Bioreductively Activatable Prodrug Conjugates of Combretastatin A-1 and Combretastatin A-4 as Anticancer Agents Targeted toward Tumor-Associated Hypoxia.

The natural products combretastatin A-1 (CA1) and combretastatin A-4 (CA4) function as potent inhibitors of tubulin polymerization and as selective vascular disrupting agents (VDAs) in tumors. Bioreductively activatable prodrug conjugates (BAPCs) can enhance selectivity by serving as substrates for reductase enzymes specifically in hypoxic regions of tumors. A series of CA1-BAPCs incorporating methyl, -methyl, and dimethyl nitrothiophene triggers were synthesized together with corresponding CA4-BAPCs, previously reported by Davis ( , 5 (11), 2886), for comparison.

Structure Guided Design, Synthesis, and Biological Evaluation of Novel Benzosuberene Analogues as Inhibitors of Tubulin Polymerization.

A promising design paradigm for small-molecule inhibitors of tubulin polymerization that bind to the colchicine site draws structural inspiration from the natural products colchicine and combretastatin A-4 (CA4). Our previous studies with benzocycloalkenyl and heteroaromatic ring systems yielded promising inhibitors with dihydronaphthalene and benzosuberene analogues featuring phenolic (KGP03 and KGP18) and aniline (KGP05 and KGP156) congeners emerging as lead agents. These molecules demonstrated dual mechanism of action, functioning both as potent vascular disrupting agents (VDAs) and as highly cytotoxic anticancer agents.

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

The natural products colchicine and combretastatin A-4 () 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 (referred to as ) 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.

Clinical trial experience with CA4P anticancer therapy: focus on efficacy, cardiovascular adverse events, and hypertension management.

Combretastatin A4-phosphate (CA4P) is a vascular-disrupting agent (VDA) in clinical development for the treatment of ovarian and other cancers. In contrast to antiangiogenic agents, such as bevacizumab, which suppress the development of new tumor vasculature, VDAs target established tumor vasculature. These differing but complementary mechanisms of action are currently being explored in clinical trials combining CA4P and bevacizumab.

The development and use of vascular targeted therapy in ovarian cancer.

Combination vascular-targeted therapies (VTTs) represent a promising approach for patients with platinum-resistant recurrent ovarian cancer (OC). VTTs include two mechanistically distinct classes of agents: anti-angiogenic agents (AAs) and vascular-disrupting agents (VDAs). AAs suppress growth of new tumor vasculature through inhibition of vascular endothelial growth factor (VEGF) and other pro-angiogenic molecules.

Synthesis and Biological Evaluation of Benzocyclooctene-based and Indene-based Anticancer Agents that Function as Inhibitors of Tubulin Polymerization.

The natural products colchicine and combretastatin A-4 () have been inspirational for the design and synthesis of structurally related analogues and spin-off compounds as inhibitors of tubulin polymerization. The discovery that a water-soluble phosphate prodrug salt of (referred to as ) is capable of imparting profound and selective damage to tumor-associated blood vessels paved the way for the development of a new therapeutic approach for cancer treatment utilizing small-molecule inhibitors of tubulin polymerization that also act as vascular disrupting agents (VDAs). Combination of salient structural features associated with colchicine and led to the design and synthesis of a variety of fused aryl-cycloalkyl and aryl-heterocyclic compounds that function as inhibitors of tubulin polymerization.

Synthesis and biological evaluation of a water-soluble phosphate prodrug salt and structural analogues of KGP94, a lead inhibitor of cathepsin L.

The magnitude of expression of cathepsin L, often upregulated in the tumor microenvironment, correlates with the invasive and metastatic nature of certain tumors. Inhibition of cathepsin L represents an emerging strategy for the treatment of metastatic cancer. A potent, small-molecule inhibitor (referred to as KGP94) of cathepsin L, and new KGP94 analogues were synthesized.

Bioreductively activatable prodrug conjugates of phenstatin designed to target tumor hypoxia.

A variety of solid tumor cancers contain significant regions of hypoxia, which provide unique challenges for targeting by potent anticancer agents. Bioreductively activatable prodrug conjugates (BAPCs) represent a promising strategy for therapeutic intervention. BAPCs are designed to be biologically inert until they come into contact with low oxygen tension, at which point reductase enzyme mediated cleavage releases the parent anticancer agent in a tumor-specific manner.

Chemosensitizing AML cells by targeting bone marrow endothelial cells.

Refractory disease is the greatest challenge in treating patients with acute myeloid leukemia (AML). Blood vessels may serve as sanctuary sites for AML. When AML cells were co-cultured with bone marrow endothelial cells (BMECs), a greater proportion of leukemia cells were in G0/G1.

Design, synthesis, and biological evaluation of water-soluble amino acid prodrug conjugates derived from combretastatin, dihydronaphthalene, and benzosuberene-based parent vascular disrupting agents.

