Aptamers: Cutting edge of cancer therapies
Authors: Sarah Shigdar, Brett Schrand, Paloma H. Giangrande, Vittorio de Franciscis
Journal: Molecular Therapy (2021)

The development of an aptamer-based therapeutic has rapidly progressed following the first two reports in the 1990s, underscoring the advantages of aptamer drugs associated with their unique binding properties. In 2004, the US Food and Drug Administration (FDA) approved the first therapeutic aptamer for the treatment of neovascular age-related macular degeneration, Macugen developed by NeXstar. Since then, eleven aptamers have successfully entered clinical trials for various therapeutic indications. Despite some of the pre-clinical and clinical successes of aptamers as therapeutics, no aptamer has been approved by the FDA for the treatment of cancer. This review highlights the most recent and cutting-edge approaches in the development of aptamers for the treatment of cancer types most refractory to conventional therapies. Herein, we will review (1) the development of aptamers to enhance anti-cancer immunity and as delivery tools for inducing the expression of immunogenic neoantigens; (2) the development of the most promising therapeutic aptamers designed to target the hard-to-treat cancers such as brain tumors; and (3) the development of "carrier" aptamers able to target and penetrate tumors and metastasis, delivering RNA therapeutics to the cytosol and nucleus. The development of an aptamer-based therapeutic has rapidly progressed following the first two reports in the 1990s, underscoring the advantages of aptamer drugs associated with their unique binding properties. In 2004, the US Food and Drug Administration (FDA) approved the first therapeutic aptamer for the treatment of neovascular age-related macular degeneration, Macugen developed by NeXstar. Since then, eleven aptamers have successfully entered clinical trials for various therapeutic indications. Despite some of the pre-clinical and clinical successes of aptamers as therapeutics, no aptamer has been approved by the FDA for the treatment of cancer. This review highlights the most recent and cutting-edge approaches in the development of aptamers for the treatment of cancer types most refractory to conventional therapies. Herein, we will review (1) the development of aptamers to enhance anti-cancer immunity and as delivery tools for inducing the expression of immunogenic neoantigens; (2) the development of the most promising therapeutic aptamers designed to target the hard-to-treat cancers such as brain tumors; and (3) the development of "carrier" aptamers able to target and penetrate tumors and metastasis, delivering RNA therapeutics to the cytosol and nucleus. Despite the proven antitumor efficacy of immune-checkpoint blockade by the combined use of anti-cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) and anti-programmed death-1 (PD-1) antibodies for the treatment of advanced melanoma, mesothelioma, and non-small cell lung cancer, only a small fraction of these tumors responds to treatment or progress after transient disease control. To improve immunotherapeutic effects, agonist and antagonist aptamers have been described. They have the unique advantage to be easily conjugated as bispecific and multispecific molecules targeting the tumor site and able to locally stimulate the immune response.1Pastor F. Berraondo P. Etxeberria I. Frederick J. Sahin U. Gilboa E. Melero I. An RNA toolbox for cancer immunotherapy.Nat. Rev. Drug Discov. 2018; 17: 751-767Crossref PubMed Scopus (69) Google Scholar,2Shu D. Zhang L. Bai X. Yu J. Guo P. Stoichiometry of multi-specific immune checkpoint RNA Abs for T cell activation and tumor inhibition using ultra-stable RNA nanoparticles.Mol. Ther. Nucleic Acids. 