Bumped Kinase Inhibitors as therapy for apicomplexan parasitic diseases: lessons learned

Choi, Ryan; Hulverson, Matthew A.; Huang, Wenlin; Vidadala, Rama Subba Rao; Whitman, Grant R.; Barrett, Lynn K.; Schaefer, Deborah A.; Betzer, Dana P.; Riggs, Michael W.; Doggett, J. Stone; Hemphill, Andrew; Ortega‐Mora, Luis Miguel; McCloskey, Molly C.; Arnold, Samuel; Hackman, Robert C.; Marsh, Kennan C.; Lynch, James J.; Freiberg, Gail; LeRoy, Bruce E.; Kempf, Dale J.; Choy, Robert K. M.; Hostos, Eugenio L. de; Maly, Dustin J.; Fan, Erkang; Ojo, Kayode K.; Voorhis, Wesley C. Van

doi:10.1016/j.ijpara.2020.01.006

Cited by 40 publications

(34 citation statements)

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“…The genes coding for these kinases originate from plants [7], and therefore constitute interesting drug targets. During the last decade, CDPK1 has been an important focus as a target for drug development against a wide range of apicomplexans such as Plasmodium falciparum, Toxoplasma gondii, Neospora caninum, Sarcocystis neurona, Besnoitia besnoiti, Babesia bovis, Theileria equi and Cryptosporidium parvum [7,8]. Bumped kinase inhibitors (BKIs) are ATP-competitive kinase inhibitors that target CDPK1 in different apicomplexans [9].…”

Section: Introductionmentioning

confidence: 99%

Neospora caninum: Differential Proteome of Multinucleated Complexes Induced by the Bumped Kinase Inhibitor BKI-1294

et al. 2020

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Background: the apicomplexan parasite Neospora caninum causes important reproductive problems in farm animals, most notably in cattle. After infection via oocysts or tissue cysts, rapidly dividing tachyzoites infect various tissues and organs, and in immunocompetent hosts, they differentiate into slowly dividing bradyzoites, which form tissue cysts and constitute a resting stage persisting within infected tissues. Bumped kinase inhibitors (BKIs) of calcium dependent protein kinase 1 are promising drug candidates for the treatment of Neospora infections. BKI-1294 exposure of cell cultures infected with N. caninum tachyzoites results in the formation of massive multinucleated complexes (MNCs) containing numerous newly formed zoites, which remain viable for extended periods of time under drug pressure in vitro. MNC and tachyzoites exhibit considerable antigenic and structural differences. Methods: Using shotgun mass spectrometry, we compared the proteomes of tachyzoites to BKI-1294 induced MNCs, and analyzed the mRNA expression levels of selected genes in both stages. Results: More than half of the identified proteins are downregulated in MNCs as compared to tachyzoites. Only 12 proteins are upregulated, the majority of them containing SAG1 related sequence (SRS) domains, and some also known to be expressed in bradyzoites Conclusions: MNCs exhibit a proteome different from tachyzoites, share some bradyzoite-like features, but may constitute a third stage, which remains viable and ensures survival under adverse conditions such as drug pressure. We propose the term “baryzoites” for this stage (from Greek βαρυσ = massive, bulky, heavy, inert).

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Section: Introductionmentioning

confidence: 99%

Neospora caninum: Differential Proteome of Multinucleated Complexes Induced by the Bumped Kinase Inhibitor BKI-1294

et al. 2020

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“…These apicomplexan CDPK1 proteins have a unique glycine 'gate-keeper' residue that confers selectivity over human kinases [88,89]; the inhibitors are termed 'bumped kinase inhibitors,' or BKIs, as they do not bind in the human ATP-binding domain. Lead optimization strategies on BKIs over the past decade have resulted in preclinical leads from three BKI scaffolds with in vivo efficacy, but varying safety and tolerability [90][91][92][93].…”

