The stabilization of DNA triplexes using the novel base analogue 2-aminopyridinone (APyO)

Triplex-forming oligonucleotides (TFOs) could position themselves at the forefront of personalised medicine in their potential to interact with defined target gene sequences in humans and in other organisms. They could alter gene expression thus providing potential new agents to combat diseases that are recalcitrant to current therapies or where no efficacious medicine currently exists. Target selection has been greatly expanded through the sequencing of the complete human genome and that of many other organisms. A practical use that we have identified is to silence beta lactamase in drug-resistant, pathogenic bacteria such as MRSA.

Application of TFOs as therapeutic agents is hampered by the pH sensitivity when pyrimidine-containing TFOs form Hoogsteen hydrogen bonds with DNA targets. We propose chemical synthesis and evaluation of a new, pH-independent base analogue for incorporation into TFOs, and evaluation of the stability and sequence selectivity.

The project will involve the chemical synthesis and evaluation of the protected, activated derivative of aminopyridinone 2’-deoxynucleoside for use in automated solid-supported synthesis of TFOs targeted to the beta lactamase gene in MRSA. Various TFOs have been prepared and studied in attempts to overcome the poor hybrid stability of triplex structures formed between TFO and duplex DNA target. Recognition of duplex DNA is achieved through Hoogsteen hydrogen bond formation between the third base and duplex target in the C⁺*G.C triplex (Figure 1). Although stable under acidic conditions, the requirement for protonation of cytosine results in triplex destabilisation of TFOs at neutral pH. To circumvent the need for acidic pH to facilitate protonation at nitrogen, we have identified APyO as having the capability to form potentially pH-independent triplexes when used in place of C⁺ (Figure 1). We will modify the known procedure used for preparation of the APyO ribonucleoside (2) by using alternative 2’-deoxyribose derivatives as starting materials, to form the corresponding 2’-deoxyribonucleoside (1) in suitably protected form as the phosphoramidite for oligo synthesis. We have optimised a multi-gram scale synthesis of the Hoffer sugar starting material from 2’-deoxyribose without need for chromatographic separation of intermediates or the use of gaseous HCl. Our publication track record demonstrates proficiencies in nucleoside chemistry, development of solid phase synthesis methods, evaluation and patent protection of active, antimicrobial compounds.

Estimated yearly cost of consumables

£4,000 per year

Person Specification

A Masters degree in a relevant subject with a 60% or higher weighted average, and/or a First or Upper Second Class Honours degree (or an equivalent qualification from an overseas institution)

Submitting an application

As part of the application, you will need to supply:

·        A copy of your current CV

·        Copies of your academic qualifications for your Bachelor degree, and Masters degree (if studied); this should include both certificates and transcripts, and must be translated in to English

·        A research proposal statement*

·        Two academic references

·        Proof of your English Language proficiency

Details of how to submit your application can be found here. 

*The application must be accompanied by a “research proposal” statement. An original proposal is not required as the initial scope of the project has been defined, candidates should take this opportunity to detail how their knowledge and experience will benefit the project and should also be accompanied by a brief review of relevant research literature.

Please include the supervisor’s name and project title in your Personal Statement.

If you require further information about the application process please contact the Postgraduate Admissions team at [Email Address Removed]