Targeting tumor vasculature represents an intriguing therapeutic strategy in the treatment of cancer. In an effort to discover new vascular disrupting agents with improved water solubility and potentially greater bioavailability, various amino acid prodrug conjugates (AAPCs) of potent amino combretastatin, amino dihydronaphthalene, and amino benzosuberene analogs were synthesized along with their corresponding water-soluble hydrochloride salts. These compounds were evaluated for their ability to inhibit tubulin polymerization and for their cytotoxicity against selected human cancer cell lines.

Structural interrogation of benzosuberene-based inhibitors of tubulin polymerization.

The discovery of 3-methoxy-9-(30,40,50-trimethoxyphenyl)-6,7-dihydro-5H-benzo[7]annulen-4-ol (a benzosuberene-based analogue referred to as KGP18) was originally inspired by the natural products colchicine and combretastatin A-4 (CA4). The relative structural simplicity and ease of synthesis of KGP18, coupled with its potent biological activity as an inhibitor of tubulin polymerization and its cytotoxicity (in vitro) against human cancer cell lines, has resulted in studies focused on new analogue design and synthesis. Our goal was to probe the relationship of structure to function in this class of anticancer agents.

Synthesis and biochemical evaluation of benzoylbenzophenone thiosemicarbazone analogues as potent and selective inhibitors of cathepsin L.

Upregulation of cathepsin L in a variety of tumors and its ability to promote cancer cell invasion and migration through degradation of the extracellular matrix suggest that cathepsin L is a promising biological target for the development of anti-metastatic agents. Based on encouraging results from studies on benzophenone thiosemicarbazone cathepsin inhibitors, a series of fourteen benzoylbenzophenone thiosemicarbazone analogues were designed, synthesized, and evaluated for their inhibitory activity against cathepsins L and B. Thiosemicarbazone inhibitors 3-benzoylbenzophenone thiosemicarbazone 1, 1,3-bis(4-fluorobenzoyl)benzene thiosemicarbazone 8, and 1,3-bis(2-fluorobenzoyl)-5-bromobenzene thiosemicarbazone 32 displayed the greatest potency against cathepsin L with low IC50 values of 9.

The vascular disrupting activity of OXi8006 in endothelial cells and its phosphate prodrug OXi8007 in breast tumor xenografts.

This study describes the vascular disrupting ability and the mechanism of action of the indole-based tubulin-binding compound, OXi8006, and its water-soluble phosphate prodrug OXi8007. Treatment of rapidly proliferating human umbilical vein endothelial cells (HUVECs), used as a model for the tumor vasculature, with OXi8006 or OXi8007, caused potent microtubule disruption followed by extensive reorganization of the cytoskeletal network. The mechanism of action involved an increase in focal adhesion formation associated with an increase in phosphorylation of both non-muscle myosin light chain and focal adhesion kinase.

Exaggerated hypertensive response to combretastatin A-4 phosphate in hypertensive rats: Effective pharmacological inhibition by diltiazem.

Combretastatin A-4 phosphate (CA4P), a tubulin depolymerizing agent, shows promise in anti-cancer therapy and is associated with dose-dependent transient hypertension. The cardiac consequence of this hypertensive effect is unknown. This study was conducted to examine the cardiotoxic effect of CA4P on a rat model of hypertension.

Synthesis of structurally diverse benzosuberene analogues and their biological evaluation as anti-cancer agents.

Diversely functionalized, fused aryl-alkyl ring systems hold a prominent position as well-established molecular frameworks for a variety of anti-cancer agents. The benzosuberene (6,7 fused, also referred to as dihydro-5H-benzo[7]annulene and benzocycloheptene) ring system has emerged as a valuable molecular core component for the development of inhibitors of tubulin assembly, which function as antiproliferative anti-cancer agents and, in certain cases, as vascular disrupting agents (VDAs). Both a phenolic-based analogue (known as KGP18, compound 39) and its corresponding amine-based congener (referred to as KGP156, compound 45), which demonstrate strong inhibition of tubulin assembly (low micromolar range) and potent cytotoxicity (picomolar range for KGP18 and nanomolar range for KGP156) are noteworthy examples of such benzosuberene-based compounds.

Synthesis of a 2-aryl-3-aroyl indole salt (OXi8007) resembling combretastatin A-4 with application as a vascular disrupting agent.

The natural products colchicine and combretastatin A-4 are potent inhibitors of tubulin assembly, and they have inspired the design and synthesis of a large number of small-molecule, potential anticancer agents. The indole-based molecular scaffold is prominent among these SAR modifications, leading to a rapidly increasing number of agents. The water-soluble phosphate prodrug 33 (OXi8007) of 2-aryl-3-aroylindole-based phenol 8 (OXi8006) was prepared by chemical synthesis and found to be strongly cytotoxic against selected human cancer cell lines (GI₅₀ = 36 nM against DU-145 cells, for example).

Synthesis and biological evaluation of indole-based, anti-cancer agents inspired by the vascular disrupting agent 2-(3'-hydroxy-4'-methoxyphenyl)-3-(3″,4″,5″-trimethoxybenzoyl)-6-methoxyindole (OXi8006).

