Enhancement of CPT-11 antitumor activity by adenovirus-mediated expression of β–glucuronidase in tumors

Huang, Peng; Kc, Chen; Prijovich, Zeljko M.; Cheng, Tao; Leu, Y-L; Roffler, Steve R.

doi:10.1038/cgt.2011.3

Cited by 20 publications

(28 citation statements)

References 42 publications

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“…2) and a range of enzymes from the glycosidase family, each inhibitor displayed selectivity toward E. coli b-glucuronidase. The selectivity of our inhibitors is particularly critical for their use in conjunction with CPT-11 because human b-glucuronidase expressed by tumor cells appears to play an important role in the antitumor efficacy of CPT-11 through the reactivation of SN-38G to SN-38 in the tumor microenvironment (Tobin et al, 2006;Huang et al, 2011). Of the four compounds described here, only Inhibitor 5 (K i and IC 50 values of 180 and 540 nM, respectively) displayed potency comparable with our previously-characterized inhibitors, which have a scaffold not shared by the four inhibitors outlined in this study (Wallace et al, 2010).…”

Section: Discussionmentioning

confidence: 99%

Molecular Insights into Microbialβ-Glucuronidase Inhibition to Abrogate CPT-11 Toxicity

et al. 2013

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Bacterial b-glucuronidases expressed by the symbiotic intestinal microbiota appear to play important roles in druginduced epithelial cell toxicity in the gastrointestinal (GI) tract. For the anticancer drug CPT-11 (irinotecan) and the nonsteroidal anti-inflammatory drug diclofenac, it has been shown that removal of the glucuronide moieties from drug metabolites by bacterial b-glucuronidases in the GI lumen can significantly damage the intestinal epithelium. Furthermore, selective disruption of bacterial b-glucuronidases by small molecule inhibitors alleviates these side effects, which, for CPT-11 {7-ethyl-10-[4-(1-piperidino)-1-piperidino]}, can be dose limiting. Here we characterize novel microbial b-glucuronidase inhibitors that inhibit Escherichia coli b-glucuronidase in vitro with K i values between 180 nM and 2 mM, and disrupt the enzyme in E. coli cells, with EC 50 values as low as 300 nM. All compounds are selective for E. coli b-glucuronidase without inhibiting purified mammalian b-glucuronidase, and they do not impact the survival of either bacterial or mammalian cells. The 2.8 Å resolution crystal structure of one inhibitor bound to E. coli b-glucuronidase demonstrates that it contacts and orders only a portion of the "bacterial loop" present in microbial, but not mammalian, b-glucuronidases. The most potent compound examined in this group was found to protect mice against CPT-11-induced diarrhea. Taken together, these data advance our understanding of the chemical and structural basis of selective microbial b-glucuronidase inhibition, which may improve human drug efficacy and toxicity.

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Section: Discussionmentioning

confidence: 99%

Molecular Insights into Microbialβ-Glucuronidase Inhibition to Abrogate CPT-11 Toxicity

et al. 2013

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“…[17]), attach it to the cell surface (e.g. [23,33]), or secrete from producing cells (e.g. [32,34]) offers a number of different potential applications.…”

Section: Discussionmentioning

confidence: 99%

“…Several strategies were successfully employed: e.g. fusion of cancer specific antibody-fragments with beta-glucuronidase [20] or tumor selective expression of the enzyme using bacteria [21] or adenoviruses [22,23]. The reporter gene properties of the enzyme were not studied as extensively in animals.…”

Section: Introductionmentioning

confidence: 99%

Bacterial glucuronidase as general marker for oncolytic virotherapy or other biological therapies

