Together

0 5 10 15100

200

300

400

500

600

Vehicle

LAPSInsulin 43.1 nmol/kgLAPSInsulin 258.30 nmol/kgLAPSInsulin 115 43.1 nmol/kgLAPSInsulin 115 129.2 nmol/kg

Time (days)

FB

G (m

g/d

L)

European Association for the Study of Diabetes 50th Annual Meeting; Vienna, Austria; September 15-19, 2014 For any questions, please contact Hanmi Pharm. Co., Ltd., Phone: +82-31-371-5141; cjchoi@hanmi.co.kr

The Long-acting Basal Insulin (LAPSInsulin 115) Offers Once-weekly Dosing Potential With Favorable PK, PD And Mitogenic ProfilesIY Choi1, SY Hwang1, , 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

P933

Hanmi Hanmi Pharm. Co., Ltd.

METHODS

RESULTS

Pharmacokinetic analysis of LAPSInsulin 115 in animal modelsTest materials were s.c. injected to normal or renal failure SD rats or normal pigs. Re-nal failure SD rat model was induced by 4 mpk of cisplatin. Serum concentration of test articles were determined using ELISA and PK parameters were calculated by a non-compartmental method.

Table 1. Receptor binding affinity, metabolic and mitogenic potencies relative to human insulin

BACKGROUND

LAPSInsulin 115 as a once weekly basal insulin

Lower peak-to-trough ratio from long duration of action Improved patient adherence by once-weekly administration Ideal combination partner with weekly GLP-1 agonist

[EASD 2014 Poster #972, Combination of LAPSInsulin 115 and LAPSCA-Exendin-4]

Insulin 115 analog conjugated with a constant region of a human immunoglobulin fragment via non-peptidyl linker.

AIMS Investigation of in vitro receptor pharmacology, metabolic, and mito-

genic activity of LAPSInsulin 115 Evaluation of pharmacokinetics and pharmacodynamics of LAPSInsulin

115 in normal and diabetic animal models Prediction of human pharmacokinetic profile of LAPSInsulin 115 based on

animals’ PK data

Test materials

Receptor binding (%) Mitogenic potency (%)

Mitogenic/Metabolic po-tency§ ratio

Insulin IGF-1 SaOS-2 MCF-7 SaOS-2 MCF-7

Human insulin 100 100 100 100 1 1

IGF-1 0.6 17,984 6,255 1,527 2,234 545

Insulin AspB10 248 449 756 912 2.5 3.0

Insulin 115 86 84 119 88 <1 <1

LAPSInsulin 115 1.7 Too low to be determined

5.3 5.4 <1 <1

Both LAPSInsulin 115 and Insulin 115 showed reduced affinities on IR and IGF-1R compared with human insulin. In addition, LAPSInsulin 115 and Insulin 115 showed mitogenic/metabolic potency which was comparable to human insulin.

Figure 2. Ex vivo T cell activation of Insulin 115 and LAPSInsulin 115

Insulin 115 showed negligible activation of T cells from 50 human donors, which is below the immunogenic threshold. In addition, LAPSInsulin 115 completely attenuated the residual T cell activation.

0

5

10

15

20

LAPSInsulin 115

Insulin 115

Ex v

ivo

T c

ell

activ

atio

n

Fre

qu

en

cy o

f re

sp

on

se

(%

)

0%

8%

0

2

4

6

8

10

LAPSInsulin 115

Insulin 115

ex vivo

T

cell activatio

n

fre

qu

en

cy o

f R

esp

on

se

(%

)

Immunogenic threshold

Key requirements for once weekly insulin

Additional requirements Excellent Device No CV safety concerns

PK matched with weekly GLP-1 for combination therapy

Longer half-life

Flat profile

Low Variability (intra-patient)

Low Risk of hypos

Flexible timing/dosing

Little or no weight gain

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

Phamacodynamic analysis of LAPSInsulin 115 in db/db miceIn an acute study, 4-hr fasting blood glucose level was measured every day after s.c administration of test articles in db/db mice. In a chronic study, HbA1c level was mea-sured after 4-wk administration in db/db mice with Q2D dosing interval to mimic human QW dosing.

