Limits...
Chemical combinations elucidate pathway interactions and regulation relevant to Hepatitis C replication.

Owens CM, Mawhinney C, Grenier JM, Altmeyer R, Lee MS, Borisy AA, Lehár J, Johansen LM - Mol. Syst. Biol. (2010)

Bottom Line: However, inhibition of the sterol pathway with statin drugs has not yielded consistent results in patients.There is a marked transition from antagonistic to synergistic antiviral effects as the combination targets shift downstream along the sterol pathway.Our findings show how combination chemical genetics can reveal critical pathway connections relevant to viral replication, and can identify potential treatments with an increased therapeutic window.

View Article: PubMed Central - PubMed

Affiliation: CombinatoRx, Incorporated, Cambridge, MA 02142, USA.

ABSTRACT
The search for effective Hepatitis C antiviral therapies has recently focused on host sterol metabolism and protein prenylation pathways that indirectly affect viral replication. However, inhibition of the sterol pathway with statin drugs has not yielded consistent results in patients. Here, we present a combination chemical genetic study to explore how the sterol and protein prenylation pathways work together to affect hepatitis C viral replication in a replicon assay. In addition to finding novel targets affecting viral replication, our data suggest that the viral replication is strongly affected by sterol pathway regulation. There is a marked transition from antagonistic to synergistic antiviral effects as the combination targets shift downstream along the sterol pathway. We also show how pathway regulation frustrates potential hepatitis C therapies based on the sterol pathway, and reveal novel synergies that selectively inhibit hepatitis C replication over host toxicity. In particular, combinations targeting the downstream sterol pathway enzymes produced robust and selective synergistic inhibition of hepatitis C replication. Our findings show how combination chemical genetics can reveal critical pathway connections relevant to viral replication, and can identify potential treatments with an increased therapeutic window.

Show MeSH

Related in: MedlinePlus

Multi-target interactions in the replicon assay. Each panel presents a schematic of the sterol pathway showing the connection to replicon replication mediated by protein prenylation, along with response matrices for combinations representing different kinds of mechanistic interactions. Dose matrices, colored to show the activity at each dose point, are shown with green markers indicating the targets of each combination, and with shape symbols to indicate the type of interaction seen (either synergy or epistasis with an arrow indicating the direction of dominance from single agent to combination activity). (A) Combinations targeting the top of the sterol pathway produced epistatic responses, where the upstream agent's response predominates at high concentrations, irrespective of the pro- or antiviral activity of the single agents. (B) Targeting the lower end of the pathway led to inhibitory synergy in both the replicon and host viability assays, especially when both agents were downstream of OSC. (C) Inhibitors of the prenylation pathway interacted weakly (close to the SPE expectation) with probes targeting enzymes at the upper end of the sterol pathway, but had significant interactions further downstream. The strongest synergies were produced in combination with agents targeting enzymes downstream of OSC.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2913396&req=5

f4: Multi-target interactions in the replicon assay. Each panel presents a schematic of the sterol pathway showing the connection to replicon replication mediated by protein prenylation, along with response matrices for combinations representing different kinds of mechanistic interactions. Dose matrices, colored to show the activity at each dose point, are shown with green markers indicating the targets of each combination, and with shape symbols to indicate the type of interaction seen (either synergy or epistasis with an arrow indicating the direction of dominance from single agent to combination activity). (A) Combinations targeting the top of the sterol pathway produced epistatic responses, where the upstream agent's response predominates at high concentrations, irrespective of the pro- or antiviral activity of the single agents. (B) Targeting the lower end of the pathway led to inhibitory synergy in both the replicon and host viability assays, especially when both agents were downstream of OSC. (C) Inhibitors of the prenylation pathway interacted weakly (close to the SPE expectation) with probes targeting enzymes at the upper end of the sterol pathway, but had significant interactions further downstream. The strongest synergies were produced in combination with agents targeting enzymes downstream of OSC.

Mentions: Chemical combinations targeting enzymes upstream of squalene epoxidase (SQLE) at the top of the sterol pathway (Figure 4A) elicited Bateson-type epistatic responses (Boone et al, 2007), where the upstream agent's response predominates over the effects of inhibiting all downstream targets. This was especially notable for combinations including simvastatin and either U18666A or squalestatin, and for squalestatin in combination with Ro48-8071. Treatment with squalestatin prevents the SQLS substrate, farnesyl pyrophosphate (FPP) from being further metabolized by the sterol pathway. As FPP concentrations increase, the metabolite can be shunted away from the sterol pathway toward farnesylation and GGPP synthetic pathways, resulting in an increase in host protein geranylgeranylation, including FBL2, and consequently replicon replication. This increase in replicon replication explains the source of the observed epistasis over Ro48-8071 treatment.


