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Proteasome Inhibition Suppresses Dengue Virus Egress in Antibody Dependent Infection.

Choy MM, Zhang SL, Costa VV, Tan HC, Horrevorts S, Ooi EE - PLoS Negl Trop Dis (2015)

Bottom Line: Many studies have identified the ubiquitin proteasome pathway (UPP) to be important for successful DENV production, but how the UPP contributes to DENV life cycle as host factors remains ill defined.Molecular and imaging analyses in β-lactone treated THP-1 cells suggest that proteasome function does not prevent virus assembly but rather DENV egress.Furthermore, bortezomib treatment of DENV-infected mice inhibited the spread of DENV in the spleen as well as the overall pathological changes.

View Article: PubMed Central - PubMed

Affiliation: Program in Emerging Infectious Diseases, Duke-National University of Singapore Graduate Medical School, Singapore, Singapore.

ABSTRACT
The mosquito-borne dengue virus (DENV) is a cause of significant global health burden, with an estimated 390 million infections occurring annually. However, no licensed vaccine or specific antiviral treatment for dengue is available. DENV interacts with host cell factors to complete its life cycle although this virus-host interplay remains to be fully elucidated. Many studies have identified the ubiquitin proteasome pathway (UPP) to be important for successful DENV production, but how the UPP contributes to DENV life cycle as host factors remains ill defined. We show here that proteasome inhibition decouples infectious virus production from viral RNA replication in antibody-dependent infection of THP-1 cells. Molecular and imaging analyses in β-lactone treated THP-1 cells suggest that proteasome function does not prevent virus assembly but rather DENV egress. Intriguingly, the licensed proteasome inhibitor, bortezomib, is able to inhibit DENV titers at low nanomolar drug concentrations for different strains of all four serotypes of DENV in primary monocytes. Furthermore, bortezomib treatment of DENV-infected mice inhibited the spread of DENV in the spleen as well as the overall pathological changes. Our findings suggest that preventing DENV egress through proteasome inhibition could be a suitable therapeutic strategy against dengue.

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Related in: MedlinePlus

Bortezomib decouples infectious DENV production from viral RNA replication in primary monocytes.Viral RNA genome was detected using qRT-PCR, but no infectious DENV2 was detected using plaque assay in the supernatant of cells treated with higher concentrations of bortezomib for (A) DENV1, (B) DENV2, (C) DENV3 and (D) DENV4. Mean ± SD. N = 4.
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pntd.0004058.g004: Bortezomib decouples infectious DENV production from viral RNA replication in primary monocytes.Viral RNA genome was detected using qRT-PCR, but no infectious DENV2 was detected using plaque assay in the supernatant of cells treated with higher concentrations of bortezomib for (A) DENV1, (B) DENV2, (C) DENV3 and (D) DENV4. Mean ± SD. N = 4.

Mentions: To ensure that our findings are not limited to THP-1 cell line or the use of β-lactone, we also explored if bortezomib, a FDA-approved reversible proteasome inhibitor used to treat multiple myeloma and mantle cell lymphoma, could inhibit DENV egress in primary monocytes at doses that cause minimal cytotoxicity. The 50% cytotoxic concentration (CC50) of bortezomib in primary monocytes is above 1 μM (S2B Fig). DENV1-4 opsonized with enhancing levels of humanized 4G2 (h4G2) monoclonal antibody was used to infect primary monocytes [21]. Indeed, although the virus genome was detected in bortezomib treated DENV-infected cells for all four serotypes of DENV, no infectious DENV was detected using plaque assay at higher concentrations of bortezomib (Fig 4A–4D). Furthermore, pre-treatment of primary monocytes with bortezomib also inhibited replication of different low-passage clinical isolates of all 4 serotypes of DENV in a dose-dependent manner (Fig 5A–5D). The maximal effective concentration of bortezomib that inhibited 50% of virus replication (EC50) is less than 20 nM for each of these isolates. Bortezomib was also able to inhibit 50% of virus production of the attenuated strain of yellow fever virus, YF17D, at a concentration of 0.5 nM, suggesting the dependence of functional UPP is not limited to DENV but may also apply to other flaviviruses (Fig 5E). Similar observations were also made when another proteasome inhibitor, epoxomicin, from which the licensed carfilzomib was derived, was used at concentrations well tolerated by primary monocytes (Figs 5F and S2C).


