Manufacturing of Cervarix

Group 6

Outline

• Cervical Cancer & HPV

• Cervarix (HPV vaccine)

• The Manufacturing process

• Quality Control

• Conclusion

Cervical Cancer & HPV

• Cervical cancer is a malignant neoplasm of the cervix uteri or cervical area.

• Worldwide, it is the twelfth most common and fifth most deadly cancer in women.

• It affects about 16 per 100,000 per year and kills about 9 per 100,000 per year globally.

Worldwide incidence of HPV

Cervarix-HPV vaccine• Cervarix is a preventive vaccine against certain types of cancer causing human papillomavirus.

• Manufactured at GlaxoSmithKline Biologicals S.A. in Belgium.

• Aims to prevent infection from HPV types 16 & 18 that cause about 70% of cervical cancer cases.

• Developed, in parallel, by researchers at Georgetown University Medical Center, the University of Rochester, the University of Queensland Australia and the US National Cancer Institute

Cervarix- HPV vaccine• Approved for use in females 10-25 years of age.

• Consists of 3 doses of 0.5 mL each, by intramuscular injection at 0, 1 and 6 months.

• The vaccine is available in 0.5 mL single dose vials and prefilled TIP-LOK syringes

• Cervarix has shown to be 92% effective and effective for more than 4 years.

• Shelf life: 3 years.

HPV Vaccine Global SalesCervical cancer vaccine sales

Annual Sales World Wide ($m)

Product Company 2008 2010 2012 2014

Gardasil Sanofi-Pasteur MSD

865 1,222 1,686 1,984

Gardasil Merck & Co 1,403 1,227 1,399 1,372

Cervarix GlaxoSmithKline

231 797 1,246 1,353

Total HPV vaccine market 2,499 3,246 4,331 4,709

Adapted from GSK: 2010 Annual Report, Sanofi 2009 Annual Report

HPV Vaccine Global Sales

Sanofi Pasteur MSD

Merck & Co

GlaxoSmithKline

0200400600800

100012001400160018002000

20082010

20122014

HPV

sal

es (

$m)

Cervarix Active Ingredient

• Cervarix, HPV-16/18 L1 AS04 vaccine contains recombinant C-terminally truncated major capsid L1 proteins of HPV types 16 and 18 as active ingredients

• The first vaccines for humans from Baculovirus Expression System

Production from cells

• The L1 proteins of HPV-16 and HPV-18 are separately produced

• Usage of a:

recombinant Baculovirus expression system

insect cell line Hi-5 Rix4446 derived from Trichoplusia ni

Advantages of Baculovirus expression system

• High levels of heterologous gene expression

• Grows well in suspension cultures, easy for large-scale production

• Safe (restricted to infection of invertebrate species)

• Cheaper as Hi-5 Rix4446 can be cultured in serum free medium

How the vaccine is made

The Manufacturing ProcessAmplification of Seed Recombinant

Baculovirus

Extraction of Recombinant Baculovirus inocula

L1 proteins are extracted by methods of osmotic shock

Purification by chromatography, nanometric filtration and

ultrafiltration

Infection of Trichoplusia ni Hi-5 production cell lines in fermenter Sterile filtration of L1 VLP

Combination of HPV 16 and 18 L1 proteins

Formulation, filling and packaging

Purification

• The cell lysate may be subjected to a microfiltration and ultra-filtration process

• Cation- exchange chromatogrophy

• Hydroxyapatite Chromatography

- hydroxyapatite as the column medium- elution with a buffer solution containing phosphate anion- removes a large amount of contaminants from a partially

purified cellular lysate- Removes contaminating biomolecules, including DNA, lipids

and proteins are removed from the lysate.

Formulation

• The L1 proteins

• AS04 adjuvant system composed of- aluminium hydroxide - 3-O-desacyl-4.- monophosphoryl lipid A (MPL)

• The MPL immunostimulant - a detoxified derivative of the lipopolysaccharide of the gram negative bacterium Salmonella minnesota R595 strain.

Quality ControlCELL SEED AND BANKING

• The MCB, WCB, MS and WS and End of Production Cells (EPC) are tested for identity, purity and safety

• The Assays are:– Tumourogenic Potential– Adventitious Agents– Genetic Stability– Specific Viral Contaminants

Quality ControlPROCESS CONTROL & VALIDATION

• The 3 unit-steps of the drug substance production process are tested for consistency: – The recombinant HPV-16/18 baculovirus inoculum

production process – The L1 single harvest production and L1 extraction – The L1 antigen purification process.

MANUFACTURING PROCESS DEVELOPMENT

• the final process purified bulks, are assessed by subjecting each type of materials to a series of physico-chemical analyses and immunogenic property tests.

Quality Control

Physico-Chemical Analyses• SDS-PAGE • Western blotting • Capillary electrophoresis• Mass spectrometry• Amino acid analysis• Peptide mapping• N-and C-terminal analysis • Infra-Red and circular

dichromism spectroscopy• Size exclusion chromatography• Electron microscopy• Disc centrifugation size analysis

Immunogenic Properties• Antigenic activity • Binding to polyclonal sera • Immune response elicited in

mice

Summary

• Cervarix is a vaccine that prevents infection from HPV types 16 and 18

• Market

• Manufacured and purified from expression of L1 proteins in the Hi-5 Rix4446 insect cells