De novo engineering of a human cystathionine-γ-lyase for systemic l-methionine depletion cancer therapy Journal Article
| Authors: | Stone, E.; Paley, O.; Hu, J.; Ekerdt, B.; Cheung, N. K.; Georgiou, G. |
| Article Title: | De novo engineering of a human cystathionine-γ-lyase for systemic l-methionine depletion cancer therapy |
| Abstract: | It has been known for nearly a half century that human tumors, including those derived from the nervous system such as glioblastomas, medulloblastoma, and neuroblastomas are much more sensitive than normal tissues to l-methionine (l-Met) starvation. More recently, systemic l-Met depletion by administration of Pseudomonas putida methionine-γ-lyase (MGL) could effectively inhibit human tumors xenografted in mice. However, bacterial-derived MGLs are unstable in serum (t 1/2 = 1.9 ± 0.2 h) and highly immunogenic in primates. Since the human genome does not encode a human MGL enzyme, we created de novo a methionine degrading enzyme by reengineering the structurally homologous pyridoxal phosphate-dependent human enzyme cystathionine-γ- lyase (hCGL). hCGL degrades l-cystathionine but displays no promiscuous activity toward l-Met. Rational design and scanning saturation mutagenesis led to the generation of a variant containing three amino acid substitutions (hCGL-NLV) that degraded l-Met with a k cat/K M of 5.6 × 10 2 M -1 s -1 and displayed a serum deactivation t 1/2 = 78 ± 5 h (non-PEGylated). In vitro, the cytotoxicity of hCGL-NLV toward 14 neuroblastoma cell lines was essentially indistinguishable from that of the P. putida MGL. Intravenous administration of PEGylated hCGL-NLV in mice reduced serum l-Met from 123 μM to <5 μM for over 30 h. Importantly, treatment of neuroblastoma mouse xenografts with PEGylated hCGL-NLV resulted in near complete cessation of tumor growth. Since the mode of action of hCGL-NLV does not require breaching the blood-brain barrier, this enzyme may have potential application for sensitive tumors that arise from or metastasize to the central nervous system. © 2012 American Chemical Society. |
| Journal Title: | ACS Chemical Biology |
| Volume: | 7 |
| Issue: | 11 |
| ISSN: | 1554-8929 |
| Publisher: | American Chemical Society |
| Date Published: | 2012-11-16 |
| Start Page: | 1822 |
| End Page: | 1829 |
| Language: | English |
| DOI: | 10.1021/cb300335j |
| PROVIDER: | scopus |
| PMCID: | PMC3500410 |
| PUBMED: | 22963240 |
| DOI/URL: | |
| Notes: | --- - "Export Date: 3 December 2012" - "CODEN: ACBCC" - "Source: Scopus" |
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