2021; 24: 426-435Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar One such proof-of-concept study demonstrated the ability to target immunomodulatory aptamers specific to targets secreted into the tumor microenvironment. Products like vascular endothelial growth factor (VEGF) and osteopontin are highly upregulated and retained in the tumor microenvironment, making them an attractive target to enhance anti-tumor immunity by allowing preferential enrichment of immunomodulatory drugs, like costimulatory 4-1BB aptamers.3Schrand B. Berezhnoy A. Brenneman R. Williams A. Levay A. Kong L.Y. Rao G. Zhou S. Heimberger A.B. Gilboa E. Targeting 4-1BB costimulation to the tumor stroma with bispecific aptamer conjugates enhances the therapeutic index of tumor immunotherapy.Cancer Immunol. Res. 2014; 2: 867-877Crossref PubMed Google Scholar Recently, Soldevilla et al.4Soldevilla M.M. Villanueva H. Meraviglia-Crivelli D. Menon A.P. Ruiz M. Cebollero J. Villalba M. Moreno B. Lozano T. Llopiz D. et al.ICOS Costimulation at the Tumor Site in Combination with CTLA-4 Blockade Therapy Elicits Strong Tumor Immunity.Mol. Ther. 2019; 27: 1878-1891Abstract Full Text Full Text PDF PubMed Scopus (11) Google Scholar demonstrated the efficacy of the combined treatment of an anti-CTLA-4 antibody with a tumor-targeted anti-inducible T cell costimulator (ICOS) aptamer in vitro and in vivo. With the multidrug resistance protein 1 (MRP1) being specifically in cancer stem cells (CSCs), the authors took advantage of a previously described MRP1-targeting aptamer to generate a MRP1-ICOS bi-specific aptamer (Figure 1A).5Soldevilla M.M. Villanueva H. Casares N. Lasarte J.J. Bendandi M. Inoges S. López-Díaz de Cerio A. Pastor F. MRP1-CD28 bi-specific oligonucleotide aptamers: target costimulation to drug-resistant melanoma cancer stem cells.Oncotarget. 2016; 7: 23182-23196Crossref PubMed Scopus (34) Google Scholar The resulting aptamer holds two binding sites: one specific for the MRP1-expressing cancer cells and another one for ICOS, which can stimulate T lymphocytes in the proximity of the tumor. These findings underscore the potentiality of using aptamers in combined therapies involving anti-CTLA-4 blockade and an ICOS agonist. In addition, strong evidence suggesting that tumor neoantigens are recognized by the immune system and that their low expression would be a possible cause for the low responsiveness of tumors to immune-checkpoint blockade therapies is emerging.6Yarchoan M. Johnson 3rd, B.A. Lutz E.R. Laheru D.A. Jaffee E.M. Targeting neoantigens to augment antitumour immunity.Nat. Rev. Cancer. 2017; 17: 569Crossref PubMed Scopus (3) Google Scholar Having previously reported the induction of neoantigens by inhibiting the nonsense-mediated messenger RNA (mRNA) decay (NMD) pathway in tumor cells,7Pastor F. Kolonias D. Giangrande P.H. Gilboa E. Induction of tumour immunity by targeted inhibition of nonsense-mediated mRNA decay.Nature. 2010; 465: 227-230Crossref PubMed Scopus (178) Google Scholar Garrido et al.8Garrido G. Schrand B. Rabasa A. Levay A. D'Eramo F. Berezhnoy A. Modi S. Gefen T. Marijt K. Doorduijn E. et al.Tumor-targeted silencing of the peptide transporter TAP induces potent antitumor immunity.Nat. Commun. 2019; 10: 3773Crossref PubMed Scopus (20) Google Scholar used an elegant and broadly applicable approach to selectively induce neoantigens in tumor cells as a promising approach to potentiate the responsiveness of patients to immunotherapies. As a targeting moiety for a small interfering (si)RNA specific for the transporter associated with antigen processing (TAP) expression, the authors took advantage of the previously characterized AS1411 aptamer.9Bates P.J. Reyes-Reyes E.M. Malik M.T. Murphy E.M. O'Toole M.G. Trent J.O. G-quadruplex oligonucleotide AS1411 as a cancer-targeting agent: Uses and mechanisms.Biochim. Biophys. Acta, Gen. Subj. 2017; 1861: 1414-1428Crossref PubMed Scopus (123) Google Scholar By folding into G-quadruplex, AS1411, originally discovered serendipitously by developing cancer-selective therapies, was shown to bind nucleolin. Nucleolin is an RNA-binding protein that is mainly localized in the nucleolus in normal cells, whereas it is found abundantly on the cell surface of most tumor cells where it can shuttle into the cytoplasm.9Bates P.J. Reyes-Reyes E.M. Malik M.T. Murphy E.M. O'Toole M.G. Trent J.O. G-quadruplex oligonucleotide AS1411 as a cancer-targeting agent: Uses and mechanisms.Biochim. 