rapid sar of peptide therapeutics for diabetes and beyond
TRANSCRIPT
Yanyu Peng, Liz Schanuel, Stephen Eisenberg, Misha Plam and Michael Stowell
AmideBio, LLC, 331 South 104th Street Louisville CO 80027, USA
BIOPURE PROCESS. AmideBio has developed a low cost
peptide production platform combining recombinant and
chemical methods for rapid SAR of complex or difficult to
manufacture peptides1. The process implements a library of
expression vectors optimized for bacterial or yeast expression
combined with an on column chemical cleavage process which
provides a highly orthogonal platform enabling the rapid high
purity production of a variety of peptides and proteins for drug
discovery. Here we describe the BioPure method and its
application to a variety of peptides from the amyloid peptide
Ab42 to the Kv1.3 specific channel blocker from Hadrurus
gertschi2.
B B I (S o y b e a n ) In h ib it io n A s s a y
3 U /m l try p s in , 0 .3 m M s u b s tra te in c u b a te 2 5o
C 3 h rs
-1 0 -9 -8 -7 -6 -5
0 .0
0 .2
0 .4
0 .6
0 .8
A m id e B io B B I_ 0 1
A m id e B io B B I_ 0 3
L o g [G ra m ]
Ab
s 4
05
nm
IC 5 0
1 1 .7 n g
IC 5 0
8 7 n g
S ig m a B B I
APPLICATION TO SINGLE CHAIN INSULIN. AmideBio has
implemented the BioPure process for the discovery of novel single
chain insulins3 (SCIs) suitable for the pump and patch market where
high concentration and long-term stability at elevated temperatures
are required. We have produced over different 75 analogues from a
library of more than 200 designed SCIs and have examined their
physical and biological properties.
APPLICATION TO GLUCAGON. AmideBio has implemented the
BioPure process for the discovery of solution stable glucagon
suitable for the treatment of hypoglycemia and for the patch and
pump market. Glucagon is inherently unstable in solution and
currently is only available in a lyophilized powder that must be
reconstituted prior to use in the case of hypoglycemia. Using a
structure based design approach we created a library of more than
100 potential candidates for testing. Using the Bio-Pure process we
manufactured 35 candidates for testing. Because solution stability it
the most critical aspect of this program we tested all analogues for
long term stability.
APPLICATION TO BOWMAN-BIRK INHIBITORS. Bowman-Birk
inhibitors are bi-functional inhibitors of both trypsin and
chymotrypsin. They have been shown to have potential therapeutic
application in cancer and a number of rare diseases4.
APPLICATION TO TOXIN THERAPEUTICS. A large number of
toxin molecules have been discovered with a diverse set of activities
and potential therapeutic and non-therapeutic applications including,
multiple sclerosis, psoriasis, rheumatoid arthritis, myasthenia gravis,
chronic pain, tumor diagnostics as well as pesticides. Because of
the large diversity of such toxins the ability to quickly and efficiently
produce various forms for screening will be important for realization
of any future therapeutic potential. AmideBio has implemented the
BioPure process to produce a variety of toxins with potential use in
autoimmune diseases.
1. The BioPure Process is covered under US Patent 8,796,431.
2. Chen et al. J. Biol Chem. 2012 Apr 20;287(17):13813-21.
3. The SCI molecules are covered under US Patent 9,006,176.
4. Grant et al. Multiple Sclerosis 2006; 12: 688697
5. Ming and Hellekant. "FEBS letters 355.1 (1994): 106-108.
www.amidebio.com
APPLICATION TO BRAZZEIN. Brazzein is a naturally occurring
peptide produced by the oublie berry (pentadiplandra brazzeana)
that is >2000 times sweeter than sugar on a mass basis. Brazzein
is a potential product for the $6B low calorie sweetener market.
Brazzein is all natural and heat stable with a “true” sugar taste that
is lacking in current sweeteners such as Stevia, Aspartame and
Sucralose5. AmideBio applied the BioPure process to brazzein to
determine the economic feasibility of manufacturing a food product.
TIME %A %B Flow
0.00 92 8 1
2.00 92 8 1
3.10 85 15 1
4.00 85 15 1
60.00 25 75 1
60.10 25 75 1
66.00 10 90 1
66.10 10 90 1
70.00 92 8 1
75.00 92 8 1
Ab42 DAEFRHDSGYEVHHQKLVFFAEDVGSNKGAIIGLMVGGVVIA
99.1% Ab42
Analysis Column: Ascentis® Express ES-Cyano, 2.7 Micron HPLC Column
Temp: 60OC, Buffer A: H2O + 0.05% TFA, Buffer B: ACN + 0.05% TFA
CONCLUSIONS. AmideBio has applied the BioPure process to a
wide range of difficult to manufacture peptides with excellent
success. The ability to rapidly manufacture high-purity peptides for a
variety of therapeutic and non-therapeutic applications facilitates
rapid SAR for a large number of current and future targets.
Rapid SAR of Peptide Therapeutics for
Diabetes and Beyond