Human Growth Hormone

CHEE450

By Leslie Davis

Human Growth Hormone

Simulates linear growth in humansEffects on the growth plates of the long

bones Increased cell growth Increased protein, carbohydrate and lipid

metabolism

Human Growth Hormone

Generally used to treat those with GH-deficiencies

Beneficial effects: burns, bone fractures and Turner’s syndrome Shown promise in treating chemotherapy and

AIDSIncreases body mass

Human Growth Hormone

Polypeptide 191 amino acid residuesMW = 22125 daltonsHighly hydrophobic proteins

Historical Perspective

Isolated hGH in human pituitary 1956Proof of effectiveness 1958Obtained from human pituitary gland only

Urgent need to obtain human pituitary glands at autopsy

High cost

Cloned from human gene in 1980s

Recombinant DNA Synthesis of hGH

Cloning of human DNAAdapting of the cloned gene for

expression in Escherichia coli E. coli did not have the biochemical

machinery 84 base pairs of ds DNA synthesized for direct

expression

Cloning of the gene

How it works

Chemical messenger produced by the pituitary gland

Promotes growth during childhood

Plays important metabolic role throughout adulthood

Pituitary Gland

Production Process

Fermentation Highly reproducible Intracellular E. coli W3110 with a pB322-derived plasmid

coding Antibiotic resistance of genetically modified E.

coli strain Typically tetracycline and ampicillin resistant

Production Process

Glucose and mineral salt mediumExponentially growing cells in batch mode

Transferred from seed fermentor to production bioreactor

10% of final volume

High cell density fermentation

Production Process

Fed-batch bioreactor Glucose feed ~ 500-600 g/L May use peptone for amino acid source Temperature – near maximum tolerance of E.

coli Agitation generally ranges 1000-1500 rpm Slightly positive pressure

Production Process

Purification Cell disruption Microfiltration Chromatographic methods

Genentech adapted anion exchange chromatography and gel filtration technology (early 80s)

Production Process

Issues with scale up Mass transfer through high density cells

OxygenGlucose feed

Inhibitory ion formation Acetate and formate

Final Product

White lyophilized powderLow oral bioavailability (<1%)Intended for intravenous, subcutaneous or

intramuscular administrationSold in vials or cartridges containing

4 - 24 mg of somatropin Preservatives Water for injection

Final Product

Approximately 28 day shelf lifeOxygen Sensitive Must be refrigeratedImprove the solubility, sodium dodecyl

sulfate (extremely surface active) or denaturing agents - urea and guanidine hydrochloride

Market

U.S. Manufacturers Genotropin by Pharmacia & Upjohn Company,

Humatrope by Eli Lilly, Norditropin by Novo Nordisk, Saizen and Serostim by Serono Laboratories, Nutropin by Genentech

Market

Costs: $75 to $200 per weekWorld-wide sales of $1.6 billionApproximately 20,000 U.S. children taking

drug Prescribed for non-GH-deficient children if

(U.S.) <3SD below mean for age category <25% growth velocity

Discussion

Psychological issues Cultural “heightism”

Increased metabolism, reduced fat, anti-aging, gain strength

Articles

References

Human Growth Hormone, edited by Salcatore Raiti and Robert A. Tolman, 1986

Human Growth Hormone: Research and Clinical Practice, edited by Roy G. Smith and Michael O. Thorner

Influence of Scale-Up on the Quality of Recombinant Human Growth Hormone, Bylund, Castan, Mikkola, Veide and Larsson

Modelling the effects of Glucose Feeding on a recombinant E. coli Fermentation, Cockshott and Bogle

Further Studies Related to the Scale-up of High Cell Density Escherichia coli Fed-Batch Fermentations: The Additional Effect of a Changing Microenvironment When Using Aqueous Ammonia to Control pH

References

http://pi.lilly.com/us/humatrope-pi.pdf http://www.hghnews.us/p/91.html http://www.hgh-humangrowthhormone.com/

hgh_human_growth_hormone_sitemap/hgh_manufacturer.html

http://www.novonordisk.com/therapy_areas/growth_hormone_therapy/default.asp

http://patft.uspto.gov/netacgi/ http://www.genotropin.com/patients/

growth_disorders/index.asp