Anticoagulant and anticancer activities of Hirudo sulukii saliva extract (HSSE): Inhibition of thrombin and PI3K/AKT/mTOR signaling in prostate cancer cell lines
European Journal of Integrative Medicine, cilt.87, 2026 (SCI-Expanded, Scopus)
- Yayın Türü: Makale / Tam Makale
- Cilt numarası: 87
- Basım Tarihi: 2026
- Doi Numarası: 10.1016/j.eujim.2026.102744
- Dergi Adı: European Journal of Integrative Medicine
- Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus, EMBASE
- Anahtar Kelimeler: Cytotoxicity, Hirudo sulukii, Leech saliva extract, Prostate cancer, PI3K/AKT/mTOR pathway, Thrombin inhibition
- Erciyes Üniversitesi Adresli: Evet
Özet
Introduction: Leech saliva is a rich natural source of bioactive molecules with well-established anticoagulant properties and emerging anticancer potential. Hirudo sulukii (H. sulukii), a recently identified Turkish leech species, remains largely uncharacterized. Understanding the biological activity of H. sulukii saliva extract (HSSE) may offer novel therapeutic options for thrombotic disorders and cancer, particularly prostate cancer, in which dysregulation of the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mechanistic target of rapamycin (mTOR) signaling pathway (PI3K/AKT/mTOR signaling) plays a critical role. This study aimed to evaluate the thrombin inhibitory activity and cytotoxic effects of HSSE in human prostate cancer cell lines. Methods: HSSE was prepared by extracting saliva from adult H. sulukii specimens maintained under controlled laboratory conditions, followed by filtration and lyophilization. Thrombin inhibition was assessed using a fluorometric thrombin activity assay, with hirudin serving as a reference standard. Cytotoxicity was evaluated in LNCaP and DU-145 prostate cancer cells using a real-time cell viability monitoring system (xCELLigence) over 48 h at various HSSE concentrations. Western blot analysis was performed to examine the expression of key signaling proteins involved in the PI3K/AKT/mTOR signaling pathway. Results: HSSE demonstrated potent thrombin inhibitory activity, with an IC₅₀ value of 21.99 ± 0.73 µg/mL, corresponding to 28.65 ± 0.81 antithrombin units (ATU)/mg. It also exerted dose-dependent cytotoxicity, with DU-145 cells showing greater sensitivity (IC₅₀ = 31.00 ± 1.63 µg/mL) than LNCaP cells (IC₅₀ = 71.08 ± 1.88 µg/mL). Western blot results revealed downregulation of phosphorylated AKT, proline-rich AKT substrate of 40 kDa (PRAS40), and ribosomal protein S6 kinase beta-1 (p70S6K), alongside upregulation of the tumor suppressor Forkhead box O3 (FOXO3). Conclusion: These findings suggest that HSSE inhibits thrombin activity and suppresses the PI3K/AKT/mTOR signaling pathway in prostate cancer cells. These findings highlight the therapeutic potential of HSSE as a promising natural candidate for the development of anticoagulant and anticancer agents. The isolation and characterization of bioactive compounds, in vivo evaluation of anticancer activity, and investigation of HSSE effects on cardiovascular diseases are strongly recommended for future studies.