acetogenins in vivo re: wu - (p 22-36)

Herbal Medicine Development inTaiwan and the Research for

WYC-01 and WYC-02

Yang-Chang Wu

Graduate Institute of Natural ProductsKaohsiung Medical University

Kaohsiung, Taiwan

1ICSB, University of Mississippi, May 3, 2007

WHA No. 56.31 Resolution

• WHA No. 56.31 resolution urges membercountries to adjust, adopt and implement the“WHO traditional medicine strategy”.

• As the basis of national planning, there are fourmain goals including:

–

–

–

–

Policymaking

Enhancement of safety

Efficacy and quality

Ensuring availability and promotion of proper use

Reference: abstracted from the China Journal of Traditional Medicine, 2002


ICSB, University of Mississippi, May 3, 20073

The Global Trend of Traditional MedicineWHO traditional Medicine Strategy 2002-2005

• Traditional Chinese Medicine (TCM) &Complementary and Alternative Medicine (CAM)

– Integrate with national health care systems

– Promote the safety, efficacy and quality

– Increase the availability and affordability

– Promote therapeutically sound use of appropriate of

TCM/CAM

Reference: WHO Traditional Medicine Strategy 2002-2005, 2002


The Developing Dynamics and Trend ofTraditional Medicine (TM) in the World

–TM/CAM integrated into national healthcare systems where appropriate

–Safety, efficacy, and quality of TM/CAMenhanced

–Availability and affordability of TM/CAMenhanced

–Rational use of TM/CAM by providersand consumers promoted

•WHO Medicines strategy 2004-2007


Recognition of WHO on Traditional Medicine Development(Current Traditional Medicine Conditions in Taiwan) (I)

• Statistics and data released by officialWHO organizations indicate that , so far,Taiwan, China, North Korea, Korea, andVietnam have established a completelyintegrated health care system with regardsto traditional medicine.

Recognition of WHO on Traditional Medicine Development(Current Traditional Medicine Conditions in Taiwan) (II)

• The National Medicinal Policy recognizestraditional medicine and its practitioners; itallows traditional medicinal products to becomeregister and regulated and, it also allowstraditional medicine to be practiced by hospitalsand public/private clinics.

• Traditional medicine is covered by healthinsurance and related researches andeducation will be made possible to all.


Recognition of WHO on Traditional Medicine Development(Current Traditional Medicine Conditions in Taiwan) (III)

• 25 countries have created state policiesconcerning the development of traditionalmedicine

• Nearly 70 countries have stipulated lawsand regulation for the registration ofmedicinal herbal products.


Important Development and Achievement ofCommittee on Chinese Medicine and Pharmacy

(CCMP), Taiwan in Recent Years


The modernization and internalization of TCM

Provide a safe

development for TCM

environment of doctorconsultation and

medication for public

5-year plan: Establishment of anenvironment for safe use of TCM

Follow the global trendand accelerate the

The proposal to promote TCM modernization andinternationalization

Establish clinical teachingprogram for TCM

Integration of CMPmodernization and

internalization and Researchon Genome of TCM



Important achievement in recent years (I)

• The Chinese medicine has been incorporatedinto the National Health Insurance.

• Complete standards and regulations of Chinesemedicine education, certification, preclinicaltraining, and accreditation are well established.

• Te overall practice of GMP in Chinese medicinehad been reached by Sep. 2005

Important achievement in recent years (II)

• The “unified formula” of Chinese medicine areannounced and implemented.

• Enhancing clinical teaching quality of Chinesemedicine

• Planning of appraisal of affiliated Chinesemedicine departments in Chinese medicinehospitals and regular hospitals

• Improving Chinese medicine personnel quality• 13 Clinical trial centers for Chinese medicine are

constructed.


Important achievement in recent years (III)

• Announcement of Pharmaceutical Affairsregulations

• Conducting supervision program on illegaladvertisement in print media with a total of 1379cases supervised and 222 cases in violation.

• Consigning a total of 73 research programs onChinese medicine and subsidizing 2 academicinterchange programs across the Strait in 2006


Important achievement in recent years (IV)

• Conducting Program of Chinese MedicineHealth Security Protection Network Technology

• Aggressively promoting academic Chinesemedicine and held 4 international symposiums

• Putting Chinese medicine evidence medicineresearch into practice.


