Product Development and Clinical Studies of Traditional Medicines

N. L. Phang Nova Laboratories Sdn. Bhd. Malaysia (www.nova.com.my)

Objectives
1. To discuss the main issues and regulatory guidelines on product development. 2. To describe develop process of a traditional medicine with therapeutic claim.
To illustrate with a case study The development of a botanical drug containing standardized Phyllanthus niruri extract EPN 797.

Successful herbal product: Development of bromhexine

Ethnobotany approach in drug development Bromhexine (Trade name: Bisolvon)
Is a popular mucolytic agent for cough.
It is derived from Indian Adhatoda vasica (the malabar nut tree) which is traditionally used in cough therapy.

Definition of herbal products

Three categories of herbal products: A. Drugs (NCE, new chemical entities) Single active ingredient pharmaceuticals originating from plants. E.g.: vinblastine, digoxin.

Definition of herbal products (Contd)

B. Botanical drugs (Multi-component botanical drugs) Botanical drugs are manufactured medicines obtained exclusively from plants to relieve, prevent or cure a disease or to alter physiological and pathological process. E.g.: None in USA, several in clinical trials. C. Dietary supplements / herbal supplements Plant components with health benefits. E.g.: Garlic or Echinacea

Why do we develop botanical drugs?

1. Diverge chemical range
Enormous propensity to synthesize diverse bioactive compounds. (Multiple and mutually potentiating therapeutic effects) Complex molecules with unique properties.

2.

Biomanufacturing factories Relatively low-cost, highly effective and complex.

Higher investment cost on chemical synthesis. Perception that phytochemicals provide a safer and more holistic approach to disease treatment and prevention.

3. 4.

Development process of botanical drugs

i. ii. iii. Development of botanical drugs is a hard and expensive task. Each new drug requires a big investment and a minimum of 5 - 10 years of work. Therefore, we have to adopt a very carefully planned strategy.

iv.

The development of botanical drug emphasizes on three

important aspects of the product:
Quality Efficacy Safety Inter-related as efficacy and safety largely depend on the quality.

Herbal product development: The 6S Process *

Selection Herb informatics Unmet health need
Sourcing Chemotaxonomy Raw material analysis Structure Identification of active constituents Method validation

Standardization Chemical profile Pharmacological profile

Safety Historical / traditional use Toxin analysis Safety studies in animals Substantiation . Review of pre-existing data Prospective clinical study

* Joseph Chang, Biochemical Pharmacology, Vol. 59, pp 211-219, 2000.

Study the guidelines for botanical drugs

The final products are strictly controlled by the regulatory authorities. It is important to study the requirements of several important regulatory guidelines.

USA FDA CDER Guidelines for botanical drugs

The Guidance for Industry: Botanical Drug Products quality standards for standardized plant extracts (botanical drugs). Guidelines for the development of drug

products from botanicals.

It is now possible to market these products under the New Drug Application (NDA) approval process.

Abbreviated preclinical and clinical testing protocols

For plants with safe history of human use USA FDA proposed abbreviated preclinical and clinical testing protocols for botanical drugs derived from plants.

Companies in North America and the UK currently involved in development of botanical drugs for clinical trials
Company Ancile pharmaceuticals. San Diego, CA CV Technologies. Edmonton, Canada GW Pharmaceuticals. Salisbury, UK Oxford Natural Products. Oxford, UK PhytoCeutica. New Haven, CT Phytomedics. Dayton, NJ Phytopharm. Godmanchester, UK Areas of clinical testing Sleep, anxiety disorders Respiratory infection Cannabis-based prescription medicines

Dysmenorrhoea, hepatitis-C symptoms, cognitive decline

Cancer, neurovascular disease Autoimmune diseases, cancer Appetite suppressant, inflammatory bowel disease, Alzheimers disease, cancer

WHO & EMEA guidelines

The WHO Guidelines for the Assessment of Herbal Remedies, adopted by the International Conference of Drug Regulatory Authorities (Ottawa, October 1991). EMEA Guidelines.

