A Myristoyl Amide Derivative of Doxycycline Potently Targets Cancer Stem Cells (CSCs) and Prevents Spontaneous Metastasis, Without Retaining Antibiotic Activity

Abstract

This study details the synthesis and biological evaluation of Doxy-Myr, a novel derivative of Doxycycline designed to selectively target cancer stem cells (CSCs). By attaching a myristic acid moiety to 9-amino-Doxycycline, researchers enhanced the compound's potency against CSCs, achieving over fivefold increased efficacy compared to the parent compound. Doxy-Myr demonstrated significant inhibition of CSC propagation in vitro and effectively prevented metastasis in vivo, without affecting the viability of normal cells or exhibiting antibiotic activity.

Key Findings

• Enhanced Potency: Doxy-Myr exhibited an IC₅₀ of 3.46 μM in inhibiting 3D mammosphere formation of MCF7 CSCs, compared to 18.1 μM for Doxycycline, indicating over fivefold increased potency.
• Selective Targeting: The compound did not affect the viability of MCF7 cells or normal fibroblasts grown as 2D monolayers, highlighting its selectivity for CSCs.
• Lack of Antibiotic Activity: Doxy-Myr showed no inhibitory effects on the growth of gram-negative (E. coli) and gram-positive (S. aureus) bacteria, suggesting a reduced risk of promoting antibiotic resistance.
• In Vivo Efficacy: In a chick embryo model, Doxy-Myr significantly inhibited metastasis of MDA-MB-231 breast cancer cells without affecting primary tumor growth or causing toxicity.
• Structure-Activity Relationship: Analogues with shorter (lauric acid) or longer (palmitic acid) fatty acid chains were less effective, indicating the 14-carbon myristic acid provides optimal activity.

Methodology

Researchers synthesized Doxy-Myr by covalently attaching a myristic acid moiety to 9-amino-Doxycycline. The compound's efficacy was assessed using 3D mammosphere assays to evaluate CSC propagation. Selectivity was tested on MCF7 cells and normal fibroblasts in 2D cultures. Antibiotic activity was evaluated against E. coli and S. aureus. In vivo efficacy was determined using a chick embryo chorioallantoic membrane (CAM) model to assess metastasis and toxicity.

Conclusion

Doxy-Myr represents a promising therapeutic candidate for targeting CSCs and preventing metastasis. Its enhanced potency, selectivity, lack of antibiotic activity, and efficacy in vivo suggest potential for further development as an anti-cancer agent with minimized risk of antibiotic resistance.