The quest for a novel anti-echinococcal drug

Echinococcosis is an often fatal zoonotic disease caused by infection with the larval stage of tapeworms from the genus Echinococcus. These parasitic worms are found across much of the globe and are particularly prevalent in the northern hemisphere, such as Europe, Central and Eastern Asia, and North America.

The life cycle of Echinococcus is a relatively complex one, involving a (usually) herbivorous mammal as an intermediate host and a carnivorous mammal, such as dogs or foxes, as the definitive host.

Humans are accidental intermediate hosts for Echinococcus, with infections occurring through ingestion of food or water containing parasite eggs, by direct contact with contaminated soil, or contact with infected animal hosts.

The two main types of Echinococcosis that appear in humans are the cystic form (CE) caused by E. granulosus, and the alveolar form (AE), caused by E. multilocularis.

Life cycle of Echinococcus multilocularis, the tapeworm responsible for Alveolar Echinococcosis. Source: CDC, Public Domain

Upon infection with E. multiolcularis, the parasite migrates to the liver and develops into the larval form (metacestode), that proliferates indefinitely. Proliferation results in multiplying parasitic tumour-like masses that can invade other tissues, such as the lungs and brain. AE poses a great threat to public health and is considered the be the most lethal helminthiasis, with a mortality rate of > 90% within 10 years if left untreated.

Unfortunately, once patients are diagnosed the disease has often progressed to the advanced stage and requires invasive surgery to remove the malignant parasitic mass, or prolonged chemotherapy with benzimidazoles. These drugs are parasitostatic, and thus have limited efficacy and often require repeat treatments. Therefore, novel drugs are necessary to overcome these deficiencies, and improved treatment of AE.

Crocus sativus, the wildly cultivated Saffron crocus is a source of crocin. Source: Wikimedia Commons, By KENPEI, CC BY-SA 3.0

A recent study by Liu and colleagues looking into the use of the compound Crocin to treat AE has returned some promising findings.

Crocus sativus & Crocin

Crocin is a naturally occurring carotenoid found in the plant Crocus sativus (or saffron), primarily responsible for the plants/spices vivid colour. Liu et al investigated the use of Crocin as a possible anti-echinococcal compound due to its demonstrated antioxidant, antitumour, antiangiogenic, anti-inflammatory, antibacterial and antiparasitic effects, as well as its liver, gallbladder and neural protective properties.

Assessment of crocin activity against E. multilocularis

The anti-echinococcal effects of crocin were tested against the metacestodes, germinal cell layer and protoscoleces of E. multilocularis in vitro, as well as its in vivo efficacy in infected mice. These three parasite tissues were targeted due to their role in the proliferation and survival of the parasite.

In vitro

In vitro analyses revealed that crocin was significantly toxic to the protoscoleces and germinal cells of E. multiocularis metacestodes. At 10μM concentration, crocin significantly inhibited germinal cell proliferation, reducing cell viability by ~45% and causing the germinal cell layer to detach from the metacestode, resulting in substantial structural damage.

Across a 7-day treatment at the same concentration, crocin was also shown to have killed ~60% of protoscoleces, thereby demonstrating a promising parasiticidal effect. Overall, crocin was determined to have a median effective concentration (EC50) of 11.36μM against E. multilocularis metacestodes.

In vivo

In the infected mice, the effectiveness of crocin was determined by analysing the parasite weight before and after treatment, and showed a significant reduction (68%) at a dose of 100mg/kg. Crocin treatment again was shown to cause considerable structural damage to the germinal cell layers of the metacestode.

In addition, treatment with crocin was revealed to significantly increase expression levels of Interleukin-2 (IL-2) and IL-4, as well as inhibiting the expression of matrix metalloproteinases (MMPs) and promoting collagen deposition in host tissues surrounding the parasite.

An increase in IL-4 has been previously shown to restrict the growth of AE metacestodes, whereas an increase in IL-2 was revealed to enhance the immune response against E. multilocularis, and reduce the parasite’s ability to escape the host’s immune system. Furthermore, MMPs act to degrade the host’s extracellular matrix, thereby allowing the metacestode to grow and proliferate. Therefore, the inhibition of MMPs, along with increased collagen deposition, restricts metacestode growth and proflieration, and promotes a greater a granulomatous response to contain the parasite.

The in vitro and in vivo assessment of crocin highlighted its considerable therapeutic efficacy against Echinoccocus multilocularis.

Graphical representation of crocin action on metacestodes. Source: Liu et al., 2021.

Assessment of crocin toxicity

In addition to being highly effective at killing the parasite, drugs must also be safe for the patient/host. The researchers tested its toxicity against Human foreskin fibroblast (HFFs) and Reuber rat hepatoma (RH) cells (in vitro), as well as after 6 weeks in the crocin-treated mice (in vivo).

In vitro, crocin was deemed to be about half as toxic to HFFs and RH cell lines than to E. multilocularis metacestodes and germinal cells. As for its in vivo toxicity, no adverse reactions were observed during treatment, nor did it alter blood indices, and upon histological analysis, no damage was seen in liver or kidneys of crocin-treated mice.

Crocin: a promising new anthelmintic?

In this work Liu and colleagues demonstrated the considerable parasiticidal activity of crocin against E. multilocularis in vitro and in vivo, as well as its limited toxicity against mammalian cell lines and treated mice. In particular, crocin demonstrated a proective effect against AE-induced liver and kidney fibrosis.

The authors suggest that the immune effects of crocin-treatment of AE need further study, but nonetheless, crocin seems to be a highly promising alternative drug for the treatment of AE that requires further investigation and development.

View the latest posts on the BugBitten homepage