A Thapsigargin Prodrug Produces Sustained Growth Inhibition and Substantial Regression of Human Breast Cancers In Vivo with Minimal Host Toxicity.
Authors: Denmeade S. , Rosen D. , Christensen S. , Dionne C. , Isaacs J. 
Published: December 2010

Abstract
Background: Thapsigargin (TG) is a cytotoxic natural product isolated in high yields from the plant Thapsia garganica. In the NCI 60 Cancer Cell Line screen, TG has a GI50 of ∼10-10 M which compares favorably with agents such as paclitaxel (10-8 M) and doxorubicin (10-7 M) in this assay. TG is a non-cell type specific cytotoxin that kills cells in a proliferation independent manner via its potent inhibition of a critical intracellular protein, the Sarcoplasmic/Endoplasmic Reticulum Calcium ATPase (SERCA) pump. Sustained inhibition of the SERCA pump by TG produces an elevation of intracellular calcium to micromolar levels which induces apoptosis via activation of the ER stress response and release of apoptotic factors from the mitochondria. TG has no therapeutic index in vivo with an LD100 of 0.2 mg/kg.Methods: In order to create a theraputic index that would allow for sytemic therapy of human breast cancer we generated prodrugs by coupling water soluble acidic amino acid containing peptides to a potent analog of thapsigargin, (12Aminododecanoyl)-8-O debutanoylthapsigargin (12ADT). In addition to solubilization, the peptide also serves to mask the cytotoxicity of the TG analog until it is released by specific active proteases present within tumor sites. One of these prodrugs (G202) was selected for further in vivo evaluation based on its solubility and ability to be hydrolyzed by the carboxypeptidase Prostate-Specific Membrane Antigen (PSMA). PSMA is membrane protein that is highly expressed by normal and malignant prostate epithelial cells. PSMA, however, is also expressed by the neovasculature within most solid tumors, but not by normal tissue vasculature. In this study we evaluated the toxicity and efficacy of this PSMA-activated thapsigargin prodrug against human MCF-7 breast cancer xenografts.Results: In preliminary studies with prostate cancer xenografts we determined that a single intravenous dose of 56 mg/kg G202 formulated in propylene glycol/Solutol could be administered safely without acute toxicity. Mice bearing MCF-7 xenografts were thus treated with one, two or three doses of 56 mg/kg G202. Three doses produced significant tumor regressions and prolonged growth inhibition but was associated with toxicity whereas a single dose of G202 had no discernible toxicity but produced only transient growth inhibition. In contrast, two consecutive doses of G202 produced only transient weight loss and resulted in significant tumor regression and sustained growth inhibition, with complete responses observed in a subset of animals. High levels (∼5 μM) of the active drug 12ADT-Asp were measurable in the tumor tissue 96 hrs post injection while negligible hydrolysis of G202 was observed in plasma. Discussion: The effectiveness of G202 against a panel of ER positive and negative breast cancer xenografts is currently being evaluated. On the basis of this encouraging preliminary data continued clinical development of G202 as therapy for metastatic breast cancer appears warranted.
Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5074.