Synthesis of dihydronaphthalene analogues inspired by combretastatin A-4 and their biological evaluation as anticancer agents

Abstract

The natural products colchicine and combretastatin A-4 (CA4) have provided inspiration for the discovery and development of a wide array of derivatives and analogues that inhibit tubulin polymerization through a binding interaction at the colchicine site on β-tubulin. A water-soluble phosphate prodrug salt of CA4 (referred to as CA4P) has demonstrated the ability to selectively damage tumor-associated vasculature and ushered in a new class of developmental anticancer agents known as vascular disrupting agents (VDAs). Through a long-term program of structure activity relationship (SAR) driven inquiry, we discovered that the dihydronaphthalene molecular scaffold provided access to small-molecule inhibitors of tubulin polymerization. In particular, a dihydronaphthalene analogue bearing a pendant trimethoxy aryl ring (referred to as KGP03) and a similar aroyl ring (referred to as KGP413) were potent inhibitors of tubulin polymerization (IC₅₀ = 1.0 and 1.2 μM, respectively) and displayed low nM cytotoxicity against human cancer cell lines. In order to enhance water-solubility for in vivo evaluation, the corresponding phosphate prodrug salts (KGP04 and KGP152, respectively) were synthesized. In a preliminary in vivo study in a SCID-BALB/c mouse model bearing the human breast tumor MDA-MB-231-luc, a 99% reduction in signal was observed with bioluminescence imaging (BLI) 4 h after IP administration of KGP152 (200 mg kg^-1) indicating reduced tumor blood flow. In a separate study, disruption of tumor-associated blood flow in a Fischer rat bearing an A549-luc human lung tumor was observed by color Doppler ultrasound following administration of KGP04 (15 mg kg^-1).

Fig. 1. Representative small-molecule inhibitors of tubulin assembly that bind to the colchicine site; including the combretastatin analogues (CA4, CA1),, dihydronaphthalene analogues (KGP03 and KGP413), isocombretastatin analogue phenstatin,, and indole analogue (OXi8006), along with the corresponding phosphate salt prodrugs.

Scheme 1. Synthesis of KGP03 from 6-methoxytetralin featuring early installation of phenolic moiety.

Scheme 2. Efficient alternative synthesis of KGP03 from 6-methoxytetralone.

Scheme 3. Synthesis of KGP413 from 6-methoxytetralin featuring a modified Shapiro coupling reaction.

Scheme 4. Efficient alternatives synthesis of KGP413 from 6-methoxytetralone featuring a Weireb amide coupling reaction.

Scheme 5. Synthesis of KGP04 and KGP152 phosphate prodrug salt.

Fig. 2. BLI monitoring of tumor response to KGP152. KGP152 (200 mg kg^–1) was administered IP to a BALB/C SCID mouse with a large MDA-MB-231-luc orthotopic breast tumor (2 cm³). (A) Images show signal intensity heat maps overlaid on gray scale photographs of mouse at successive time points. Following KGP152 administration, the BLI signal was dramatically lower at 4 h (99% signal reduction) and recovered slightly at 24 h (81% signal reduction from baseline). (B) Corresponding dynamic time courses of total flux obtained following luciferin administration at the three time points.

Fig. 3. Color Doppler ultrasound images show reduction of tumor blood perfusion. Images show baseline and 2 h after the administration of KGP04 (15 mg kg^–1, IP) to a nude rat with a subcutaneous A549 human tumor xenograft (0.5 cm³). Red and blue indicate magnitude of flow into and out of plane. The layer of skin is apparent at the top of the images, together with structural heterogeneity in the tumor. Vascular shutdown is indicated by yellow arrow.