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In vitro selection of DNA aptamers against human osteosarcoma

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Summary

Background. Osteosarcoma is a highly malignant bone tumor, most frequently occurring in the rapid bone growth phase. Effective treatment of this disease is hindered by the lack of specific probes for early diagnosis and the fast cancer widespread. Methods. To find such probes, the cell-Systematic Evolution of Ligands by EXponential enrichment (cell-SELEX) methodology was implemented against the human osteosarcoma MG-63 cell line towards the selection of new specific aptamers. After 10 rounds of selection, the aptamer DNA pool was Sanger sequenced and the sequences were subjected to a bioinformatic analysis that included sequence alignment, phylogenetic relationship, and secondary structure prediction. Results. A DNA aptamer (OS-7.9), with a dissociation constant (Kd) value in the nanomolar range (12.8 ± 0.9 nM), revealed high affinity against the target cells at the physiological temperature. Furthermore, the selected aptamer also recognized lung carcinoma and colon colorectal adenocarcinoma cell lines, which are reported as common metastasis sites of osteosarcoma. Conclusions. These results suggest that OS-7.9 could recognize a common protein expressed in these cancer cells, possibly becoming a potential molecular probe for early diagnosis and targeted therapies for metastatic disease. Moreover, to the best of our knowledge, this was the first attempt to generate a DNA aptamer (OS-7.9 aptamer) against the MG-63-cell line by cell-SELEX.

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Abbreviations

CAR-T:

Chimeric antigen receptor T

BSA:

Bovine serum albumine

DAPI:

4′,6-Diamidino-2-phenylindole

DMEM:

Dulbecco’s Modified Eagle Medium

DG2:

Disialoganglioside

FBS:

Fetal Bovine Serum

\({{{K}}}_{{{d}}}\) :

Dissociation constant

mAb:

Monoclonal antibody

OS:

Osteosarcoma

PBS:

Phosphate-buffered saline

PFA:

Paraformaldehyde

RNA:

Ribonucleic acid

SD:

Standard deviation

SELEX:

Systematic Evolution of Ligands by EXponential Enrichment

SL:

Stem-loop

ssDNA:

Single-stranded deoxyribonucleic acid

VEGF:

Vascular endothelial growth factor

WB:

Washing buffer

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Funding

This work was supported by the Hacettepe University Scientific Research Projects Coordination Unit (project no. FHD-2018–17245) and by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UIDB/04469/2020unit. Diana A. Sousa (DAS) acknowledge FCT for the grant PD/BD/139083/2018. Débora Ferreira (DF) is recipient of a fellowship supported by a doctoral advanced training (call NORTE-69–2015-15) funded by the European Social Fund under the scope of Norte2020—Programa Operacional Regional do Norte.

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Author notes

  1. Khaliunsarnai Tsogtbaatar and Diana A. Sousa these authors contributed equally to this work as first authors

Authors and Affiliations

  1. Institute of Science, Hacettepe University, Bioengineering Division, 06800, Ankara, Turkey

    Khaliunsarnai Tsogtbaatar, Atakan Tevlek, Halil Murat Aydın & Eda Çelik

  2. CEB - Centre of Biological Engineering, University of Minho, Campus de Gualtar, 4710-057, Braga, Portugal

    Khaliunsarnai Tsogtbaatar, Diana A. Sousa, Debora Ferreira & Ligia Rodrigues

  3. Centre for Bioengineering, Hacettepe University, 06800, Ankara, Turkey

    Halil Murat Aydın

  4. Department of Chemical Engineering, Hacettepe University, 06800, Ankara, Turkey

    Eda Çelik

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  1. Khaliunsarnai Tsogtbaatar
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Contributions

LR conceived the study. DF, AT, KT and DAS performed the experiments. KT, DAS, DF, AT, LR and ECA analyzed the data and wrote the manuscript. LR, ECA, HMA and DF provided suggestions on the experimental design. All authors reviewed the results and approved the final version of the manuscript.

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Correspondence to Ligia Rodrigues.

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Tsogtbaatar, K., Sousa, D.A., Ferreira, D. et al. In vitro selection of DNA aptamers against human osteosarcoma. Invest New Drugs 40, 172–181 (2022). https://doi.org/10.1007/s10637-021-01161-y

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