Product Development and Clinical Studies of Traditional Medicines

1. Product Development and Clinical Studies of Traditional Medicines N. L. Phang Nova Laboratories Sdn. Bhd. Malaysia (www.nova.com.my)

2. Objectives • To discuss the main issues and regulatory guidelines on product development. • 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.

3. 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.

4. 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.

5. Definition of herbal products (Cont’d) 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. • Dietary supplements / herbal supplements Plant components with health benefits. E.g.: Garlic or Echinacea

6. Why do we develop botanical drugs? • Diverge chemical range Enormous propensity to synthesize diverse bioactive compounds. (Multiple and mutually potentiating therapeutic effects) Complex molecules with unique properties. • Biomanufacturing factories – Relatively low-cost, highly effective and complex. 3. Higher investment cost on chemical synthesis. 4. Perception that phytochemicals provide a safer and more holistic approach to disease treatment and prevention.

7. Development process of botanical drugs • 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. • 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.

8. 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 Herbal product development:The 6S Process * * Joseph Chang, Biochemical Pharmacology, Vol. 59, pp 211-219, 2000.

9. 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.

10. 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.

11. 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.

12. Companies in North America and the UK currently involved in development of botanical drugs for clinical trials

13. 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.

14. Pharmacopoeial monographs: Quality specification • Define the quality standards of herbal products. • E.g: USP NF (USA), Indian Pharmacopoeia, Chinese Pharmacopoeia.

15. USP NF • 21 monographs for medicinal plants and medicinal extracts.

16. Structure of USP botanical monograph

17. 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

18. Chinese Pharmacopoeia Volume 1:Monograph for Ginkgo -- Describes assay method (HPLC) for flavonol glycosides.

19. 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.

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

21. 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 …”

22. 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.

23. 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.

24. Two phases of development process: Pre-clinical and clinical studies • Pre-clinical studies (Animal studies) • Evaluate the pharmacological activities. • Evaluate the toxicity. • Clinical studies (Human studies) • Evaluate the efficacy. • Evaluate the toxicity.

25. Flow chart of development of HEPAR-P Capsule (1) Medical plant Extract Bioassays Fractions Toxicology Chemical characterization

26. Flow chart of development of HEPAR-P Capsule (2 - Cont’d) Chemical standardization Qualitative & quantitative analytical methods Standardized extract – EPN 797 Pure compound Drug Delivery Technology New chemical entity Manufacturing technology for Finished product Stability studies

27. Flow chart of development of HEPAR-P Capsule (3 - Cont’d) • 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

28. Flow chart of development of HEPAR-P Capsule (4 - Cont’d) • 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

29. Specification of a botanical drug • Chemical standardization • Identification of chemical constituents • Measurement of marker compounds • Biological standardization • In vitro anti-HBsAg activity (ELISA method) • In vivo liver protective activity in rats • Stability of the finished product • Accelerated stability study • Real time stability study

30. Chemical structure:Corilagin & Phyllanthus flavonoids Corilagin – Polyphenol (Anti-viral & liver protective) Phyllanthus flavonoids (Liver protective) Chemical structure of rutin, one of the Phyllanthus total flavonoids. Chemical structure of corilagin.

31. TLC fingerprint TLC identification of Phyllanthin and fingerprints of HEPAR-P capsule Niranthin Hypophyllanthin Phyllanthin

32. TLC fingerprints (Cont’d) TLC identification of corilagin in HEPAR-P capsule TLC identification of rutin in HEPAR-P capsule Corilagin Rutin

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

34. HPLC analysis of corilagin Chromatogram of Phyllanthus niruri extract EPN 797

35. 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.

36. HEPAR-P Capsule:Quality control • Chemical standardization • Ensures batch-to-batch consistency in chemical composition. • Biological standardization • Ensures batch-to-batch reproducible biological activities.

37. Biological standardization • Liver protective – In vivo (animal study) • Anti-viral – In vitro (ELISA test)

38. Result of liver protective study Effect of Phyllanthus on CCL4 induced Liver injury in rats. Effect of Phyllanthus on CCL4 induced Liver injury in rats. AST (U/L) ALT (U/L)

39. Result of Inactivation of HBsAg study In vitro inhibitory activity against Hepatitis B surface antigen (HBsAg) by HEPAR-P™

40. 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

41. Specification of HEPAR-P Capsule Specification Appearance description Identity Impurities Potency Contaminants Quality Heavy metals Microbial Pesticide residues Color Smell Product related Process related

42. Safety of HEPAR-P Capsule:Animal toxicology studies Toxicology evaluation on the finished product: • 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)

43. 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.

44. 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.

45. 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.

46. Conclusion • 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. 3. 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).

47. Thank you