Ex vivo T cell activation (Antitope limited, UK)Test articles were incubated for 8 days with PBMC from 50 donors. Cell proliferation was meausred by [3H]-Thymidine incorporation and T-cell activation was identified with IL-2 secretion by Elispot assay.

Figure 1. Mitogenic signaling in MCF-7 cells

IGF-1R

p-ERK1/2

ERK1/2

p-IGF-1R

p-AKT

AKT

Actin

h-Insu

lin

Control

IGF-1

Asp B

10In

sulin

115

IGla

rgin

e

pIGF-1R (10 min)

0

5

10

15

Untreated

IGF-1

Human insulin

AspB10

Insulin A15

Control HumanInsulin

AspB10

Insulin115

IGF-1

Insulin glargine

IGlar

IGF

-1R

ph

osp

ho

ryla

tio

n

(F

old

of c

on

tro

l)

IGF-1R phosphorylation

pAkt (10 min)

0

5

10

Untreated

IGF-1

Human insulin

AspB10

Insulin A15

Control HumanInsulin

AspB10

Insulin115

IGF-1

Insulin glargine

IGlar

Akt p

ho

sp

ho

ryla

tio

n

(F

old

of c

on

tro

l)

AKT phosphorylation pERK (10 min)

0

2

4

6

8

Untreated

IGF-1

Human insulin

AspB10

Insulin A15

Control HumanInsulin

AspB10

Insulin115

IGF-1

Insulin glargine

IGlar

Erk1/2

ph

osp

ho

ryla

tio

n

(F

old

of c

on

tro

l)

ERK1/2 phosphorylation

Insulin

IGF-1R

IRS-1/2

PI3K

Apoptosis

SHC

RAS/RAF

Cell proliferation

IGF-1 receptor signaling

IGF-1

AKT ERK1/2

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

In vitro receptor binding affinity and mitogenic signaling in MCF-7Binding affinity of test materials was measured in competition between unlabeled and 125I-labeled materials on the IR- or IGF-1R/CHO membrane. Mitogenic signaling was assayed in MCF-7 cells and the phosphorylation level was analyzed by Western blot.

In vitro Binding, Proliferation, and ex vivo T Cell Activation

§ Metabolic potencies were obtained from lipogenesis assay from rat primary adipocytes

65.1 nmol/kg

0 2 4 6 80.1

1

10

100

1000

LAPS-Insulin 65.1nmol/kg [kv-3119]LAPS-Insulin A15 65.1nmol/kg [kv-3119]

20K-Insulin 65.1 nmol/kg [kv-3107]

Degludec 55.8 nmol/kg [kv-3059]

Time (days)

Seru

m c

on

c.

(nM

)

LAPSInsulin 115t1/2 = 44.1 hrs

LAPSInsulint1/2 = 18.5 hrs20K PEG Insulin

t1/2 = 6.7 hrsIDegt1/2 = 2.9 hrs

65.1 nmol/kg

0 2 4 6 80.1

1

10

100

1000

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)

Time (days)

Seru

m co

nc. (n

M)

Figure 3. PK in normal and renal failure model rats

0 2 4 6 80.01

0.1

1

10

100

-40

-20

0

20

Time (days)S

eru

m c

on

c.

(nM

)

FB

G (m

g/d

L)

LAPSInsulin 115 showed PK/PD correlation in normal pigs.

Figure 4. PK/PD in pigs (n=3, s.c.)

0 2 4 6 80.01

0.1

1

10

100

-40

-20

0

20

PK_LAPSInsulin 3.5 nmol/kg

PK_LAPSInsulin 115 3.5 nmol/kg

FBG_vehicle

FBG_LAPSInsulin 3.5 nmol/kg

FBG_LAPSInsulin 115 3.5 nmol/kg

Time (days)

Seru

m co

nc. (n

M)

FB

G (m

g/d

L)

0 2 4 6 80.01

0.1

1

10

100

-40

-20

0

20

PK_LAPSInsulin 3.5 nmol/kg

PK_LAPSInsulin 115 3.5 nmol/kg

FBG_vehicle

FBG_LAPSInsulin 3.5 nmol/kg

FBG_LAPSInsulin 115 3.5 nmol/kg

Time (days)

Seru

m co

nc. (n

M)

FB

G (m

g/d

L)

LAPS Insulin 115t1/2 = 76.6 hrs

LAPS Insulint1/2 = 23 hrs

(a) PK in normal rats (n=5, s.c.)

