Document Type

Dissertation - Open Access

Award Date


Degree Name

Doctor of Philosophy (PhD)

Department / School

Chemistry and Biochemistry

First Advisor

Fathi T. Halaweish


Cucurbitacins (CUCS) are natural products with highly oxygenated tetracyclic triterpenes produced mostly by Cucurbitaceae family plant. They are known for their therapeutic efficiency with different biological activities, such as anti-inflammatory, hepatoprotective and anti-cancer. Hepatocellular carcinoma (HCC) is the third leading cause of death worldwide. Previous reports have shown the ability of CUCS to inhibit the growth of hepatocellular carcinoma cell lines (HepG-2) significantly. Structural activity relationship studies suggested the potential of the 23, 24 enone side chain of CUCS to bind to the Epidermal Growth Factor Receptor (EGFR). Due to the limited quantities of CUCS upon isolation and the challenges of total synthesis of CUCS, therefore estrone skeleton were used as a starting scaffold to synthesize CUCS-inspire estrone analogues (CIEA) targeting HCC. Molecular docking study of cucurbitacin-inspired estrone analogs was conducted using 1M17 (EGFR receptor) co-crystallized with Erolitinib (known EGFR HCC anti-cancer chemotherapeutic drug) and several analogs were identified from the docking study and were processed for multiple steps organic synthesis. Novel CUCUSinspired estrone analogs with aliphatic enone side chain such as MMA102, MMA132 were synthesized by installing the CUCS side chain at C17 of estrone scaffold. In addition, various CUCUS-inspired estrone analogs with different aliphatic, aromatic and heterocyclic pharmacophores at C-3, C-16 and C-25 were synthesized. The novel analogs showed a comparable affinity to EGFR receptor based on the docking study and improved binding through hydrophobic filling of the binding of EGFR pocket and hydrogen bonding interactions. Cell proliferation inhibition assay results demonstrated the ability of analogues MMA102 and MMA132 to inhibit HCC cell line (HepG2) proliferation with IC50 3μM and 2μM, respectively in comparison to Erlotinib IC50 (of 25 μM). Western blot experiments proved that compounds MMA132 has the ability to bind to the EGFR-TK and inhibit its phosphorylation by 90%. Flow-cytometry/cell cycle arrest study showed that our lead compound MMA132 induce a significant change in G1/S phases at different IC50 concentrations (1μM, 2μM and 3μM). The novel synthesized cucurbitacin-inspired estrone analogs showed a significant 12 times more cytotoxicity than standard chemotherapy and bind through hydrogen bonding to the same amino acids that Erlotinib binds to in the 1M17 EGFR pocket. Compound MMA311, which consist of 2-bromothiophene enone side chain along with methoxy at C-3 and double bond at C-16 and C-17, showed a significant and potent anti-proliferation activity with IC50 value of 0.7 μM. Due to the high hydrophobic character of the first set of compounds and the expected metabolism of estrone by Odealkylation of methoxy group at C-3, which may trigger estrone side effects, C-3 methoxy group were substituted with sulfamoyl moiety to improve the pharmacokinetic profile of the synthesized analogues. Several CIEA analogs that contain sulfamoyl group at C-3 and various aliphatic, aromatic and heterocyclic enone side chains at C-17 were synthesized and biologically tested as EGFR inhibitors. Compounds that contain heterocyclic enone side chains at C-17 along with sulfamoyl moiety or hydroxyl group at C-3 such as MMA297, MMA314, MMA313 and MMA312 showed an outstanding cytotoxicity with IC50 1 μM, 1.5 μM, 9 μM and 8 μM; respectively, in comparison to the Erlotinib IC50 of 25 μM. The potential of our novel CIEA to overcome cancer resistance to current chemotherapy was explored and identified their ability to decrease the drug resistance by inhibiting MRP1 utilizing high-content based assay in presence of calcein-AM as MRP1 substrate. Compounds MMAmix, MMA242, MMA132, MMA335, MMA337 and MMA320 showed potential inhibitory activity on MRP1 with inhibitory activity of 70%, 63%, 46.2%, 46%, 30% and 22%; respectively, in comparison to MK-571, which known MRP1 inhibitor. Our study demonstrated the design, synthesis of novel CIEA analogs of potent antiproliferation/ anticancer activities toward hepatocellular carcinoma and potential application to overcome cancer resistance to current chemotherapeutic agents.

Library of Congress Subject Headings

Liver -- Cancer -- Treatment.


Includes bibliographical references



Number of Pages



South Dakota State University



Rights Statement

In Copyright