Evaluation of in vitro antileishmanial efficacy of cyclosporin A and its non-immunosuppressive derivative, dihydrocyclosporin A

Abstract

Background: New therapeutic drugs are urgently needed against visceral leishmaniasis because current drugs, such as pentavalent antimonials and miltefosine, produce severe side effects and development of resistance. Whether cyclosporine A (CsA) and its derivatives can be used as therapeutic drugs for visceral leishmaniasis has been controversial for many years. Methods:In this study, we evaluated the efficacy of CsA and its derivative, dihydrocyclosporin A (DHCsA-d), against promastigotes and intracellular amastigotes of Leishmania donovani. Sodium stibogluconate (SSG) was used as a positive control. Results:Our results showed that DHCsA-d was able to inhibit the proliferation of L. donovani promastigotes (IC 50 : 21.24 μM and 12.14 μM at 24 h and 48 h, respectively) and intracellular amastigotes (IC 50 : 5.23 μM and 4.84 μM at 24 and 48 h, respectively) in vitro, but CsA treatment increased the number of amastigotes in host cells. Both DHCsA-d and CsA caused several alterations in the morphology and ultrastructure of L. donovani, especially in the mitochondria. However, DHCsA-d showed high cytotoxicity towards cells of the mouse macrophage cell line RAW264.7, with CC50 values of 7.98 μM (24 h) and 6.65 μM (48 h). Moreover, DHCsA-d could increase IL-12, TNF-α and IFN-γ production and decrease the levels of IL-10, IL-4, NO and H 2 O 2 in infected macrophages. On the contrary, CsA decreased IL-12, TNF-α, and IFN-γ production and increased the levels of IL-10, IL-4, NO and H 2 O 2 in infected macrophages. The expression of L. donovani cyclophilin A (LdCyPA) in promastigotes and intracellular amastigotes and the expression of cyclophilin A (CyPA) in RAW 264.7 cells were found to be significantly downregulated in the CsA-treated group compared to those in the untreated group. However, no significant changes in LdCyPA and CyPA levels were found after DHCsA-d or SSG treatment. Conclusions:Our findings initially resolved the dispute regarding the efficacy of CsA and DHCsA-d for visceral leishmaniasis treatment. CsA showed no significant inhibitory effect on intracellular amastigotes. DHCsA-d significantly inhibited promastigotes and intracellular amastigotes, but it was highly cytotoxic. Therefore, CsA and DHCsA-d are not recommended as antileishmanial drugs.

Integrative genomic, proteomic and phenotypic studies of Leishmania donovani strains revealed genetic features associated with virulence and antimony-resistance

“…Stationary phase promastigotes (~ 5.0 × 10 6 cells/per well) of HCZ isolate, DD8 and 9044 strains were added into wells, and incubated at 37 °C, 5% CO 2 for 6 h. Next, the non-infected promastigotes were removed, and the infected macrophages were incubated at 37 °C in 5% CO 2 with fresh medium without drugs for 24 h. The medium was then removed, and fresh culture medium supplemented with 10 μM SSG (cf. [ 33 ]) were added into wells for 2 days, with PBS used as control. At 24 and 48 h, the glass coverslips were fixed in methanol (Solarbio, Beijing, China) and stained with Wright’s stain (Solarbio) to calculate the number of intracellular amastigotes under light microscopy by counting 200 macrophages per slide.…”

The Antiviral Properties of Cyclosporine. Focus on Coronavirus, Hepatitis C Virus, Influenza Virus, and Human Immunodeficiency Virus Infections

“…Cyclosporine acted on promastigotes and amastigotes via the blockage of calcineurin phosphatase and cyclophilin-dependent thermotolerance, respectively [65]. Conversely, other studies proved there was no significant inhibiting effect of cyclosporine on amastigotes, and the authors did not recommend cyclosporine as an anti-leishmanial drug [66].…”

An overview of the treatment of cutaneous leishmaniasis

“…evaluated the efficacy of both drugs against promastigotes and amastigotes of L. donovani versus SSG as a positive control. Intracellular amastigotes decreased compared to the untreated group after the administration of DHCsA-d or SSG 51 . Also, the intracellular amastigotes increased after the administration of CsA, and it appears that only the last was found to promote intracellular amastigotes 51 .…”

“…Cyclosporin A (CsA) exhibits its immunosuppressive action by inhibiting the production of calcineurin through binding to cyclophilin A (CyPA). L. donovani expresses a variant of CyPA (LdCyPA) that is different to those of humans and has a role in the survival of parasites in the tissues 51 . Dihydrocyclosporin A (DHCsA-d) (a co-metabolite of CsA) has minimum immunosuppressive activity and also inhibits L. donovani in vivo and in vitro .…”