Selection of RNA-aptamers as anti-malaria agents
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Publication Number
US-9309517-B2
Publication Date
2016-04-12
Expiration Date
Abstract
The present invention relates to an aptamer or an active fragment thereof raised against the semi-conserved duffy binding ligand domain 1α, DBL1α, region of the Plasmodium falciparum erythrocyte membrane protein 1, PfEMPI, which aptamer has an effect against malaria, in particular severe cerebral malaria.
Core Innovation
The invention relates to selection and characterization of anti-malaria RNA aptamers that bind to the semi-conserved DBL1α region of PfEMP1 from Plasmodium falciparum. It identifies RNA aptamer clones selected against recombinant DBL1α, including conserved aptamer clones such as d12, b02, and e05, and sequence identifiers SEQ ID NO: 1–6 and fluorinated derivatives SEQ ID NO: 7–12.
Recombinant DBL1α (His-tag) is produced and used in an 8-round SELEX with a 2′F-modified random RNA library. Sequencing identifies the selected aptamer clones, and fluorinated aptamers are provided for enhanced blood stability. The aptamers are further characterized for association with recombinant DBL1α and binding to live infected erythrocytes, including fluorescence-based labeling.
Functionally, the selected aptamers disrupt rosetting in vitro and exhibit serum stability. The document further outlines use in treating severe cerebral malaria and in diagnosis by measuring blood sample response to the aptamers, including the use of live cell binding observations and aptamer response as a basis for the diagnostic approach.
Claims Coverage
The provided material includes two independent claims directed to specific RNA aptamers, including a non-fluorinated aptamer defined by SEQ ID NO: 6 and a fluorinated aptamer defined by SEQ ID NO: 12. The inventive features are the explicitly defined aptamer sequence content and, for the second claim, fluorination of the aptamer.
Aptamer consisting of SEQ ID NO: 6
An aptamer comprising SEQ ID NO: 6, with the dependent refinement stating that the aptamer consists of SEQ ID NO: 6.
Fluorinated aptamer consisting of SEQ ID NO: 12
A fluorinated aptamer of SEQ ID NO: 6, having the specified sequence (SEQ ID NO: 7–12), with the dependent refinement stating that the fluorinated aptamer consists of SEQ ID NO: 12.
Across the independent claims, the inventive scope is grounded in explicitly defined RNA aptamer sequences: one defined by SEQ ID NO: 6 and one defined as a fluorinated aptamer that is narrowed to SEQ ID NO: 12.
Stated Advantages
Enhanced blood stability for fluorinated aptamers.
Serum stability.
Disruption of rosetting in vitro by the selected aptamers.
Documented Applications
Treating severe cerebral malaria.
Diagnosis by measuring a blood sample response to the aptamers.
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