*Article*

#### **Anticancer Effects of New Ceramides Isolated from the Red Sea Red Algae** *Hypnea musciformis* **in a Model of Ehrlich Ascites Carcinoma: LC-HRMS Analysis Profile and Molecular Modeling**

**Sameh S. Elhady 1,†, Eman S. Habib 2,†, Reda F. A. Abdelhameed 3, Marwa S. Goda 2, Reem M. Hazem 4, Eman T. Mehanna 5, Mohamed A. Helal 6,7, Khaled M. Hosny 8, Reem M. Diri 9, Hashim A. Hassanean 2, Amany K. Ibrahim 2, Enas E. Eltamany 2, Usama Ramadan Abdelmohsen 10,11 and Safwat A. Ahmed 2,\***

	- Department of Medicinal Chemistry, Faculty of Pharmacy, Suez Canal University, Ismailia 41522, Egypt
	- Department of Pharmacy Practice, Faculty of Pharmacy, King Abdulaziz University, Jeddah 21589, Saudi Arabia; rdiri@kau.edu.sa

**Abstract:** Different classes of phytochemicals were previously isolated from the Red Sea algae *Hypnea musciformis* as sterols, ketosteroids, fatty acids, and terpenoids. Herein, we report the isolation of three fatty acids—docosanoic acid **4**, hexadecenoic acid **5**, and alpha hydroxy octadecanoic acid **6**—as well as three ceramides— **A** (**1**), **B** (**2**), and **C** (**3**)—with 9-methyl-sphinga-4,8-dienes and phytosphingosine bases. Additionally, different phytochemicals were determined using the liquid chromatography coupled with electrospray ionization high-resolution mass spectrometry (LC-ESI-HRMS) technique. Ceramides **A** (**1**) and **B** (**2**) exhibited promising in vitro cytotoxic activity against the human breast adenocarcinoma (MCF-7) cell line when compared with doxorubicin as a positive control. Further in vivo study and biochemical estimation in a mouse model of Ehrlich ascites carcinoma (EAC) revealed that both ceramides **A** (**1**) and **B** (**2**) at doses of 1 and 2 mg/kg, respectively, significantly decreased the tumor size in mice inoculated with EAC cells. The higher dose (2 mg/kg) of ceramide **B**(**2**) particularly expressed the most pronounced decrease in serum levels of vascular endothelial growth factor -B (VEGF-B) and tumor necrosis factor-α (TNF-α) markers, as well as the expression levels of the growth factor midkine in tumor tissue relative to the EAC control group. The highest expression of apoptotic factors, p53, Bax, and caspase 3 was observed in the same group that received 2 mg/kg of ceramide **B** (**2**). Molecular docking simulations suggested that ceramides **A** (**1**) and **B**(**2**) could bind in the deep grove between the H2 helix and the Ser240-P250 loop of p53, preventing its interaction with MDM2 and leading to its accumulation. In conclusion, this study reports the

**Citation:** Elhady, S.S.; Habib, E.S.; Abdelhameed, R.F.A.; Goda, M.S.; Hazem, R.M.; Mehanna, E.T.; Helal, M.A.; Hosny, K.M.; Diri, R.M.; Hassanean, H.A.; et al. Anticancer Effects of New Ceramides Isolated from the Red Sea Red Algae *Hypnea musciformis* in a Model of Ehrlich Ascites Carcinoma: LC-HRMS Analysis Profile and Molecular Modeling. *Mar. Drugs* **2022**, *20*, 63. https://doi.org/10.3390/ md20010063

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Academic Editors: Marco García-Vaquero and Brijesh K. Tiwari

Received: 21 December 2021 Accepted: 7 January 2022 Published: 10 January 2022

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**Copyright:** © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

cytotoxic, apoptotic, and antiangiogenic effects of ceramides isolated from the Red Sea algae *Hypnea musciformis* in an experimental model of EAC.

**Keywords:** ceramides; MCF-7; Ehrlich ascites carcinoma; VEGF-B; TNF-α; midkine; apoptosis; p53; LC-ESI-HRMS; *Hypnea musciformis*
