Discovery of natural product scaffolds with anthelmintic activity against Haemonchus contortus

Abstract

The present thesis focused on discovering new candidate compounds in natural products against economically important parasitic worms of livestock to circumvent current problems associated with resistance to commercially-available drugs (anthelmintics) (cf. Chapter 1). One of the most pathogenic parasites of small ruminants, Haemonchus contortus (barber’s pole worm), was used as the screening tool. This whole-organism bioassay was used to identify active compounds/extracts that reduced motility and/or development or altered the morphology (phenotype) of H. contortus larvae in vitro. Selected compounds were purified from active extracts using a bioassay-guided fractionation approach.

The screening of two compound libraries containing a total of 1,000 natural products, natural product-inspired or synthetic compounds identified 34 compounds with notable activity against H. contortus (Chapters 2 and 3). Most of the ‘hits’ (n=32) identified from the first library were analogues of arylpyrrole (a natural product scaffold), while deguelin and rotenone (two plant rotenoids) were identified from the second library. Encouraged by these findings, a library of 7,500 extracts from different plant species was screened (Chapter 4). This screen identified three active extracts from the leaves and roots of Cryptocarya novoguineensis and the roots of Piper methysticum. Bioassay-guided fractionation of active extracts yielded four known alpha-pyrones, namely goniothalamin from C. novoguineensis, and dihydrokavain, desmethoxyyangonin and yangonin (= kavalactones) from P. methysticum. Three kavalactones induced a lethal ‘evisceration’ phenotype in treated larvae of H. contortus, and had limited toxicity on mammalian epithelial (MCF10A) cells. This is the first report on the activity of such natural compounds on a parasitic nematode of animals.

The work was extended to explore compounds from Australian marine species. A library of 2,000 extracts from marine invertebrates was screened (Chapter 5) and identified three active extracts from marine sponges - one from Monanchora unguiculata and two extracts from Haliclona sp. The compounds with moderate activity against H. contortus were fomiamycalin from Monanchora unguiculata and halaminol A from Haliclona sp., although the activity of these compounds was not entirely selective.

Subsequently, a structure-activity relationship (SAR) investigation of one of the identified natural product scaffolds, alpha-pyrone was conducted (Chapter 6). This work identified three analogues (designated W-408, W-415 and W-417) with anthelmintic activity similar to, or greater than the synthetic parent kavalactones (desmethoxyyangonin or yangonin), with the most potent analogue W-408 achieving a marked increase in potency (7-fold) against and selectivity (> 21) for H. contortus.

Taken together, this drug discovery effort identified or purified 40 natural products representing distinct chemical classes (Chapters 2-5), and the SAR investigation of the alpha-pyrone scaffold identified three key analogues with enhanced potency and/or selectivity (Chapter 6). Collectively, the findings of this thesis indicate that some of the identified natural product scaffolds have the potential to be developed as ‘lead’ candidates. Developing such anthelmintic candidates via future chemical optimisation, efficacy and safety assessments, broad spectrum activity assessments, and target identification represents an exciting prospect (cf. Chapter 7) and, if successful, could pave the way to subsequent pre-clinical and clinical evaluations.