The discovery of a 2-aryl-3-aroyl indole-based small-molecule inhibitor of tubulin assembly (referred to as OXi8006) inspired the design, synthesis, and biological evaluation of a series of diversely functionalized analogues. In the majority of examples, the pendant 2-aryl ring contained a 3-hydroxy-4-methoxy substitution pattern, and the fused aryl ring featured a 6-methoxy group. Most of the variability was in the 3-aroyl moiety, which was modified to incorporate methoxy (33-36), nitro (25-27), halogen (28-29), trifluoromethyl (30), or trifluoromethoxy (31-32) functionalities.

An Amino-Benzosuberene Analogue That Inhibits Tubulin Assembly and Demonstrates Remarkable Cytotoxicity.

The recent discovery of a small-molecule benzosuberene-based phenol that demonstrates remarkable picomolar cytotoxicity against selected human cancer cell lines and strongly inhibits tubulin polymerization (1-2 µM) inspired the design and synthesis of a variety of new, structurally diverse benzosuberene derivatives. An efficient synthetic route to functionalized benzosuberenes was developed. This methodology utilized a Wittig reaction, followed by a selective alkene reduction and ring-closing cyclization to form the core benzosuberone structure.

Small-molecule inhibitors of cathepsin L incorporating functionalized ring-fused molecular frameworks.

Cathepsin L is a cysteine protease that is upregulated in a variety of malignant tumors and plays a significant role in cancer cell invasion and migration. It is an attractive target for the development of small-molecule inhibitors, which may prove beneficial as treatment agents to limit or arrest cancer metastasis. We have previously identified a structurally diverse series of thiosemicarbazone-based inhibitors that incorporate the benzophenone and thiochromanone molecular scaffolds.

Initial evaluation of the antitumour activity of KGP94, a functionalized benzophenone thiosemicarbazone inhibitor of cathepsin L.

Kinetic analysis of the mode of inhibition of cathepsin L by KGP94, a lead compound from a privileged library of functionalized benzophenone thiosemicarbazone derivatives, demonstrated that it is a time-dependent, reversible, and competitive inhibitor of the enzyme. These results are consistent with the formation of a transient covalent bond, and are supported by molecular modeling that places the thiocarbonyl of the inhibitor in proximity to the thiolate moiety of the enzyme active site Cys25. KGP94 significantly decreased the activity of cathepsin L toward human type I collagen, and impeded both migration and invasion of MDA-MB-231 human breast cancer cells.

Synthesis and biochemical evaluation of thiochromanone thiosemicarbazone analogues as inhibitors of cathepsin L.

A series of 36 thiosemicarbazone analogues containing the thiochromanone molecular scaffold functionalized primarily at the C-6 position were prepared by chemical synthesis and evaluated as inhibitors of cathepsins L and B. The most promising inhibitors from this group are selective for cathepsin L and demonstrate IC50 values in the low nanomolar range. In nearly all cases, the thiochromanone sulfide analogues show superior inhibition of cathepsin L as compared to their corresponding thiochromanone sulfone derivatives.

Regioselective synthesis of water-soluble monophosphate derivatives of combretastatin A-1.

The natural products combretastatin A-4 (CA4) and combretastatin A-1 (CA1) are potent cancer vascular disrupting agents and inhibitors of tubulin assembly (IC₅₀ = 1-2 μM). The phosphorylated prodrugs CA4P and CA1P are undergoing human clinical trials against cancer. CA1 is unique due to its incorporation of a vicinal phenol, which has afforded the opportunity to prepare both diphosphate and regioisomeric monophosphate derivatives.

Functionalized benzophenone, thiophene, pyridine, and fluorene thiosemicarbazone derivatives as inhibitors of cathepsin L.

A series of thiosemicarbazone analogs based on the benzophenone, thiophene, pyridine, and fluorene molecular frameworks has been prepared by chemical synthesis and evaluated as small-molecule inhibitors of the cysteine proteases cathepsin L and cathepsin B. The two most potent inhibitors of cathepsin L in this series (IC(50)<135 nM) are brominated-benzophenone thiosemicarbazone analogs that are further functionalized with a phenolic moiety (2 and 6). In addition, a bromo-benzophenone thiosemicarbazone acetyl derivative (3) is also strongly inhibitory against cathepsin L (IC(50)=150.

Regio- and stereospecific synthesis of mono-beta-d-glucuronic acid derivatives of combretastatin A-1.

Synthetic routes have been established for the preparation of regio- and stereoisomerically pure samples of the mono-beta-d-glucuronic acid derivatives of combretastatin A-1, referred to as CA1G1 (5a) and CA1G2 (6a). Judicious choice of protecting groups for the catechol ring was required for the regiospecific introduction of the glucuronic acid moiety. The tosyl group proved advantageous in this regard.

Design, synthesis, and biological evaluation of potent thiosemicarbazone based cathepsin L inhibitors.

A small library of 36 functionalized benzophenone thiosemicarbazone analogs has been prepared by chemical synthesis and evaluated for their ability to inhibit the cysteine proteases cathepsin L and cathepsin B. Inhibitors of cathepsins L and B have the potential to limit or arrest cancer metastasis. The six most active inhibitors of cathepsin L (IC50<85 nM) in this series incorporate a meta-bromo substituent in one aryl ring along with a variety of functional groups in the second aryl ring.