Taiwan’s National Health Insurance (NHI)cover around 99% of population

1.33%

InsuredUninsured

98.67%

As of Dec. 2003, 22 million Taiwan people are covered in the NHI


The Chinese medicine doctor education system

• There are two Chinese Medical education systems – 8 year systemsand post Bachelor program (5 years)

College SchoolTotal

study year

7

8

RegistrationAnnual

enrollment

120

120

Accumulative no. ofStudents

1,804

219

No. ofgraduates

2,358

108

ChineseMedicine

ChinaMedical

University

ChangGung

university

ChinaMedical

University

1966-1995

1996~

ChineseMedicine

PostBachelorChineseMedicine

8 1998~ 50 0 0

8 1984 100 1.250 998


Promoting GMP in ConventionalTraditional Medicine Manufacturers

• The committee on Chinese Medicine andPharmacy has spared no efforts in the pastyears to promote the practice of GMP inconventional pharmaceutical manufacturers. It isconsidered imperative that these factoriespractice the GMP; and the overall practice ofGMP by manufacturers had been reach by 2005.

• 103 have passed the review of the GMP for thetime being.

• 5,000 licenses; 200 standard formula


Improve the quality and safety ofTraditional Medicine

• A total of 203 items ofTraditional Medicinalmaterials were compiled in a“TM Pharmacopoeia” andpublished on 2004.Theyprovide bases for the testingmethods and specificationsof raw materials.


Establishment of Traditional medicine clinicaltrials and legislative environment in Taiwan

• The clinical trials should keep the characteristicsof traditional medicine.

• The department of Health announced its Adviceon Herbal Medicine NDA Application on 1998

• The Application Guidelines on Herbal MedicinalClinical Trials (IND) and guidelines on HerbalMedicine NDA Applications (NDA) are to beresearched and drawn up.


New traditional Medicine Clinical TrialApplication case (2001-2004)

• 22 cases (partial ones are center test)

• 11 cases have been passed theexamination and verification for clinicaltrail.

• 6 cases under submitting anotherapplication and examination

• 2 cases have been withdrawn

• The first NDA approval in 2005


Kaohsiung Medical UniversityKaohsiung, Taiwan


Natural Products Chemistry and TCM Analysis

• Discover of new lead compound from natural products

• Total/partial synthesis of bioactive compounds

• TCM fingerprint, qualitative &quantitative analysis


Pharmacology and BioactivityAnti-cancer, anti-platelet agents

screening and mechanisms research


The Characteristics of

WYC01


The structural features ofAnnonaceous acetogenins

some oxygen-bearing moieties, such aszero to three tetrahydrofuran (THF)

and/or tetrahydropyran (THP) rings and

several hydroxyl groups

the spacer moiety linking the two

rings with hydroxyl groups

a terminal γ-lactone ring

a terminal aliphatic side chain

Annonaceous acetogenins are unique second metabolites of Annonaceous plantscontaining C35-C37, 0-3 THF rings, a γ-lactone ring, and a long aliphatic regions.Till now, more than 450 annonaceous acetogenins were isolated from this family.


Biological activity against human cancer cellsThe IC50 values of WYC-01, Doxorubicin and Taxol against Various Human

Cancer Cell Lines.

Cell lines

Hepatoma

HepG2

Lung

Calu-1

H23

H23/0.3

Colon

MES-SA/Dx5

COLO205

Ovary

SKOV3

Breast

BT474

MDA-MB-453

MDA-MB-231

MCF-7

Prostate

PC-3

LNCaP.FGC

Gastric

TSGH9201

27.42

37.89

0.3571

3.57

5.02

3.87

60.45

23.5

34.18

<0.01

18.0357

3.6866

27.7

5.94

0.48

54.86

1.14

0.82

1.37

0.82

1.03

1.04

0.5942

0.9587

0.7396

6.19

0.0775

0.0065

7.09

2.54

0.006

0.015

15.01

0.0102

0.002

-

-

-

0.0098

WYC-01(μg/ml)

36.4

Doxorubicin (μM)

1.49

Taxol (μM)

11.7


Toxicity test

The hepatic, renal functions, and hematological values ofthe tested mice by oral administration of WYC-01 for 14 and 28 days

20.019.0

18.017.016.015.0

14.013.012.0

11.010.09.0

8.00.80.60.40.20.0

0 1 2

250GOT

200

BUN

150

25.024.023.022.021.020.019.018.017.016.015.014.013.012.011.010.09.08.00.80.60.40.20.0

