Difference between revisions of "Part:BBa K2688003"
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<partinfo>BBa_K2688003 short</partinfo>
 
<partinfo>BBa_K2688003 short</partinfo>
  
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'''Executive summary''' : This BioBrick is a full translation unit for cpg2, an enzyme that degrades the anticancer drug methotrexate. When expressed in E. coli, it can rapidly eliminate methotrexate from the culture medium. This is proven conclusively by HPLC, and a bioassay that measures residual toxicity.
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'''Executive summary''' : This BioBrick is a full translation unit encoding CPG2, an enzyme that converts the anticancer drug methotrexate (MTX) into less toxic compounds. When expressed in E. coli, we have shown that it can rapidly remove MTX from the culture medium. This is proven conclusively by HPLC, and a bioassay that measures residual toxicity.
  
 
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The carboxypeptidase G2 is an hydrolase that cleaves the (poly)glutamate tail off folates and analogues, leaving behind a pteroate ring system<ref name="Roswell1997" />. There is broad substrate specificity, including endogenous folates, and notably the chemotherapeutic drug methotrexate (MTX). Its metabolite DAMPA has little antifolate activity in vitro, and none of the relevant clinical effects of the parent drug<ref name="Widemann2000" />.
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The Pseudomonas carboxypeptidase G2 (CPG2 or ‘glucarpidase’) is an hydrolase that cleaves the (poly)glutamate tail off folates and analogs, leaving behind a pteroate ring system<ref name="Roswell1997" />). There is broad substrate specificity, including endogenous folates, and notably the chemotherapeutic drug MTX (MTX). Its metabolite DAMPA (2,4-diamino-N10-methylpteroic acid) (Figure 1) has little antifolate activity in vitro, and norelevant clinical effects compared to MTX<ref name="Widemann2000" />.  
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This has led to its use as both an antidote in case of MTX poisoning<ref name="Widemann2010" />, and as a general purpose platform for novel drug delivery methods, where carboxypeptidase G2 would activate polyglutamated, soluble prodrugs in situ<ref name="Masterson2006" />.
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This has led to its use as both an antidote in case of MTX poisoning<ref name="Widemann2010" />, and as a general purpose platform for novel drug delivery methods, where CPG22 would activate polyglutamated, soluble prodrugs in situ<ref name="Masterson2006" />.
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[[File:T--GO_Paris-Saclay--cpg2.png|500px|frame|center| Figure 1 : The MTX biotransformation by Pseudomonas CPG2]]
  
  
 
<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
 
<partinfo>BBa_K2688003 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K2688003 SequenceAndFeatures</partinfo>
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Revision as of 17:03, 12 October 2018

cpg2_tu

Executive summary : This BioBrick is a full translation unit encoding CPG2, an enzyme that converts the anticancer drug methotrexate (MTX) into less toxic compounds. When expressed in E. coli, we have shown that it can rapidly remove MTX from the culture medium. This is proven conclusively by HPLC, and a bioassay that measures residual toxicity.

The Pseudomonas carboxypeptidase G2 (CPG2 or ‘glucarpidase’) is an hydrolase that cleaves the (poly)glutamate tail off folates and analogs, leaving behind a pteroate ring system[1]). There is broad substrate specificity, including endogenous folates, and notably the chemotherapeutic drug MTX (MTX). Its metabolite DAMPA (2,4-diamino-N10-methylpteroic acid) (Figure 1) has little antifolate activity in vitro, and norelevant clinical effects compared to MTX[2].

This has led to its use as both an antidote in case of MTX poisoning[3], and as a general purpose platform for novel drug delivery methods, where CPG22 would activate polyglutamated, soluble prodrugs in situ[4].

Figure 1 : The MTX biotransformation by Pseudomonas CPG2


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 132
    Illegal NgoMIV site found at 820
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 919




  1. ↑ Rowsell, S., Pauptit, R. A., Tucker, A. D., Melton, R. G., Blow, D. M., & Brick, P. (1997). Crystal structure of carboxypeptidase G2, a bacterial enzyme with applications in cancer therapy. Structure (London, England : 1993), 5(3), 337–347. https://doi.org/10.1016/S0969-2126(97)00191-3
  2. ↑ Widemann, B. C., Sung, E., Anderson, L., Salzer, W. L., Balis, F. M., Monitjo, K. S., … Adamson, P. C. (2000). Pharmacokinetics and metabolism of the methotrexate metabolite 2, 4-diamino-N(10)-methylpteroic acid. The Journal of Pharmacology and Experimental Therapeutics, 294(3), 894–901.
  3. ↑ Widemann, B. C., Balis, F. M., Kim, A. R., Boron, M., Jayaprakash, N., Shalabi, A., … Adamson, P. C. (2010). Glucarpidase, leucovorin, and thymidine for high-dose methotrexate-induced renal dysfunction: Clinical and pharmacologic factors affecting outcome. Journal of Clinical Oncology, 28(25), 3979–3986. https://doi.org/10.1200/JCO.2009.25.4540
  4. ↑ Masterson, L. A., Spanswick, V. J., Hartley, J. A., Begent, R. H., Howard, P. W., & Thurston, D. E. (2006). Synthesis and biological evaluation of novel pyrrolo[2,1-c][1,4]benzodiazepine prodrugs for use in antibody-directed enzyme prodrug therapy. Bioorganic & Medicinal Chemistry Letters, 16(2), 252–256. https://doi.org/10.1016/J.BMCL.2005.10.017