www.ias2013.org kuala lumpur, malaysia, 30 june - 3 july 2013 protein engineering of the lectin...
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www.ias2013.org Kuala Lumpur, Malaysia , 30 June - 3 July 2013
Protein engineering of the lectin BanLec reduces its mitogenic activity and prevents
vaginal HIV transmission
Michael SwansonUniversity of North Carolina
www.ias2013.org Kuala Lumpur, Malaysia , 30 June - 3 July 2013
• BanLec binds to high-mannose on gp120
• Broad, potent activity anti-HIV activity
• Microbicide Candidate• Can its stimulatory
activity be reduced?– Substitution of amino
acids in binding sites
Engineering BanLec to Reduce its Mitogenic and Stimulatory Activity
Inhibition of HIV-1 Infection of TZM-bl cells
0.01 0.1 1 10 100 10000
102030405060708090
100
0.23
IC50 (nM)
Lectin (nM)
% L
uci
fera
se A
ctiv
ity
WT BanLec
Lectin (nM)
Sti
mu
lati
on
Ind
ex(P
erip
her
al B
loo
d L
ymp
ho
cyte
Pro
lifer
atio
n)
0.1 1 10 100 10000
10
20
30
BanLec WT
Lectin (nM)
Sti
mu
lati
on
Ind
ex(P
erip
her
al B
loo
d L
ymp
ho
cyte
Pro
lifer
atio
n)
0.1 1 10 100 10000
10
20
30
BanLec WTBanLec H84T
Lectin (nM)
Sti
mu
lati
on
Ind
ex(P
erip
her
al B
loo
d L
ymp
ho
cyte
Pro
lifer
atio
n)
0.1 1 10 100 10000
10
20
30
BanLec WTBanLec H84TGRFT
Inhibition of HIV-1 Infection of TZM-bl cells
0.01 1 100 100000
102030405060708090
100WT BanLecH84T
0.23
IC50 (nM)
6.95
Lectin (nM)
% L
ucife
rase
Act
ivity
TFV 3.2 µM
Strategy Outline• cDNA for BanLec• Introduce amino
substitutions Express in E. coli
• Purify variants• Assess stimulatory
activity
www.ias2013.org Kuala Lumpur, Malaysia , 30 June - 3 July 2013
Topically-applied BanLec H84T Prevents Vaginal HIV TransmissionApply 75 µg
BanLec H84T Topically Expose Vaginally with HIV-1JR-CSF
10 minutes laterMeasure Viral Load
0 1 2 3 4 5 60
102030405060708090
100
Weeks Post Exposure
% o
f M
ice
Pro
tect
ed
Control n=8BanLec H84T n=7
p=0.0359
www.ias2013.org Kuala Lumpur, Malaysia , 30 June - 3 July 2013
Conclusions
• BanLec H84T has reduced mitogenicity and is effective ant preventing vaginal HIV transmission in vivo
• BanLec H84T should be investigated as a potential microbicide.
• Engineering of mitogenic, anti-HIV lectins could lead to more microbicide candidates
www.ias2013.org Kuala Lumpur, Malaysia , 30 June - 3 July 2013
AcknowledgementsUniversity of Leuven, Rega Institute for Medical Research-Dominique Schols-Dana HuskensUniversity of Michigan-David Markovitz-Dan BodreauxUniversity of North Carolina-J. Victor GarciaNIH Grants: AI100775