Novel lactate dehydrogenase inhibitors with in vivo efficacy against Cryptosporidium parvum

Cryptosporidium parvum is a highly prevalent zoonotic and anthroponotic protozoan parasite that causes a diarrheal syndrome in children and neonatal livestock, culminating in growth retardation and mortalities. Despite the high prevalence of C. parvum, there are no fully effective and safe drugs for treating infections, and there is no vaccine. We have previously reported that the bacterial-like C. parvum lactate dehydrogenase (CpLDH) enzyme is essential for survival, virulence and growth of C. parvum in vitro and in vivo. In the present study, we screened compound libraries and identified inhibitors against the enzymatic activity of recombinant CpLDH protein in vitro. We tested the inhibitors for anti-Cryptosporidium effect using in vitro infection assays of HCT-8 cells monolayers and identified compounds NSC158011 and NSC10447 that inhibited the proliferation of intracellular C. parvum in vitro, with IC50 values of 14.88 and 72.65 μM, respectively. At doses tolerable in mice, we found that both NSC158011 and NSC10447 consistently significantly reduced the shedding of C. parvum oocysts in infected immunocompromised mice’s feces, and prevented intestinal villous atrophy as well as mucosal erosion due to C. parvum. Together, our findings have unveiled promising anti-Cryptosporidium drug candidates that can be explored further for the development of the much needed novel therapeutic agents against C. parvum infections.

微小隐孢子虫（Cryptosporidium parvum）是一种兼具动物传人与传人动物特性的高流行率人畜共患原生动物寄生虫，可引发儿童与新生家畜罹患腹泻综合征，最终导致宿主生长迟缓乃至死亡。尽管该寄生虫感染的流行率极高，但目前尚无完全有效且安全的治疗药物，亦无可用疫苗。我们此前的研究表明，类细菌微小隐孢子虫乳酸脱氢酶（CpLDH）对该虫在体内外的存活、致病力与生长均至关重要。本研究通过筛选化合物库，体外鉴定出可抑制重组CpLDH蛋白酶活性的抑制剂。我们采用HCT-8细胞单层体外感染实验评估上述抑制剂的抗隐孢子虫效果，筛选出NSC158011与NSC10447两种化合物，二者可在体外抑制细胞内微小隐孢子虫的增殖，半数抑制浓度（IC50）分别为14.88 μM与72.65 μM。在小鼠可耐受的给药剂量下，研究发现NSC158011与NSC10447均能持续显著降低感染免疫缺陷小鼠粪便中微小隐孢子虫卵囊的排出量，同时可缓解该感染引发的小肠绒毛萎缩与黏膜糜烂。综上，本研究发现了极具潜力的抗隐孢子虫候选药物，可为亟需的微小隐孢子虫感染新型治疗药物的开发提供可行方向。