Chemical combinations elucidate pathway interactions and regulation relevant to Hepatitis C replication.

Owens CM, Mawhinney C, Grenier JM, Altmeyer R, Lee MS, Borisy AA, Lehár J, Johansen LM - Mol. Syst. Biol. (2010)

Multi-target interactions in the replicon assay. Each panel presents a schematic of the sterol pathway showing the connection to replicon replication mediated by protein prenylation, along with response matrices for combinations representing different kinds of mechanistic interactions. Dose matrices, colored to show the activity at each dose point, are shown with green markers indicating the targets of each combination, and with shape symbols to indicate the type of interaction seen (either synergy or epistasis with an arrow indicating the direction of dominance from single agent to combination activity). (A) Combinations targeting the top of the sterol pathway produced epistatic responses, where the upstream agent's response predominates at high concentrations, irrespective of the pro- or antiviral activity of the single agents. (B) Targeting the lower end of the pathway led to inhibitory synergy in both the replicon and host viability assays, especially when both agents were downstream of OSC. (C) Inhibitors of the prenylation pathway interacted weakly (close to the SPE expectation) with probes targeting enzymes at the upper end of the sterol pathway, but had significant interactions further downstream. The strongest synergies were produced in combination with agents targeting enzymes downstream of OSC.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2913396&req=5

f4: Multi-target interactions in the replicon assay. Each panel presents a schematic of the sterol pathway showing the connection to replicon replication mediated by protein prenylation, along with response matrices for combinations representing different kinds of mechanistic interactions. Dose matrices, colored to show the activity at each dose point, are shown with green markers indicating the targets of each combination, and with shape symbols to indicate the type of interaction seen (either synergy or epistasis with an arrow indicating the direction of dominance from single agent to combination activity). (A) Combinations targeting the top of the sterol pathway produced epistatic responses, where the upstream agent's response predominates at high concentrations, irrespective of the pro- or antiviral activity of the single agents. (B) Targeting the lower end of the pathway led to inhibitory synergy in both the replicon and host viability assays, especially when both agents were downstream of OSC. (C) Inhibitors of the prenylation pathway interacted weakly (close to the SPE expectation) with probes targeting enzymes at the upper end of the sterol pathway, but had significant interactions further downstream. The strongest synergies were produced in combination with agents targeting enzymes downstream of OSC.
Mentions: Chemical combinations targeting enzymes upstream of squalene epoxidase (SQLE) at the top of the sterol pathway (Figure 4A) elicited Bateson-type epistatic responses (Boone et al, 2007), where the upstream agent's response predominates over the effects of inhibiting all downstream targets. This was especially notable for combinations including simvastatin and either U18666A or squalestatin, and for squalestatin in combination with Ro48-8071. Treatment with squalestatin prevents the SQLS substrate, farnesyl pyrophosphate (FPP) from being further metabolized by the sterol pathway. As FPP concentrations increase, the metabolite can be shunted away from the sterol pathway toward farnesylation and GGPP synthetic pathways, resulting in an increase in host protein geranylgeranylation, including FBL2, and consequently replicon replication. This increase in replicon replication explains the source of the observed epistasis over Ro48-8071 treatment.

Bottom Line: However, inhibition of the sterol pathway with statin drugs has not yielded consistent results in patients.There is a marked transition from antagonistic to synergistic antiviral effects as the combination targets shift downstream along the sterol pathway.Our findings show how combination chemical genetics can reveal critical pathway connections relevant to viral replication, and can identify potential treatments with an increased therapeutic window.

View Article: PubMed Central - PubMed

Affiliation: CombinatoRx, Incorporated, Cambridge, MA 02142, USA.

ABSTRACT
The search for effective Hepatitis C antiviral therapies has recently focused on host sterol metabolism and protein prenylation pathways that indirectly affect viral replication. However, inhibition of the sterol pathway with statin drugs has not yielded consistent results in patients. Here, we present a combination chemical genetic study to explore how the sterol and protein prenylation pathways work together to affect hepatitis C viral replication in a replicon assay. In addition to finding novel targets affecting viral replication, our data suggest that the viral replication is strongly affected by sterol pathway regulation. There is a marked transition from antagonistic to synergistic antiviral effects as the combination targets shift downstream along the sterol pathway. We also show how pathway regulation frustrates potential hepatitis C therapies based on the sterol pathway, and reveal novel synergies that selectively inhibit hepatitis C replication over host toxicity. In particular, combinations targeting the downstream sterol pathway enzymes produced robust and selective synergistic inhibition of hepatitis C replication. Our findings show how combination chemical genetics can reveal critical pathway connections relevant to viral replication, and can identify potential treatments with an increased therapeutic window.

Show MeSH
Related in: MedlinePlus