Transcript In HIV-1/HBV Co-Infection Model, CPI-431

Novel Cyclophilin Inhibitor CPI-431-32 Shows Broad Spectrum Antiviral
Activity by Blocking Replication of HCV, HBV and HIV-1 Viruses
Philippe A. Gallay1, Udayan Chatterji1, Michael Bobardt1, Daren Ure2, Dan Trepanier2, Robert Foster2 and Cosme Ordonez2
1Department of Immunology & Microbial Science, The Scripps Research Institute, La Jolla, California 92037, USA. 2Ciclofilin Pharmaceuticals
BACKGROUND & AIMS
HCV/HBV-related liver disease is the main cause of
morbidity and mortality of HIV-1 patients co-infected
with HCV and/or HBV. Despite the recent advent of
anti-HCV DAAs, the treatment of HCV/HBV/HIV-1 coinfected patients remains a challenge, as these
patients are less responsive to treatment, have
higher rates of re-infection, and develop liver
fibrosis, cirrhosis and liver cancer more often than
mono-infected patients. In this study, we used a
novel in vitro co-infection model to demonstrate
that CPI-431-32, a novel cyclophilin A (CypA)
inhibitor, simultaneously blocks replication of HCV,
HBV and HIV-1 in human cells.
RESULTS
Fig 1: HIV-1/HCV and HIV-1/HBV Co-Infection/CoCulture Systems
Fig 3: In HIV-1/HBV Co-Infection Model, CPI-431-32
Simultaneously Inhibits HIV-1 and HBV
METHODS
HIV
Using a unique and novel in vitro co-infection model,
we examined whether CPI-431-32 interferes i) with
HCV RNA synthesis of isolated replication complexes
using a quantitative replicase assay, ii) with early
steps of viral replication of HIV-1 primary isolates,
and iii) HBV virus entry (NTCP binding) and viral
replication. We also tested by ELISA the CPI-431-32
inhibition of the interaction between CyPA and the
viral proteins NS5A and p24gag of HCV and HIV-1
viruses, respectively, to explore the mechanism of
action of this drug.
RESULTS
We found that CPI-431-32 blocked viral
replication of HCV, HBV and HIV-1. CPI-431-32
was more effective than alisporivir (ALV)
(another CyPA inhibitor) inhibiting replication of
each of these three viruses. We also
demonstrated that CPI-431-32 blocks nuclear
import of HIV-1 virus. CPI-431-32 blocked
binding of CyPA to HCV NS5A and HIV-1 p24gag
more efficiently than ALV, and was more
effective than ALV against established resistantvariants of HIV-1. No other antiviral agent tested
in our assays was able to show such a broadspectrum of antiviral activity.
HBV
HIV-1 JR-CSF-infected PBMCs
and HBV AD38-infected NTCPHuh 7 cells (duplicates) were
exposed to a single dose (2 µM)
of CPI-431-32 and then
incubated together for 3 days.
Viral replication was quantified
by HIV-1 capsid/p24 ELISA and
HBV HBeAg ELISA. Data are
representative of 2 independent
experiments.
Fig 5. CPI-431-32 Blocks Formation of HBV cccDNA
Huh7, HepG2 or NTCP-Huh7
cells were transfected with HBV
in the presence of DMSO, ALV (2
µM) or CIP-431-32 (2 µM).
Three days post-transfection,
amounts of HBV cccDNA were
quantified by PCR as described
previously (Gao and Hu, J. Virol.
2007).
Fig 2: The Cyclophilin Inhibitor CPI-431-32, but not the HCV NS5A Inhibitor Daclatasvir nor the HIV-1 Protease
Inhibitor Nelfinavir, Simultaneously Inhibits HCV/HIV-1 Viral Replication in a Co-Infection Model
HCV JFH-1-infected hepatoma Huh7.5.1 cells and HIV-1 JR-CSF-infected PBMCs (duplicates) were incubated together for 3 days and then
exposed to a single dose (2 µM) of the cyclophilin inhibitor CPI-431-32, the HCV NS5A inhibitor daclatasvir, the HIV-1 protease inhibitor
Nelfinavir or drug combinations. HCV and HIV-1 replications were quantified by HCV core and HIV-1 capsid/p24 ELISA. Data are representative
of 2 independent experiments.
Fig 4: Dual Block of HBV Infection
HepaRG cells (duplicates) were
exposed to HBV AD38 together
with ALV or CPI-431-32 and viral
replication was quantified by
HBV HBeAg ELISA at day 3.
NTCP-Huh7 cells (duplicates)
were incubated with HBV preS1FITC peptide together with ALV
or CPI-431-32. HBV preS1
binding to cell surface NTCP was
quantified by FACS.
CONCLUSIONS
The unique broad-spectrum of antiviral activity
shown by CPI-431-32 might be related to the role
of CypA in viral infection. CPI-431-32 prevents
binding of CypA, a protein-folding enzyme to viral
proteins such as HCV NS5A and HIV p24gag,
which are critical for HCV and HIV-1 replication.
We hypothesize that the inhibition of CypA by
CPI-431-32 is responsible for its broad-spectrum
antiviral activity by preventing activation of
specific viral proteins required for viral replication.
Overall, this study suggests that CPI-431-32, a
novel CypA inhibitor, could potentially become a
candidate medicine for the treatment of HIV-1
patients co-infected with HCV and/or HBV.
Fig 6. Mechanism of Anti-HBV Effect for CPI-431-32
ACKNOWLEDGEMENTS
We thank F. Chisari for the Huh-7.5.1 cells, T.
Pietschmann, T. Wakita and R. Bartenschlager for
the Luc-JFH-1 plasmid, and C. Seeger for
HepAD38 cells . This work was supported by the
U.S. Public Health Service grant no. AI087746
from the National Institute of Allergy and Infectious
Diseases (NIAID) and a research grant from the
Canadian Institutes of Health Research (CIHR).