Pharmacopoeial monographs: Quality specification

Define the quality standards of herbal products. E.g: USP NF (USA), Indian Pharmacopoeia, Chinese Pharmacopoeia.

USP NF
21 monographs for medicinal plants and medicinal extracts.

Structure of USP botanical monograph

Monograph for Gingko (USP26 NF21): i. Definition of herbal product iv. Microbial limits

ii.

Identification tests

v.

Limit tests For soil and sand contamination

Heavy metals

iii.

Content tests Quantitative determination of marker compounds

a. b. Content of flavonol glycosides by HPLC Content of terpene lactones by HPLC

vi.

vii.. Pesticide residues

Indian Herbal Pharmacopoeia Volume 2: Monograph for Phyllanthus

Describes analytical methods (HPLC) of marker compounds: Phyllanthin and Hypophyllanthin
phyllanthin hypophyllanthin phyllanthin

hypophyllanthin

(a) Reference standards

(b) Sample preparation

Chinese Pharmacopoeia Volume 1: Monograph for Ginkgo

-- Describes assay method (HPLC) for flavonol glycosides.

Main specification requirement of botanical drugs

Chemical standardization: i. quality identification of the product. ii. quantitative determination of the marker compound(s) of the product.

Chemical standardization
Chemical standardization emphasizes the importance of determination of the content of the herbal products.

Importance of measurement
A quotation from Lord Kelvin:
When you can measure what you are speaking about, and express it in numbers, you know something about it

Malaysian guidelines
Guidelines for Standardization of Herbal products Guidelines for Levels and Kinds of Evidence to Support Claims for Therapeutic Products

-- Published by National Committee For Research And Development In Herbal Medicine (NRDHM) -- Similar to FDA guidelines, they allow the development of botanical drugs with therapeutic claim.

HEPAR-P capsule: Approved for clinical trial

HEPAR-P Capsule contains standardized

Phyllanthus niruri extract. 1st local product approved by Medical Research & Ethics Committee, Ministry of Health Malaysia.

For clinical testing to evaluate antiviral

activities in patients with chronic hepatitis B.

Two phases of development process: Pre-clinical and clinical studies

1. Pre-clinical studies (Animal studies)
Evaluate the pharmacological activities. Evaluate the toxicity. 2. Clinical studies (Human studies) Evaluate the efficacy.

Evaluate the toxicity.

Flow chart of development of HEPAR-P Capsule (1)

Medical plant

Extract

Bioassays

Fractions

Toxicology

Chemical characterization

Flow chart of development of HEPAR-P Capsule (2 - Contd)

Chemical standardization
Qualitative & quantitative analytical methods

Standardized extract EPN 797 Drug Delivery Technology Manufacturing technology for Finished product Stability studies

Pure compound New chemical entity

Flow chart of development of HEPAR-P Capsule (3 - Contd)

Quality control protocol Chemical standardization Biological standardization Complete monograph (of herbal product) Approved herbal supplement Animal toxicology studies (on the finished product) -- Rodents & Non-rodents -- According to FDA / WHO / Malaysian guidelines

Flow chart of development of HEPAR-P Capsule (4 - Contd)

LD 50 Acute toxicology studies Sub-acute toxicology studies Chronic toxicology studies Completion of pre-clinical studies

Submission to National Committee for Research and Development In Herbal Medicine (NRDHM)
Approval to conduct clinical trial at appointed hospitals Report of clinical trial by clinical investigators Recommendation by NRDHM Application to DCA for registration with therapeutic claim

Specification of a botanical drug

i. Chemical standardization
Identification of chemical constituents Measurement of marker compounds

ii. Biological standardization

In vitro anti-HBsAg activity (ELISA method) In vivo liver protective activity in rats

iii. Stability of the finished product

Accelerated stability study Real time stability study

Chemical structure: Corilagin & Phyllanthus flavonoids

Corilagin Polyphenol
(Anti-viral & liver protective)

Phyllanthus flavonoids
(Liver protective)

Chemical structure of corilagin.

