Scientists report on the development of modified bisphosphonate drug compounds that kill malaria in mice. The lipophilic bisphosphonates are very similar to zoledronate and risedronate, but unlike the two marketed bone strengthening drugs, they are capable of entering infected host red blood cells to target a newly validated malarial parasite enzyme, GGPPS.
Reporting their work in PNAS, University of Illinois chemistry professor Eric Oldfield, Ph.D., and colleagues, say the new compounds are active at very low concentrations and don’t appear toxic at levels tested in mice.
They claim their work is the first to show that the GGPPS enzyme is a valid target for malaria. “Our work gives a new direction to find new antimalarial drugs,” comments co-author Yonghui Zhang. The researchers describe their research in a paper titled “Lipophilic analogs of zoledronate and risedronate inhibit Plasmodium GGPPS and exhibit potent anti-malarial activity.”
Previous work has indicated that bisphosphonate drugs akin to zoledronate and risedronate can also inhibit the growth of a range of parasitic protozoa including species responsible for leishaminiasis, toxoplasmosis, trypanosomiasis, and malaria. However, unlike the nitrogen-containing bone disease compounds, the most effective bisphosphonates against these parasites appear to be more lipophilic.
Although the target of such drugs hadn’t previously been determined, another line of research recently identified, cloned, and crystallized a Plasmodium vivax (Pv) protein, geranylgeranyl diphosphate synthase (PvGGPPS), which is inhibited by bisphosphonates, and thus represented a promising candidate as a target for compounds previously identified.
To investigate this further the university team recently evaluated the IC50 values for 25 bisphosphonates against PvGGPPS, but found that the IC50 values didn’t correlated with P. falciparum growth inhibition. This indicated that while bisphosphonates may well be good inhibitors of PvGGPPS, they can’t necessarily get into infected red blood cells. Another problem with bisphosphonates, the investigators point out, is that they tightly bind to bone and are rapidly removed from the bloodstream, which isn’t a good feature of a proposed anti-infective.
To address such issues the team has been developing lipophilic bisphosphonates, in which the 1-OH group on the bisphosphonate backbone is removed to prevent bone binding, and replaced by a variety of hydrophobic side-chains. Prior work by the team has already suggested such compounds are more potent both in vitro and in vivo in tumor cell growth inhibition assays, and against malaria parasites.
Their latest work set out to screen an in-house library of 560 prenyl-synthase inhibitors, developed over the last 10 years as potential anticancer drug leads and antibacterials, for their ability to inhibit P. falciparum growth inside red blood cells. Initial results showed that the most potent Plasmodium inhibitors with low toxicity to human cells were all lipophilic analogs of zoledronate and risedronate. In vitro studies confirmed that hit compounds BPH-703 and BPH-715, inhibited P. vivax GGPPS, while Plasmodium growth inhibition assays confirmed that these inhibitor effects could be rescued by treatment with geranylgeranoil.
Interestingly, the team notes, neither risedronate nor zoledronate were picked up by the screen, even though they are known to be extremely potent Plasmodium GGPPS inhibitors. In fact, the two drugs have little activity in P. falciparum growth assays, presumably because of their poor cell permeability, the researchers remark.
They subsequently carried out a P. chabaudi suppressive test to evaluate the in vivo activity of two of the lipophilic bisphosphonates, PBH-703 and BPH-811. BPH-703 is a lipophilic analog of zoledronate in which the 1-OH group is removed and the ring alkylated. BPH-811 is an analog of risedronate in which, again, the 1-OH group is removed and a lipophilic tail added.
Mice were injected with parasites, and then treated with injections of either BPH-703 or BPH-811 for four days. Initial results using relatively high doses of either compound showed that treatment with BPH-703 led to a 100% reduction in parasitaemia 100% survival, at day 14, while treatment using BPH-811 resulted in an 80% reduction in parasitemia and 80% survival. When the therapeutic dose of bisphosphonate was reduced by more than two-thirds, PBH-703 again provided the best results. Treated animals demonstrated only 8% parasitemia and still 100% survival.
“Overall, the results presented here are of interest because they show that GGPPS is a promising target for antimalarials and that the lipophilic analogs of the bone-resorption drugs zoledronate and risedronate have activity both in vitro as well as in vivo,” the authors conclude.
Notably, the lipophilic bisphosphonate inhibitors also appear to bind in a similar manner to Plasmodoum GGPPS as their parent compounds bind to both GGPPS and FPPS, and have similar enzyme activity and ΔG values for GGPPS binding, they add. “It thus seems likely that these and related lipophilic bisphosphonates will have improved activity over that already reported with more hydrophilic bisphosphonates against other parasitic protozoa, such as T. brucei, T. cruzi, Leishmania spp., T. gondii, C. parvum, and E. histolytica ... These results are of broad general interest because they indicate that it may be possible to overcome barriers to cell penetration of existing bisphosphonate drugs in this and other systems by simple covalent modification to form lipophilic analogs that retain their enzyme-inhibition activity and are also effective in vitro and in vivo.”