Transcript Early Days of Growth Hormone Therapy

EARLY DAYS OF GROWTH
HORMONE THERAPY:
IT PAYS TO TAKE CARE
Phil Lowry
Leeds University 1962-1969
Ciba/Geigy 1969-1973
St Bartholomew’s Hospital “Barts” 1973-86
Research funded by Medical Research Council
DHS therapeutic hGH research 1974-1981
University of Reading 1986-2009 (MRC funded)
PITUITARY
GLAND
REPRODUCTION
GROWTH
STRESS
THYROID
Human Growth Hormone:
From Cadaver to Clinic
Height (cm)
hGH
5mg/gland
Late 60s simple
precipitation…no
chromatography
Impure hGH
Pain after injection
• The original extraction methods started with acetone
dried human pituitary glands
• Raben method used hot glacial acetic acid
• Wilhelmi method used alkali pH
• Isolation was achieved by precipitation
• Neither method used any chromatography
• Both methods caused some denaturation
• Both preparations were associated with significant
antibody generation which blocked hGH bioactivity
• 1984 first cases of CJD (Creutzfeldt-Jakob Disease)
hGH (ex-cadaver) therapy discontinued
An example of therapeutic
hGH isolated without
using chromatography
containng 60%
“non-specific junk”
Deceasing in size
• At Barts we used frozen pituitary glands
• Extracted in mild alkali, then by isoelectric
precipitation in mild acid/10% ethanol
• Filtered through 0.22 micron membrane
• Gel chromatographic step
• Ion exchange chromatographic step
• Freeze dried. 5mg/gland … 2.5 times purer
At Wellcome Laboratories
• Redissolved and refiltered through 0.22
micron membrane
• Freeze dried in ampoules for clinical use
Therapeutic human growth hormone purification at Barts 1975-1981
Ion exchange
chromatographic
column
Gel filtration
chromatographic
cloumn
Transmitted Spongioform Encephalopathies…TSEs
Originally called “Slow Viruses” up to 14 years after infection
“Scrapie” endemic in sheep whose brains were fed to cows
Bovine Spongioform Encephalopathy “BSE” or mad cow disease
Creutzfeldt Jacob Disease was very rare in humans before
BSE infected meat in hamburgers lead to new variant CJD
“Fore” tribe in New Guinea ate brains of their dead..KURU
All these diseases are transmitted by an infectious protein
called PRION
Prion protein found in membrane of neural cells but
in its infectious state has folded into an extremely
stable form which can withstand autoclaving and
digestion by proteases.
When ingested (BSE, new variant CJD) or injected
(hGH therapy) finds its way into neural tissue
where it very slowly catalyses the conversion of
normal prion proteins into the infectious form
leading to spongioform encephalophathy.
Can also get infected from e.g. corneal transplant
and blood transfusion.
An example of therapeutic
hGH isolated without
using chromatography
containng 60%
“non-specific junk”
contained infectious particles
Deceasing in size
The Lancet, 1985 Aug 3; 260-2.
1978 Alan Dickinson contacted
MRC Head Office about
possibility of contracting CJD
from hGH extracted from
human pituitary glands
load
Polyacrylamide gel electrophoresis, under
non-dissociating conditions, of a sample of
Barts therapeutic human Growth Hormone
Since production at Barts stopped in 1981 there have
been no cases of CJD associated in patients
treated exclusively with this preparation. Antibody
titres dropped after switching over to Barts hGH.
De-amidated forms of human growth hormone
But in 1995 I ended up on the stand in the High Court !
+ Since 1986 hGH purified from genetically engineered bacteria
1986 Human Growth Hormone Cloned:
purified recombinant hGH used in Clinic
Pituitary gland
mRNA
Clone
rhGH