Proteasome Inhibition Suppresses Dengue Virus Egress in Antibody Dependent Infection.

Choy MM, Zhang SL, Costa VV, Tan HC, Horrevorts S, Ooi EE - PLoS Negl Trop Dis (2015)

Bortezomib decouples infectious DENV production from viral RNA replication in primary monocytes.Viral RNA genome was detected using qRT-PCR, but no infectious DENV2 was detected using plaque assay in the supernatant of cells treated with higher concentrations of bortezomib for (A) DENV1, (B) DENV2, (C) DENV3 and (D) DENV4. Mean ± SD. N = 4.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4643959&req=5

pntd.0004058.g004: Bortezomib decouples infectious DENV production from viral RNA replication in primary monocytes.Viral RNA genome was detected using qRT-PCR, but no infectious DENV2 was detected using plaque assay in the supernatant of cells treated with higher concentrations of bortezomib for (A) DENV1, (B) DENV2, (C) DENV3 and (D) DENV4. Mean ± SD. N = 4.
Mentions: To ensure that our findings are not limited to THP-1 cell line or the use of β-lactone, we also explored if bortezomib, a FDA-approved reversible proteasome inhibitor used to treat multiple myeloma and mantle cell lymphoma, could inhibit DENV egress in primary monocytes at doses that cause minimal cytotoxicity. The 50% cytotoxic concentration (CC50) of bortezomib in primary monocytes is above 1 μM (S2B Fig). DENV1-4 opsonized with enhancing levels of humanized 4G2 (h4G2) monoclonal antibody was used to infect primary monocytes [21]. Indeed, although the virus genome was detected in bortezomib treated DENV-infected cells for all four serotypes of DENV, no infectious DENV was detected using plaque assay at higher concentrations of bortezomib (Fig 4A–4D). Furthermore, pre-treatment of primary monocytes with bortezomib also inhibited replication of different low-passage clinical isolates of all 4 serotypes of DENV in a dose-dependent manner (Fig 5A–5D). The maximal effective concentration of bortezomib that inhibited 50% of virus replication (EC50) is less than 20 nM for each of these isolates. Bortezomib was also able to inhibit 50% of virus production of the attenuated strain of yellow fever virus, YF17D, at a concentration of 0.5 nM, suggesting the dependence of functional UPP is not limited to DENV but may also apply to other flaviviruses (Fig 5E). Similar observations were also made when another proteasome inhibitor, epoxomicin, from which the licensed carfilzomib was derived, was used at concentrations well tolerated by primary monocytes (Figs 5F and S2C).

Bottom Line: Many studies have identified the ubiquitin proteasome pathway (UPP) to be important for successful DENV production, but how the UPP contributes to DENV life cycle as host factors remains ill defined.Molecular and imaging analyses in β-lactone treated THP-1 cells suggest that proteasome function does not prevent virus assembly but rather DENV egress.Furthermore, bortezomib treatment of DENV-infected mice inhibited the spread of DENV in the spleen as well as the overall pathological changes.

View Article: PubMed Central - PubMed

Affiliation: Program in Emerging Infectious Diseases, Duke-National University of Singapore Graduate Medical School, Singapore, Singapore.

ABSTRACT
The mosquito-borne dengue virus (DENV) is a cause of significant global health burden, with an estimated 390 million infections occurring annually. However, no licensed vaccine or specific antiviral treatment for dengue is available. DENV interacts with host cell factors to complete its life cycle although this virus-host interplay remains to be fully elucidated. Many studies have identified the ubiquitin proteasome pathway (UPP) to be important for successful DENV production, but how the UPP contributes to DENV life cycle as host factors remains ill defined. We show here that proteasome inhibition decouples infectious virus production from viral RNA replication in antibody-dependent infection of THP-1 cells. Molecular and imaging analyses in β-lactone treated THP-1 cells suggest that proteasome function does not prevent virus assembly but rather DENV egress. Intriguingly, the licensed proteasome inhibitor, bortezomib, is able to inhibit DENV titers at low nanomolar drug concentrations for different strains of all four serotypes of DENV in primary monocytes. Furthermore, bortezomib treatment of DENV-infected mice inhibited the spread of DENV in the spleen as well as the overall pathological changes. Our findings suggest that preventing DENV egress through proteasome inhibition could be a suitable therapeutic strategy against dengue.

Show MeSH
Related in: MedlinePlus