34 35 36 37 38 39 40 41 42 431

10

100

LAPSInsulin 3.5 nmol/kgLAPSInsulin 115 0.88 nmol/kgLAPSInsulin 115 3.5 nmol/kg

Time (days)

Seru

m c

on

c.

(nM

)

Figure 6. Human PK simulation of LAPSInsulin115 by weekly injection

Pharmacokinetics in humans was predicted based on PK of three different animals. LAPSInsulin 115 demonstrated extended half-life and lower peak-to-trough ratio than LAPSInsulin. In addition, inter-day variability of LAPSInsulin 115 was ranged from 1.0-1.1.

REFERENCES

CONCLUSIONS In vitro biological properties showed that Insulin 115 and LAPSInsulin 115

did not increase the mitogenic potency when compared with insulin.

An extended PK profiles and prolonged glucose lowering efficacies sug-gest that LAPSInsulin 115 may achieve a basal insulin profile suitable for once weekly use.

The extended human PK modeling suggests that LAPSInsulin 115 is an ideal combination partner with LAPSCA-Exendin-4 [EASD 2014 Poster #972].

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.

Figure 5. Dose-sparing and glucose lowering in db/db mice (n=7, s.c.)

LAPSInsulin 115 showed prolonged glucose lowering effect compared to LAPSInsulin in db/db mice at the same dose and even with a 6 fold lower dose.

LAPSInsulin 115 achieved a similar HbA1c reduction with a 4 fold lower dose.

HbA1c-28

6

8

10

12

Vehicle

LAPSInsulin 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

Hb

A1c (%

)

HbA1c-28

6

8

10

12

Vehicle

LAPSInsulin 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

Hb

A1c (

%)

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

**

***

*

*** ***

-1.0

-2.0

-3.4

-1.8

-2.6

-3.3

4 fold reduced dose

(b) 4-week repeated dose (a) Single dose Together

0 5 10 15100

200

300

400

500

600

Vehicle

LAPS-Insulin 43.05 nmol/kg (1X HED)LAPS-Insulin 258.30 nmol/kg (6X HED)

HM12470A 43.05 nmol/kg

HM12470A 129.15 nmol/kg

Time (days)

FB

G (

mg

/dL

)

6 foldreduced dose

0 7 14 21 28 35 42 49 56 630.1

1

10

100

1000

Time (day)

Ser

um

co

nc.

(nM

)

Weekly injected steady state profile Single week Profile

LAPSInsulin 115, t1/2 = 132 hrs

Weekly Peak-to-Trough Ratio = 1.6

LAPSInsulin, t1/2 = 55 hrs

Weekly Peak-to-Trough Ratio = 4.0

LAPSInsulin 115,

Daily Peak-to-Trough Ratio = ~1.1

Figure 7. Development plan of LAPSInsulin 115 and QUANTUM project

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

PK/PD Correlation and Dose Sparing Effect

Human PK and Developmental Plan

LAPSInsulin ,

Daily Peak-to-Trough Ratio = ~1.31.1 1.0 1.1 1.1 1.1 1.1

HM12470

0 2 4 6 81

10

100

1000Normal ratsRenal failure rats

Time (days)

Seru

m c

on

c.

(nM

)

LAPSInsulin 115 showed extended half-life compared with LAPSInsulin and daily insulins.LAPSInsulin 115 showed a comparable PK profile in a renal failure rat model

(4 mg/kg of cisplatin injected, ip, rats) compared with normal rats.

(b) PK comparison in normal and renal failure model rats (n=5, s.c.)

Insulin 115 (172 nmol/kg, sc)

LAPSInsulin 115 (22.3 nmol/kg, sc)

Insulin

0 1 2 3 41

10

100

1000NormalRenal failure

Time (hrs)

Serum

conc

. (nM)

Insulin115

0 1 2 3 41

10

100

1000NormalRenal failure

Time (hrs)

Serum

conc

. (nM)

HM12470

0 50 100 150 2001

10

100

1000Normal SD ratsRenal failure rats

Time (hrs)

Ser

um

co

nc.

(n

M)

Insulin (172 nmol/kg, sc)