0 1

350GOT

BUN300

250

200

150

GPT100

Creatinine 50

00 1 2 3 4

100

GPT

Creatinine50

00 1 2 3 43 4 2 3 4

300

250

200

150

100HCT MCV

50

201612840

0

WBC RBCHGB

MCHMCHC

TestParametersPLT

858075706560555045403530252015543210

0

Neturo-Seg

TestParameters

550

500

450

400

350

300

250

200Lymphocytes 150

100

50201612840

0 1 2 3 4 5 6 7 8 9

TestParameters

TestParameters

Monocytes

90858075706560555045403530252015543210

0 1

TestParameters

1 2 3 4 5 6 7 8 9 10 1 2

TestParameters

3 410 2

TestParameters

3 4

TestParameters

14 days Normal

Vesicle formulation

AS 25mg/kg

AS 20mg/kg

AS 15mg/kg

28 daysNormal

Vesicle formulation

AS 25mg/kg

AS 20mg/kgAS 15mg/kg

1. The values of WYC01 are stable as that of control.2. WYC01 is quite low toxicity to test mice.


Toxicity test

The 15th day The 29th day

normal control 25 20 15 normal control 25 20 15

(mg/kg/10ml)

No differences in mice organs, including liver, renal, heart,lung, and gastrointestinal between tested and control groups


In Vivo test

Some side effects on the skin of mice, such as skin rough, skin bits and erythma(red spots), were present after oral administrating WYC-01 for 21 days, butthese phenomena disappeared after stopping drug administration for six days.


Anti-angiogenesis activity

1.5 hr ~ 2hr → 24hrhuman umbilical vascular endothelial cell (HUVEC) cord formation

Cisplatin with a dose of 333μM cannot affect the formation of HUVECDoxorubicin was reported to inhibit the anti-angiogenesis activity.


Anti-angiogenesisactivity (4 Hr)

Anti-angiogenesisactivity (24 Hr)

WYC-01 2.5 µg/ml WYC-01 25 µg/ml WYC-01 2.5 µg/ml WYC-01 25 µg/ml

Doxorubicin 17.7 µMDoxorubicin 177 µMDoxorubicin 17.2 µM Doxorubicin 172 µM

Both, WYC-01 and Doxorubicin, show similar effects on anti-angiogenesis.


Anti-Solid Lung Tumor Experiment

Post-treatment model (Therapy)

Procedures1. Culture tumor cell2. Implant tumor cell3. Treat with drugs

After induced tumor 4 days

Nude mice 10 wksWYC-01 P.O route

7.5, 10, 12.5mg/kg ; qd x 30


Drug resistantis a problemfor cancer

therapyMulti-Drug resistant solid lung tumor

H23/0.3 test in nude mice model

Post-treatment (therapeutic) model

Pre-treatment (prevent) model


Anti-Solid Lung Tumor ExperimentPre-treatment model (Prevention)

Procedures

Pre-treatment WYC-01

1. Treat with drugs2. Culture tumor cell3. Implant tumor cell4. Treat with drugs again

Nude mice 8 wks :WYC-01 P.O route10mg/kg ; qd x 10

Nude mice 8 wks :Cisplatin I.V. route10mg/kg ; q3d x 2

After induced tumor 5 days

Nude mice 10 wks :WYC-01 P.O route10mg/kg ; qd x 42

Nude mice 10 wks :Cisplatin I.V. route10mg/kg ; q3d x 2


Post-treatment model results

800

700Control

7.5mg/kg

10mg/kg

12.5mg/kg

25

24

500

400

300

200

100

0

1day

3day

5day

9day

12da

y

16da

y

19da

y

22da

y

26da

y

30da

y

600 23

22

21

20

19

18

17

16ay ayay ay ayay ay ay ay ay

1d 3d 5d 9 d 12 d 16 d 19 d 22 d 26 d 30 d

Days after treatment

1. Tumor size is smaller than control

2. Optimization dose: 10 mg / kg

Days after treatment

Body weight is not obviously change


Physiological observation

• Activity observation - normal

• During drug administration, nude miceobserve hair growth in each group.


Pre-treatment model results-IWYC-01 10mg/kg treatment group

#3-1 10 days after stop treatment #4-3 10 days after stop treatment

After stop treatment 10 days, the tumor is disappear

#3-1 80 days after stop treatment #4-3 80 days after stop treatment

After 80 days, the tumor is still not found in nude mice36


acetogenins in vivo re: wu - (p 22-36)

Health & Medicine

traditional medicine

academic chinese medicine

herbal medicine development

university of mississippi

alternative medicine

traditional medicinal

important development

tcm modernization