Preparation of Mammalian Expression Vectors Incorporating Site-Specifically Platinated-DNA Lesions
FDA-approved platinum-based anticancer drugs, cisplatin, carboplatin, and oxaliplatin, are some of the most effective chemotherapies in clinical use. The cytotoxic action of these compounds against cancer requires a combination of processes including cell entry, drug activation, DNA binding, and tra...
| Main Authors: | , , |
|---|---|
| Other Authors: | |
| Format: | Article |
| Language: | en_US |
| Published: |
American Chemical Society (ACS)
2013
|
| Online Access: | http://hdl.handle.net/1721.1/82151 https://orcid.org/0000-0002-2693-4982 |
| _version_ | 1811087159886610432 |
|---|---|
| author | Ang, Wee Han Brown, William Wesley Lippard, Stephen J. |
| author2 | Massachusetts Institute of Technology. Department of Chemistry |
| author_facet | Massachusetts Institute of Technology. Department of Chemistry Ang, Wee Han Brown, William Wesley Lippard, Stephen J. |
| author_sort | Ang, Wee Han |
| collection | MIT |
| description | FDA-approved platinum-based anticancer drugs, cisplatin, carboplatin, and oxaliplatin, are some of the most effective chemotherapies in clinical use. The cytotoxic action of these compounds against cancer requires a combination of processes including cell entry, drug activation, DNA binding, and transcription inhibition resulting in apoptotic cell death. The drugs form Pt lesions with nuclear DNA, leading to the arrest of key cellular functions and triggering a variety of cellular responses. DNA probes containing Pt−DNA conjugates are important tools for studying the molecular mechanisms of these processes. In order to facilitate investigation of specific Pt−DNA lesion processing within live cells, we devised a strategy for constructing plasmids containing a single site-specific Pt−DNA adduct. The method involves the use of nicking restriction enzymes to create closely spaced tandem gaps on the plasmid followed by removal of the intervening doubly nicked DNA strand to form a short single-stranded gap. Synthetic platinated oligonucleotides were incorporated into the gapped plasmid construct to generate a covalently closed circular platinated plasmid in good yield. We discuss the application of this methodology to prepare plasmids containing a platinum 1,2-d(G*pG*) or 1,3-d(G*pTpG*) intrastrand cross-link, two notable adducts formed by the three clinically approved drugs. |
| first_indexed | 2024-09-23T13:41:00Z |
| format | Article |
| id | mit-1721.1/82151 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T13:41:00Z |
| publishDate | 2013 |
| publisher | American Chemical Society (ACS) |
| record_format | dspace |
| spelling | mit-1721.1/821512022-09-28T15:29:23Z Preparation of Mammalian Expression Vectors Incorporating Site-Specifically Platinated-DNA Lesions Ang, Wee Han Brown, William Wesley Lippard, Stephen J. Massachusetts Institute of Technology. Department of Chemistry Ang, Wee Han Brown, William Wesley Lippard, Stephen J. FDA-approved platinum-based anticancer drugs, cisplatin, carboplatin, and oxaliplatin, are some of the most effective chemotherapies in clinical use. The cytotoxic action of these compounds against cancer requires a combination of processes including cell entry, drug activation, DNA binding, and transcription inhibition resulting in apoptotic cell death. The drugs form Pt lesions with nuclear DNA, leading to the arrest of key cellular functions and triggering a variety of cellular responses. DNA probes containing Pt−DNA conjugates are important tools for studying the molecular mechanisms of these processes. In order to facilitate investigation of specific Pt−DNA lesion processing within live cells, we devised a strategy for constructing plasmids containing a single site-specific Pt−DNA adduct. The method involves the use of nicking restriction enzymes to create closely spaced tandem gaps on the plasmid followed by removal of the intervening doubly nicked DNA strand to form a short single-stranded gap. Synthetic platinated oligonucleotides were incorporated into the gapped plasmid construct to generate a covalently closed circular platinated plasmid in good yield. We discuss the application of this methodology to prepare plasmids containing a platinum 1,2-d(G*pG*) or 1,3-d(G*pTpG*) intrastrand cross-link, two notable adducts formed by the three clinically approved drugs. National Cancer Institute (U.S.) (CA34992) 2013-11-18T13:34:42Z 2013-11-18T13:34:42Z 2009-04 2009-02 Article http://purl.org/eprint/type/JournalArticle 1043-1802 1520-4812 http://hdl.handle.net/1721.1/82151 Ang, Wee Han, William Wesley Brown, and Stephen J. Lippard. “Preparation of Mammalian Expression Vectors Incorporating Site-Specifically Platinated-DNA Lesions.” Bioconjugate Chemistry 20, no. 5 (May 20, 2009): 1058-1063. https://orcid.org/0000-0002-2693-4982 en_US http://dx.doi.org/10.1021/bc900031a Bioconjugate Chemistry Article is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use. application/pdf American Chemical Society (ACS) PMC |
| spellingShingle | Ang, Wee Han Brown, William Wesley Lippard, Stephen J. Preparation of Mammalian Expression Vectors Incorporating Site-Specifically Platinated-DNA Lesions |
| title | Preparation of Mammalian Expression Vectors Incorporating Site-Specifically Platinated-DNA Lesions |
| title_full | Preparation of Mammalian Expression Vectors Incorporating Site-Specifically Platinated-DNA Lesions |
| title_fullStr | Preparation of Mammalian Expression Vectors Incorporating Site-Specifically Platinated-DNA Lesions |
| title_full_unstemmed | Preparation of Mammalian Expression Vectors Incorporating Site-Specifically Platinated-DNA Lesions |
| title_short | Preparation of Mammalian Expression Vectors Incorporating Site-Specifically Platinated-DNA Lesions |
| title_sort | preparation of mammalian expression vectors incorporating site specifically platinated dna lesions |
| url | http://hdl.handle.net/1721.1/82151 https://orcid.org/0000-0002-2693-4982 |
| work_keys_str_mv | AT angweehan preparationofmammalianexpressionvectorsincorporatingsitespecificallyplatinateddnalesions AT brownwilliamwesley preparationofmammalianexpressionvectorsincorporatingsitespecificallyplatinateddnalesions AT lippardstephenj preparationofmammalianexpressionvectorsincorporatingsitespecificallyplatinateddnalesions |