200

300

400

500

600

Time (days)

FBG

(mg/

dL)

1/6reduced dose

(a) Single dose

FBG_vehicleFBG_LAPSInsulin 3.5 nmol/kgFBG_LAPSInsulin 115 3.5 nmol/kg

Once weekly Insulin + GLP-1 combination LAPSInsulin 115 + LAPSCA-Exendin-4

Figure 9. Development plan of QUANTUM project

QUANTUM is a proprietary name of Hanmi’s diabetes & obesity pipeline

0 2 4 6 80.01

0.1

1

10

Time (Days)

LAPS

CA

-Exd

-4 S

erum

Con

c. (n

M)

0 2 4 6 80.1

1

10

100

Time (Days)LAP

S Insu

lin 1

15 S

erum

Con

c. (n

M)

Figure 7. PK profile of combination vs mono in single-shotformulation (SD rats, n=6)

Single platform-derived combination Easy to combine into single formulation No pharmacokinetic interactions

LAPSInsulin 115 or LAPSCA-Exendin-4 did not have any interference in single formulation.

LAPSInsulin 115 onlyCombination formulation

LAPSCA-Exendin-4 onlyCombination formulation

LAPS Insulin 115t1/2 = 76.6 hrs

LAPS Insulint1/2 = 23 hrs

In vitro receptor binding affinity and mitogenic signaling in MCF-7Binding affinity of test materials was measured in competition between unlabeled and125I-labeled materials on the IR- or IGF-1R/CHO membrane. Mitogenic signaling wasdetected in MCF-7 cells and the phosphorylation level was analyzed by Western blot.

Pharmacokinetic prediction in humanHuman serum concentration-time was predicted by Css-MRT method from PKparameters of mice, rats and dogs. Human CL was derived from rule of exponentmethods, and human Vd was from simple allometry applied correction factor.

PK and PD analysis of LAPSInsulin 115 for once weekly basal insulinSerum concentration of test articles were determined using the a modified ELISA andPK parameters were calculated by a non-compartmental method. In an acute study,4-hr fasting blood glucose level was measured every day after s.c administration oftest articles in db/db mice. In a chronic study, HbA1c level was measured after 4-wkadministration in db/db mice with Q2D interval for human mimetic condition.

PK and PD of LAPSInsulin 115 + LAPSCA-Exendin-4 for QW combinationSerum concentration of LAPSInsulin 115 and/or LAPSCA-Exendin-4 were determinedusing modified ELISA. In a chronic study, LAPSInsulin 115 and/or LAPSCA-Exendin-4were administrated to db/db mice with Q2D interval. HbA1c level was measured after5 wk-treatment. Body weight change was monitored every 2 days.

Insulin 115 triggered levels of mitogenic signals comparable with human insulin,whereas positive controls (IGF-1 and AspB10) had significantly higher mitogenic signals.

0 2 4 6 80.1

1

10

100

0.1

1

10

Time (days)

LAPS

Insu

lin 1

15 S

erum

Con

c. (n

M) LA

PSCA

-Exd-4 Serum C

onc. (nM)

LAPSInsulin 115 5.5 nmol/kg (weekly)LAPSCA-Exd-4 2.5 nmol/kg (weekly)

IDegludec 55.8 nmol/kg (daily)Liraglutide 50.0 nmol/kg (daily)

Figure 6. PK profile comparison with long-acting GLP-1R agonist in SD rats (n=3~6, s.c.) Project 2014 2015 2016 2017 2018 2019 2020 NDA Submission

LAPSCA-Exendin-4 2018

LAPSInsulin115 2020

P3P2

2021

LAPSInsulin Combo 2021P3P2P1

P3P2P1

2013

P1