AS1411, a Novel DNA Aptamer as a Potential Treatment of Acute Myelogenous Leukaemia (AML).
Authors: Shah Kirti , Djeha Hakim , Richie Christy , McGeever Grainne , Green Colin 
Published: October 2019

AS1411 (formerly AGRO100) is the first nucleic acid aptamer to enter oncology trials and is currently in a Phase I study in renal and lung patients. AS1411 is a 26mer unmodified phosphodiester-based guanosine rich oligonucleotide which, as a dimer, forms a serum-resistant quartet structure. It is postulated to exert anti-cancer activity by binding to cell surface nucleolin resulting in S-phase cell cycle arrest. We have previously shown that AS1411 inhibits growth of a wide range of solid tumours, including lung, renal, prostate and colorectal with EC50s of 2 to 8 μM. In this series of studies, we investigated the potential of AS1411 in the treatment of acute myelogenous leukaemia (AML). Expression of cell-surface and intracellular expression of nucleolin was assessed by indirect immunocytochemical staining. Four AML cell lines (MV4-11, KG1, HL60 and K562) and two solid tumour cell lines (A549 - lung; DU145 - prostate) were used. Permeabilised or non-permeabilised cells were either stained in situ on chamber glass slides or as cytospin preparations. Three anti-nucleolin monoclonal antibodies and the Vector immunoperoxidase system were used. Cells were qualitatively assessed for extent and intensity of staining. In the KG1, HL60 and MV4-11 cell lines, intense, homogeneous staining was seen in 100% of non-permeabilised cells, indicating high levels of surface-expressed nucleolin; in K562 cell lines, very intense, but heterogeneous surface staining was seen. In the A549 lung and DU145 prostate lines, moderate surface staining for nucleolin was seen. In the permeabilised AML and solid tumour cells, all lines showed similar levels of homogeneous, intense intracellular staining, as expected for an ubiquitous intracellular and intranuclear protein. In SRB in vitro cytotoxicity assays, AS1411 resulted in cell death with an EC50 of 0.6 to 5 μM; the MV4-11 line was most sensitive; the EC50 for solid tumour cell lines ranged between 2 and 8 μM. Since exposure of cells to AS1411 results in S-phase cell cycle arrest; we postulated that combination of AS1411 with S-phase dependent chemotherapeutic agents may result in increased cytotoxicity and synergistic activity. An early study of the combination of AS1411 with cytosine arabinoside (AraC) indicates that there is a synergistic effect between the two drugs; the data also suggest that the synergy may be schedule dependent and requires prior exposure to AS1411 before treatment with AraC. The distribution of the aptamer in the mouse was examined using tritiated-AS1411. A total dose of 158 kBq of 3H-AS1411 was given by intravenous injection to mice bearing solid human tumour xenografts and samples of tumours and normal tissue taken between 1 hour and 4 days post injection. Approximately 1.6 to 3.1% of the injected dose was measured in the bone and bone marrow 24 hours after dosing. The ratio of the amount of 3H-AS1411 in the bone marrow to that in the blood ranged between 1.3 and 3.7 throughout the 4-day sampling period. In conclusion, we have demonstrated that nucleolin appears to be intensely expressed on the surface of AML cell lines. AML cell lines are killed by treatment with AS1411 and synergistic cytotoxicity is seen when the AS1411 is combined with AraC. In addition, the bone marrow compartment is accessible to AS1411. Taken together these findings support investigating the anti-cancer effects of AS1411 in AML patients either alone or in combination with standard AraC therapy.