et al. 2011

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BackgroundOncolytic viral tumor therapy is an emerging field in the fight against cancer with rising numbers of clinical trials and the first clinically approved product (Adenovirus for the treatment of Head and Neck Cancer in China) in this field. Yet, until recently no general (bio)marker or reporter gene was described that could be used to evaluate successful tumor colonization and/or transgene expression in other biological therapies.MethodsHere, a bacterial glucuronidase (GusA) encoded by biological therapeutics (e.g. oncolytic viruses) was used as reporter system.ResultsUsing fluorogenic probes that were specifically activated by glucuronidase we could show 1) preferential activation in tumors, 2) renal excretion of the activated fluorescent compounds and 3) reproducible detection of GusA in the serum of oncolytic vaccinia virus treated, tumor bearing mice in several tumor models. Time course studies revealed that reliable differentiation between tumor bearing and healthy mice can be done as early as 9 days post injection of the virus. Regarding the sensitivity of the newly developed assay system, we could show that a single infected tumor cell could be reliably detected in this assay.ConclusionGusA therefore has the potential to be used as a general marker in the preclinical and clinical evaluation of (novel) biological therapies as well as being useful for the detection of rare cells such as circulating tumor cells.

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“…Ad-αDNS and Ad-βG were prepared as previously described. 22 Flow Cytometry. Cells were stained with a rat monoclonal antibody against murine beta-glucuronidase (7G7) 43 or a rat monoclonal antibody against HA (Roche, Mannheim, Germany) followed by a goat antibody against rat IgG (H +L) conjugated with FITC (Jackson ImmunoResearch Laboratories Inc., West Grove, PA, USA).…”

Section: ■ Materials and Methodsmentioning

confidence: 99%

“…15 Beta-glucuronidase activity in tumors can also be artificially elevated by immunoenzyme therapy 16−20 or by expressing beta-glucuronidase in cancer cells. 21,22 The design of drugs that can be selectively activated by beta-glucuronidase in the tumor microenvironment is therefore a rational and promising approach to increase cancer chemotherapy efficacy.…”

Section: ■ Introductionmentioning

confidence: 99%

Selective Cancer Therapy by Extracellular Activation of a Highly Potent Glycosidic Duocarmycin Analogue

et al. 2013

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Conventional cancer chemotherapy is limited by systemic toxicity and poor selectivity. Tumor-selective activation of glucuronide prodrugs by beta-glucuronidase in the tumor microenvironment in a monotherapeutic approach is one promising way to increase cancer selectivity. Here we examined the cellular requirement for enzymatic activation as well as the in vivo toxicity and antitumor activity of a glucuronide prodrug of a potent duocarmycin analogue that is active at low picomolar concentrations. Prodrug activation by intracellular and extracellular beta-glucuronidase was investigated by measuring prodrug 2 cytotoxicity against human cancer cell lines that displayed different endogenous levels of beta-glucuronidase, as well as against beta-glucuronidase-deficient fibroblasts and newly established beta-glucuronidase knockdown cancer lines. In all cases, glucuronide prodrug 2 was 1000-5000 times less cytotoxic than the parent duocarmycin analogue regardless of intracellular levels of beta-glucuronidase. By contrast, cancer cells that displayed tethered beta-glucuronidase on their plasma membrane were 80-fold more sensitive to glucuronide prodrug 2, demonstrating that prodrug activation depended primarily on extracellular rather than intracellular beta-glucuronidase activity. Glucuronide prodrug 2 (2.5 mg/kg) displayed greater antitumor activity and less systemic toxicity in vivo than the clinically used drug carboplatin (50 mg/kg) to mice bearing human lung cancer xenografts. Intratumoral injection of an adenoviral vector expressing membrane-tethered beta-glucuronidase dramatically enhanced the in vivo antitumor activity of prodrug 2. Our data provide evidence that increasing extracellular beta-glucuronidase activity in the tumor microenvironment can boost the therapeutic index of a highly potent glucuronide prodrug.

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Enhancement of CPT-11 antitumor activity by adenovirus-mediated expression of β–glucuronidase in tumors

Cited by 20 publications

References 42 publications

Molecular Insights into Microbialβ-Glucuronidase Inhibition to Abrogate CPT-11 Toxicity

Molecular Insights into Microbialβ-Glucuronidase Inhibition to Abrogate CPT-11 Toxicity

Bacterial glucuronidase as general marker for oncolytic virotherapy or other biological therapies

Selective Cancer Therapy by Extracellular Activation of a Highly Potent Glycosidic Duocarmycin Analogue

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