Specific targeting of cytosine deaminase to solid tumors by engineered Clostridium acetobutylicum

Abstract

The presence of severe hypoxia and necrosis in solid tumors offers the potential to apply an anaerobic bacterial enzyme/prodrug approach in cancer treatment. In this context the apathogenic C. acetobutylicum was genetically engineered to express and secrete E. coli cytosine deaminase (CDase). Considerable levels of functional cytosine deaminase were detected in lysates and supernatants of recombinant C acetobutylicum cultures. After administration of the recombinant Clostridium to rhabdomyosarcoma bearing rats used as a model, cytosine deaminase could be detected at the tumor site. Moreover, following administration of the vascular targeting agent combretastatin A-4 phosphate significantly increased levels of cytosine deaminase were detected at the tumor site as a consequence of enlarged tumor necrosis and subsequently improved growth of C. acetobutylicum. The results provide evidence for the potential application of Clostrisdium-based therapeutic protein transfer to tumors in anticancer therapy.

Repeated cycles of Clostridium-directed enzyme prodrug therapy result in sustained antitumour effects in vivo

“…To evaluate the in vivo efficacy of this novel enzyme, transfer of the NTR-H enzyme to a Clostridium strain with enhanced tumour colonisation properties was absolutely necessary. Indeed, although previous CDEPT work undoubtedly established the safety and feasibility of the approach, proof of in vivo antitumour efficacy has so far been limited (Lemmon et al, 1997;Theys et al, 2001a;Liu et al, 2002). The major reason for this failure has been attributed to the low tumour colonisation efficiency of the employed strains.…”

Potential therapeutic applications of recombinant, invasive E. coli

“…34 Other types of bacteria are currently being developed as anticancer agents. [6][7][8][9][10]35 In these strategies, spores of anaerobic bacteria are injected and selectively grow within the necrotic regions of tumours where they find a niche to grow. For example, C. novyi has shown some dramatic antitumour effects in combination with conventional chemotherapeutic drugs 35 and Salmonella are capable of targeting, colonizing and eliciting growth suppression of tumours in mice.…”

“…More recently, this concept has been re-evaluated using attenuated strains of Salmonella 6,7 and Clostridium. [8][9][10] A genetically attenuated strain of Salmonella (VNP20009) 7 was recently used in a clinical trial. 11 VNP20009 demonstrated marked toxicity when injected above 3 Â 10 8 CFU/m 2 .…”

Microenvironmental regulation of biomacromolecular therapies

“…Tremendous efforts have been made to modify the therapeutic biomacromolecules with targeting and functional cues, and to design new delivery systems, leading to significant enhancement of therapeutic efficiency. For example, therapeutic proteins and nucleotides have been hybridized through chemical fusion or complexation with diverse targeting 17,18 and delivery molecules to enhance infiltration into tissues and cells 19,20 and tissuespecific localization [21][22][23][24] . Furthermore, therapeutic biomacromolecules have been loaded into various biomaterials to enable a sustained and localized delivery manner while preserving bioactivity, as extensively reviewed elsewhere 5,[25][26][27][28][29][30] .…”