Lipid nanocarriers of a lipid-conjugated estrogenic derivative inhibit tumor growth and enhance cisplatin activity against triple-negative breast cancer: pharmacokinetic and efficacy evaluation.

Breast cancer is the leading cause of malignancies among women globally. The triple negative breast cancer (TNBC) subtype is the most difficult to treat and accounts for 15% of all cases. Targeted therapies have been developed for TNBC but come short of clinical translation due to acquired tumor resistance. An effective therapy against TNBC must combine properties of target specificity, efficient tumor killing, and translational relevance. The objective of this study was to formulate a nontoxic, cationic, lipid-conjugated estrogenic derivative (ESC8), with demonstrated anticancer activity, for oral delivery in mice bearing triple negative breast cancer (TNBC) as xenograft tumors. The in vitro cell viability, Caco-2 permeability, and cell cycle dynamics of ESC8-treated TNBC cells were investigated. ESC8 was formulated as liposomes, solid lipid nanoparticles (SLNs), and nanostructured lipid carriers (NLCs) and characterized for size, zeta potential, entrapment efficiency, size stability, and tumor biodistribution. Pharmacokinetic modeling of plasma concentration-time course data was carried out following intravenous and oral administration in Sprague-Dawley rats. In vivo efficacy investigation of ESC8-SLNC was carried out in Nu/Nu mice bearing MDA-MB-231 TNBC as xenograft tumors, and the molecular dynamics modulating tumor growth inhibition was analyzed by Western blot. In vitro ESC8 inhibited TNBC and non-TNBC cell viability with IC50 ranging from 1.81 to 3.33 µM. ESC8 was superior to tamoxifen and Cisplatin in inhibiting MDA-MB-231 cell viability; and at 2.0 µM ESC8 enhanced Cisplatin cytotoxicity 16-fold. Intravenous ESC8 (2.0 mg/kg) was eliminated at a rate of 0.048 ± 0.01 h(-1) with a half-life of 14.63 ± 2.95 h in rats. ESC8 was orally bioavailable (47.03%) as solid lipid nanoparticles (ESC8-SLN). ESC8-SLN (10 mg/kg/day, ×14 days, p.o.) inhibited breast tumor growth by 74% (P < 0.0001 vs control) in mice bearing MDA-MB-231 cells as xenografts; and when given in combination with Cisplatin (2.0 mg/kg/biweekly, ×2 weeks, IV), tumor growth was inhibited by 87% (P = 0.0002, vs ESC8-SLN; 10 mg/kg/day, ×14 days, p.o). ESC8-SLN tumor growth inhibition was associated with increased expression of p21 and Caspase-9; as well as by inhibition of EGFR, Slug, p-Akt1, Vimentin, NFkß, and IKKγ. These results show the promise of ESC8 as an oral adjuvant or neoadjuvant against triple negative breast cancer.

Development of new estradiol-cationic lipid hybrids: ten-carbon twin chain cationic lipid is a more suitable partner for estradiol to elicit better anticancer activity.

The present study illustrates the synthesis and anticancer evaluation of six, ten, twelve and fourteen carbon chain containing cationic lipidated-estradiol hybrids. Previously, we have established the lipidation strategy to introduce anticancer activities in various pharmacophores including estradiol (ES). In this structure activity study the length of the carbon chain is narrowed down between C6-C14 to screen out the most potent anticancer molecule among the class. Among the newly developed ES-cationic lipid conjugates, ten-carbon chain containing derivative, ES-C10 (5c) exhibited 4-12 folds better anticancer activity than the previously established derivative, ES-C8 (5b) in various cancer cells of different origin. Moreover cytotoxicity of this molecule was not observed in non-cancer cells. Notably, in spite of bearing estrogenic moiety, ES-C10 exhibited anticancer activity irrespective of estrogen receptor (ER) expression status. ES-C10 exhibited prominent sub-G0 arrest of cancer cells with concomitant induction of apoptosis and demonstrated significant inhibition of tumor growth in mouse melanoma model. Collectively, ES-C10 exemplifies the development of an anticancer agent with broader activity against cancer cells of different origins.

A lipid-modified estrogen derivative that treats breast cancer independent of estrogen receptor expression through simultaneous induction of autophagy and apoptosis.

It is a challenge to develop a universal single drug that can treat breast cancer at single- or multiple-stage complications, yet remains nontoxic to normal cells. The challenge is even greater when breast cancer-specific, estrogen-based drugs are being developed that cannot act against multistaged breast cancer complications owing to the cells differential estrogen receptor (ER) expression status and their possession of drug-resistant and metastatic phenotypes. We report here the development of a first cationic lipid-conjugated estrogenic derivative (ESC8) that kills breast cancer cells independent of their ER expression status. This ESC8 molecule apparently is nontoxic to normal breast epithelial cells, as well as to other noncancer cells. ESC8 induces apoptosis through an intrinsic pathway in ER-negative MDA-MB-231 cells. In addition, ESC8 treatment induces autophagy in these cells by interfering with the mTOR activity. This is the first example of an estrogen structure-based molecule that coinduces apoptosis and autophagy in breast cancer cells. Further in vivo study confirms the role of this molecule in tumor regression. Together, our results open new perspective of breast cancer chemotherapy through a single agent, which could provide the therapeutic benefit across all stages of breast cancer.

17ß-estradiol-linked nitro-L-arginine as simultaneous inducer of apoptosis in melanoma and tumor-angiogenic vascular endothelial cells.

Aggressive melanoma is commonly associated with rapid angiogenic growth in tumor mass, tumor cells acquiring apoptosis resistance, inhibition of cellular differentiation etc. Designing a single anticancer molecule which will target all these factors simultaneously is challenging. In the pretext of inciting anticancer effect through inhibiting nitric oxide synthase (NOS) via estrogen receptors (ER) in ER-expressing skin cancer cells, we developed an estrogen-linked L-nitro-arginine molecule (ESAr) for inciting anticancer effect in melanoma cells. ESAr showed specific anticancer effect through diminishing aggressiveness and metastatic behavior in melanoma cells and tumor. In comparison, ESAr showed significantly higher antiproliferative effect than parent molecule L-nitroarginine methyl ester (L-NAME, a NOS inhibitor) through induction of prominent apoptosis in melanoma cells. ESAr-pretreated aggressive melanoma cells could not form tumor possibly because of transformation/differentiation into epithelial-type cells. Furthermore, its antiangiogenic effect was demonstrated through ESAr-induced antiproliferation in HUVEC cells and apoptosis-induction in tumor-associated vascular endothelial cells, thereby significantly restricting severe growth in melanoma tumor. The targeting moiety, estrogen, at the therapeutic concentration of ESAr has apparently no effect in tumor-growth reduction. Albeit, no specific NOS-inhibition was observed, but ESAr could simultaneously induce these three cancer-specific antiaggressiveness factors, which the parent molecule could not induce. Our data rationalize and establish a new use of estrogen as a ligand for potentially targeting multiple cellular factors for treating aggressive cancers.