Chemical structure of rutin, one of the Phyllanthus total flavonoids.

TLC fingerprint
TLC identification of Phyllanthin and fingerprints of HEPAR-P capsule

Niranthin Hypophyllanthin Phyllanthin

TLC fingerprints (Contd)

TLC identification of corilagin in HEPAR-P capsule TLC identification of rutin in HEPAR-P capsule

Corilagin

Rutin

Chemical standardization: HEPAR-P Capsule

Content of one HEPAR-P Capsule 250 mg of Phyllanthus niruri extract EPN 797 standardized to contain: i. Corilagin 10 mg ii. Total flavonoids 45 mg

HPLC analysis of corilagin

Chromatogram of Phyllanthus niruri extract EPN 797

Biological standardization
Chemical standardization is inadequate. Botanical drug contains a complex mixture of chemical compounds. Chemical standardization does not give a complete picture of a herbal product. We have combined biological assays with chemical fingerprints to provide assurance of efficacy and consistency.

HEPAR-P Capsule: Quality control

Chemical standardization
Ensures batch-to-batch consistency in

chemical composition.

Biological standardization
Ensures batch-to-batch reproducible biological activities.

Biological standardization
Liver protective In vivo (animal study)

Anti-viral In vitro (ELISA test)

Result of liver protective study

Effect of Phyllanthus on CCL4 induced Liver injury in rats. ALT (U/L)
200

Effect of Phyllanthus on CCL4 induced Liver injury in rats. AST (U/L)

200

150

150

100

100

50

50

0
Control CCL4 control Treatment with Phyllanthus

0
Control CCL4 control Treatment with Phyllanthus

Result of Inactivation of HBsAg study

In vitro inhibitory activity against Hepatitis B surface antigen (HBsAg) by HEPAR-P

HEPAR-P Capsule: Monograph

Quality specification (as compared to USP , IHP , CP) Definition of product Chemical identification Chemical assays for characteristic marker compounds Assays for biological activity (or biological assay) Heavy metals Microbial limits

Specification of HEPAR-P Capsule

Specification

Appearance description
Color Smell

Identity

Impurities

Potency

Contaminants

Quality

Product related

Heavy metals Microbial Pesticide residues

Process related

Safety of HEPAR-P Capsule: Animal toxicology studies

Toxicology evaluation on the finished product:

i.
ii. iii. iv.

Acute studies (14 days, rodents and non-rodents)

Sub-acute studies (90 days , rodents and non-rodents) Chronic study Rodents (180 days) Chronic study Non-rodents (270 days)

Pre-clinical studies for HEPAR-P Capsule (Animal studies)

Identification The active fraction. Characterization The marker compound(s). Establishment Chemical standardization methods. (Optional Biological standardization methods) Animal toxicology studies. Stability studies. Formulation development. To establish a complete monograph of the product. Medical Research & Ethnics Committee approval to conduct clinical trial.

Completion of pre-clinical studies: Approval for clinical trial

-- Approval by National Committee For
Research And Development In Herbal

Medicine (NRDHM).
-- Clinical trial at Selayang General

Hospital on Jan / Feb., 2005.

Clinical studies (Human studies)

Clinical evaluation of safety and efficacy in human

subjects by appointed clinical investigators. Report of the clinical evaluation by the investigators. Recommendation by National Committee for Research and Development in Herbal Medicine. Submission to DCA for registration of product as botanical drugs with approved therapeutic claim.

Conclusion
1. The development of botanical drugs is likely to be a major area of plant biotechnology expansion in the 21st century. The future of botanical drugs will depend on consumer and regulatory acceptance. The important challenge is to provide science-based evidence to consumers and regulatory authority on: i. Efficacy (By clinical evaluation in human), ii. Quality (Establish monograph of the standardized extract), and iii. Safety (By toxicological evaluations in animals and in human).

2.

3.

Thank you