Sterol profiling of Leishmania parasites using a new HPLC-tandem mass spectrometry-based method and antifungal azoles as chemical probes reveals a key intermediate sterol that supports a branched ergosterol biosynthetic pathway.
Sterol profiling of Leishmania parasites using a new HPLC-tandem mass spectrometry-based method and antifungal azoles as chemical probes reveals a key intermediate sterol that supports a branched ergosterol biosynthetic pathway.
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DOI：
10.1016/j.ijpddr.2022.07.003
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发表时间：
2022-12
影响因子：
4
通讯作者：
Wang, Michael Zhuo
中科院分区：
文献类型：
--
作者：
Feng, Mei;Jin, Yiru;Yang, Sihyung;Joachim, Arline M.;Ning, Yu;Mori-Quiroz, Luis M.;Fromm, Jacob;Perera, Chamani;Zhang, Kai;Werbovetz, Karl A.;Wang, Michael Zhuo
人类利什曼病是由利什曼原虫寄生虫引起的传染病。目前针对这种致命疾病的化疗方案有很大的局限性。麦角甾醇生物合成途径已被确定为利什曼原虫的药物靶标。然而，据报道，抑制麦角甾醇生物合成的抗真菌唑类药物在治疗利什曼病方面的功效存在显着差异。为了更好地了解利什曼原虫的甾醇生物合成途径并阐明抗真菌唑类药物差异功效的机制，我们开发了一种新的 LC-MS/MS 方法来研究三种利什曼原虫前鞭毛体的甾醇谱，包括两种利什曼原虫、一种利什曼原虫、一种利什曼原虫菌株和一种利什曼原虫菌株。不同的前体离子质量和 LC 保留时间的组合允许使用新开发的 LC-MS/MS 方法特异性检测羊毛甾醇和麦角甾醇之间的 16 种中间甾醇。尽管泊沙康唑和氟康唑都是已知的真菌羊毛甾醇 14α-去甲基酶 (CYP51) 抑制剂，但只有泊沙康唑导致经唑类处理的杜氏乳杆菌前鞭毛体中羊毛甾醇大量积累。此外，当这些寄生虫用泊沙康唑和氟康唑处理时，关键中间体甾醇分别积累了40倍和7倍，通过高分辨率质谱和核磁共振波谱测定为4α,14α-二甲基酵母甾醇。 4α,14α-二甲基酵母甾醇的鉴定支持利什曼原虫中的分支麦角甾醇生物合成途径，其中羊毛甾醇 C4- 和 C14- 去甲基化反应并行而不是顺序发生。我们的结果表明，选择性抑制利什曼原虫 CYP51 不足以有效防止寄生虫生长，可能需要 CYP51 和未知甾醇 C4 去甲基酶的双重抑制剂才能获得最佳抗寄生虫效果。据报道，抗真菌唑类药物治疗利什曼病的疗效存在显着差异。开发了一种新的 LC-MS/MS 方法来研究三种利什曼原虫前鞭毛体的甾醇谱。通过质谱和核磁共振波谱测定，关键中间体甾醇为4α,14α-二甲基酵母甾醇。 4α,14α-二甲基酵母甾醇的鉴定支持利什曼原虫中的分支麦角甾醇生物合成途径。为了获得最佳的抗利什曼效应，可能需要同时抑制 CYP51 和未知的甾醇 C4 去甲基酶。
Human leishmaniasis is an infectious disease caused by Leishmania protozoan parasites. Current chemotherapeutic options against the deadly disease have significant limitations. The ergosterol biosynthetic pathway has been identified as a drug target in Leishmania. However, remarkable differences in the efficacy of antifungal azoles that inhibit ergosterol biosynthesis have been reported for the treatment of leishmaniasis. To better understand the sterol biosynthetic pathway in Leishmania and elucidate the mechanism underlying the differential efficacy of antifungal azoles, we developed a new LC-MS/MS method to study sterol profiles in promastigotes of three Leishmania species, including two L. donovani, one L. major and one L. tarentolae strains. A combination of distinct precursor ion masses and LC retention times allowed for specific detection of sixteen intermediate sterols between lanosterol and ergosterol using the newly developed LC-MS/MS method. Although both posaconazole and fluconazole are known inhibitors of fungal lanosterol 14α-demethylase (CYP51), only posaconazole led to a substantial accumulation of lanosterol in azole-treated L. donovani promastigotes. Furthermore, a key intermediate sterol accumulated by 40- and 7-fold when these parasites were treated with posaconazole and fluconazole, respectively, which was determined as 4α,14α-dimethylzymosterol by high resolution mass spectrometry and NMR spectroscopy. The identification of 4α,14α-dimethylzymosterol supports a branched ergosterol biosynthetic pathway in Leishmania, where lanosterol C4- and C14-demethylation reactions occur in parallel rather than sequentially. Our results suggest that selective inhibition of leishmanial CYP51 is insufficient to effectively prevent parasite growth and dual inhibitors of both CYP51 and the unknown sterol C4-demethylase may be required for optimal antiparasitic effect. Remarkable differences in the efficacy of antifungal azoles have been reported for the treatment of leishmaniasis. A new LC-MS/MS method has been developed to study sterol profiles in promastigotes of three Leishmania species. A key intermediate sterol was determined as 4α,14α-dimethylzymosterol by mass spectrometry and NMR spectroscopy. The identification of 4α,14α-dimethylzymosterol supports a branched ergosterol biosynthetic pathway in Leishmania. Dual inhibition of both CYP51 and the unknown sterol C4-demethylase may be required for optimal antileishmanial effect.
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影响因子：
3.8
作者：
Hailu A;Musa A;Wasunna M;Balasegaram M;Yifru S;Mengistu G;Hurissa Z;Hailu W;Weldegebreal T;Tesfaye S;Makonnen E;Khalil E;Ahmed O;Fadlalla A;El-Hassan A;Raheem M;Mueller M;Koummuki Y;Rashid J;Mbui J;Mucee G;Njoroge S;Manduku V;Musibi A;Mutuma G;Kirui F;Lodenyo H;Mutea D;Kirigi G;Edwards T;Smith P;Muthami L;Royce C;Ellis S;Alobo M;Omollo R;Kesusu J;Owiti R;Kinuthia J;Leishmaniasis East Africa Platform (LEAP) group
通讯作者：
Leishmaniasis East Africa Platform (LEAP) group
影响因子：
5.2
作者：
de Macedo Silva, Sara Teixeira;Visbal, Gonzalo;Fernandes Rodrigues, Juliany Cola
通讯作者：
Fernandes Rodrigues, Juliany Cola
影响因子：
3.5
作者：
Delfin, Dawn A.;Morgan, Rachel E.;Werbovetz, Karl A.
通讯作者：
Werbovetz, Karl A.
影响因子：
--
作者：
de Menezes JP;Guedes CE;Petersen AL;Fraga DB;Veras PS
通讯作者：
Veras PS
影响因子：
3.1
作者：
Fritsche, Claudia;Sitz, Mandy;Pohl, Hans-Dieter
通讯作者：
Pohl, Hans-Dieter