COP9 signalosome is an essential and druggable parasite target that regulates protein degradation

Abstract

Understanding how the protozoan protein degradation pathway is regulated could uncover new parasite biology for drug discovery. We found the COP9 signalosome (CSN) conserved in multiple pathogens such as Leishmania, Trypanosoma, Toxoplasma, and used the severe diarrhea-causing Entamoeba histolytica to study its function in medically significant protozoa. We show that CSN is an essential upstream regulator of parasite protein degradation. Genetic disruption of E. histolytica CSN by two distinct approaches inhibited cell proliferation and viability. Both CSN5 knockdown and dominant negative mutation trapped cullin in a neddylated state, disrupting UPS activity and protein degradation. In addition, zinc ditiocarb (ZnDTC), a main metabolite of the inexpensive FDA-approved globally-available drug disulfiram, was active against parasites acting in a COP9-dependent manner. ZnDTC, given as disulfiram-zinc, had oral efficacy in clearing parasites in vivo. Our findings provide insights into the regulation of parasite protein degradation, and supports the significant therapeutic potential of COP9 inhibition.

Revisiting Drug Development Against the Neglected Tropical Disease, Amebiasis

“…In the same study, the CSN5 gene was silenced using high-efficiency inducible RNA interference technology to develop knockout trophozoites which exhibited reduced proliferation and were eventually killed. Expression of dominant negative mutant of CSN5 also showed the same results ( Ghosh et al., 2020 ). Also, the drug zinc-ditiocarb (ZnDTC) was tested in vitro against wild type trophozoites and it was found to reduce their viability.…”

“…CSN5 (EHI_050500) contains a JAMM (JAB1/MPN/Mov34 metalloenzyme) motif. Other components of the signalosome complex such as CSN2 (EHI_174890), CSN1 (EHI_182890), CSN3 (EHI_103560), CSN6 (EHI_068470) were also co-purified while putative genes for CSN4 and CSN8 were identified in the E. histolytica genome ( Ghosh et al., 2020 ). In the same study, the CSN5 gene was silenced using high-efficiency inducible RNA interference technology to develop knockout trophozoites which exhibited reduced proliferation and were eventually killed.…”

“…Also, the drug zinc-ditiocarb (ZnDTC) was tested in vitro against wild type trophozoites and it was found to reduce their viability. The drug was even tested in vivo using mouse models that mimicked human amebic colitis and showed positive results for pathogen clearance ( Ghosh et al., 2020 ). The ZnDTC is already an FDA approved drug which is used for treating alcoholism and can be repurposed as an anti-amebic agent.…”

Drug Repurposing of the Alcohol Abuse Medication Disulfiram as an Anti-Parasitic Agent

“…Because proteasomemediated degradation is essential for parasite survival, this pathway has become an attractive drug target for protozoan parasites (Khare et al, 2016;Li et al, 2016). The anti-amebic activity of ZnDTC was shown to involve inhibition of the ubiquitin-proteasome pathway (Ghosh et al, 2020). E3 ubiquitin ligases catalyze the ubiquitination of proteins destined for proteasomal degradation (Cromm and Crews, 2017;Wertz and Wang, 2019).…”

“…A new preclinical study using a mouse model that simulates human amebic colitis showed that oral disulfiram combined with zinc was highly effective in clearing parasites ( Figure 1 ) ( Ghosh et al., 2020 ). Zinc-ditiocarb complex (ZnDTC) had high potency and was active against E. histolytica parasites at low nanomolar concentrations ( Ghosh et al., 2020 ), significantly below the serum and tissue level achieved with disulfiram therapy at recommended doses ( Johansson, 1992 ; Skrott et al., 2017 ), and 1000-fold-lower less than the EC 50 of metronidazole, the current drug of choice to treat amebiasis.…”