Section: Repositioning Validated Parasite Drug Targetsmentioning

confidence: 99%

Phenotypic screening techniques forCryptosporidiumdrug discovery

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McNamara

2020

Expert Opinion on Drug Discovery

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Introduction: Two landmark epidemiological studies identified Cryptosporidium spp. as a significant cause of diarrheal disease in pediatric populations in resource-limited countries. Notably, nitazoxanide is the only approved drug for treatment of cryptosporidiosis but shows limited efficacy. As a result, many drug discovery efforts have commenced to find improved treatments. The unique biology of Cryptosporidium presents challenges for traditional drug discovery methods, which has inspired new assay platforms to study parasite biology and drug screening. Areas covered: The authors review historical advancements in phenotypic-based assays and techniques for Cryptosporidium drug discovery, as well as recent advances that will define future drug discovery. The reliance on phenotypic-based screens and repositioning of phenotypic hits from other pathogens has quickly created a robust pipeline of potential cryptosporidiosis therapeutics. The latest advances involve new in vitro culture methods for oocyst generation, continuous culturing capabilities, and more physiologically relevant assays for testing compounds. Expert opinion: Previous phenotypic screening techniques have laid the groundwork for recent cryptosporidiosis drug discovery efforts. The resulting improved methodologies characterize compound activity, identify, and validate drug targets, and prioritize new compounds for drug development. The most recent improvements in phenotypic assays are poised to help advance compounds into clinical development.

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“…Calcium-dependent protein kinases (CDPKs) are especially interesting drug targets in that homologs of CDPKs are not found in mammals. One of these kinases is CDPK1, homologs of which are intensively studied in target-based drug development for a wide range of apicomplexans including Plasmodium, Cryptosporidium, Eimeria, Babesia, Theileria, Toxoplasma, and Neospora among others [reviewed by (12,13)]. In N. caninum and T. gondii, CDPK1 is essential for motility, host cell invasion, and egress of tachyzoites from the host cell, through its role in the signaling events that control microneme secretion (14,15).…”

Section: Introductionmentioning

confidence: 99%

The Impact of BKI-1294 Therapy in Mice Infected With the Apicomplexan Parasite Neospora caninum and Re-infected During Pregnancy

et al. 2020

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Exposure of Neospora caninum tachyzoites to BKI-1294 in vitro results in the formation of long-lived multinucleated complexes (MNCs). However, in vivo treatment of BALB/c mice with BKI-1294 shortly after N. caninum infection during pregnancy was safe and profoundly reduced pup mortality and vertical transmission. We hypothesized that the formation of MNCs could trigger immune responses that contribute to BKI efficacy in vivo. In this study, mice were first vaccinated with a sublethal dose of N. caninum tachyzoites and were treated with BKI-1294. We then investigated the effects of these treatments after mating and re-infection during pregnancy. Effects on fertility, pup survival, vertical transmission, and parasite load in dams were evaluated. Cytokines in sera or splenocyte culture supernatants were assessed by either ELISA or the Luminex TM 200 system, and humoral immune responses against tachyzoite and MNC antigens were compared by ELISA, Western blotting and immunoproteomics. Our results showed that BKI-1294 treatment of live-vaccinated mice reduced the cerebral parasite load in the dams, but resulted in higher neonatal pup mortality and vertical transmission. In live-vaccinated mice, cytokine levels, most notably IFN-y, IL-10, and IL-12, were consistently lower in BKI-1294 treated animals compared to non-treated mice. In addition, comparative Western blotting identified two protein bands in MNC extracts that were only recognized by sera of live-vaccinated mice treated with BKI-1294, and were not found in tachyzoite extracts. We conclude that treatment of live-vaccinated mice with BKI-1294 influenced the cellular and humoral immune responses against infection, affected the safety of the live-vaccine, and decreased protection against re-infection and vertical transmission during pregnancy.

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Bumped Kinase Inhibitors as therapy for apicomplexan parasitic diseases: lessons learned

Cited by 40 publications

References 54 publications

Neospora caninum: Differential Proteome of Multinucleated Complexes Induced by the Bumped Kinase Inhibitor BKI-1294

Neospora caninum: Differential Proteome of Multinucleated Complexes Induced by the Bumped Kinase Inhibitor BKI-1294

Phenotypic screening techniques forCryptosporidiumdrug discovery

The Impact of BKI-1294 Therapy in Mice Infected With the Apicomplexan Parasite Neospora caninum and Re-infected During Pregnancy

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