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However, TAP inhibition leads to the upregulation of alternative pathways leading to the presentation of neoantigens, named T cell epitopes associated with impaired peptide processing (TEIPP) that can elicit effective CD8+ T cell responses.12Vigneron N. Ferrari V. Van den Eynde B.J. Cresswell P. Leonhardt R.M. Cytosolic Processing Governs TAP-Independent Presentation of a Critical Melanoma Antigen.J. Immunol. 2018; 201: 1875-1888Crossref PubMed Scopus (11) Google Scholar, 13Marijt K.A. Blijleven L. Verdegaal E.M.E. Kester M.G. Kowalewski D.J. Rammensee H.G. Stevanović S. Heemskerk M.H.M. van der Burg S.H. van Hall T. Identification of non-mutated neoantigens presented by TAP-deficient tumors.J. Exp. Med. 2018; 215: 2325-2337Crossref PubMed Scopus (34) Google Scholar, 14Doorduijn E.M. Sluijter M. Marijt K.A. Querido B.J. van der Burg S.H. van Hall T. T cells specific for a TAP-independent self-peptide remain naïve in tumor-bearing mice and are fully exploitable for therapy.OncoImmunology. 2017; 7: e1382793Crossref PubMed Scopus (12) Google Scholar, 15Doorduijn E.M. Sluijter M. Querido B.J. Oliveira C.C. Achour A. Ossendorp F. van der Burg S.H. van Hall T. TAP-independent self-peptides enhance T cell recognition of immune-escaped tumors.J. Clin. Invest. 2016; 126: 784-794Crossref PubMed Scopus (38) Google Scholar The authors show that the AS1411-siTAP chimera (AsiC), named Nucl-TAP, exhibited high selectivity for tumor cells, leading to the targeted downregulation of TAP and increased antigenicity of tumor cells (Figure 1B). TAP downregulation promoted an antitumor immune response in mice, inhibiting tumor growth in the absence of measurable toxicity. Based on the antitumor response elicited in cancer cells upon TAP downregulation, Garrido and coworkers16Garrido G. Schrand B. Levay A. Rabasa A. Ferrantella A. Da Silva D.M. D'Eramo F. Marijt K.A. Zhang Z. Kwon D. et al.Vaccination against Nonmutated Neoantigens Induced in Recurrent and Future Tumors.Cancer Immunol. Res. 2020; 8: 856-868Crossref PubMed Scopus (2) Google Scholar further investigated a novel anti-tumor strategy of vaccination against TEIPP induced by TAP downregulation. To this end, the authors took advantage of a CpG oligonucleotide as the targeting moiety for a TAP-specific siRNA. Short CpG oligodeoxyribonucleotides (ODNs) have been previously shown to preferentially target and activate in vivo the Toll-like receptor 9 (TLR9)-expressing dendritic cells (DCs). Upon binding to TLR9, the CpG-TAP siRNA is taken up by the receptor-expressing antigen-presenting cells, leading to TAP downregulation and the induction of TEIPP neo-antigens. The authors concluded that vaccination against TEIPP antigens is effective at enhancing the antitumor response elicited against tumor cells treated with the Nucl-TAP aptamer conjugate to induce TEIPP.16Garrido G. Schrand B. Levay A. Rabasa A. Ferrantella A. Da Silva D.M. D'Eramo F. Marijt K.A. Zhang Z. Kwon D. et al.Vaccination against Nonmutated Neoantigens Induced in Recurrent and Future Tumors.Cancer Immunol. Res. 2020; 8: 856-868Crossref PubMed Scopus (2) Google Scholar Given the limited success of current immunotherapies against solid tumors, combinatorial and synergistic therapies might be required to fight the most difficult-to-treat tumors. A recent study by Zhang et al.17Zhang Y. Xie X. Yeganeh P.N. Lee D.-J. Valle-Garcia D. Meza-Sosa K.F. Junqueira C. Su J. Luo H.R. Hide W. Lieberman J. Immunotherapy for breast cancer using EpCAM aptamer tumor-targeted gene knockdown.Proc. Natl. Acad. Sci. USA. 2021; 118 (e2022830118)Google Scholar set out to combine different aptamer-siRNA chimeras targeting different tumor cell regulatory pathways to enhance immunotherapy. In this study, the authors used the previously described murine and human targeting epithelial cell adhesion molecule (EpCAM) aptamer, a target frequently upregulated on triple-negative breast cancer (TNBC). They conjugated an siRNA against the following: (1) Upf2, a target regulating the aforementioned NMD pathway to induce neoantigens, (2) CD47, a cell-surface marker frequently upregulated on tumor cells that acts as a "don't eat me signal" to the immune system, (3) myeloid cell leukemia 1 (MCL1), a TNBC dependency gene, or (4) poly(ADP-ribose) polymerase 1 (PARP1), a DNA damage repair gene that, when downregulated, can lead to genomic instability and initiate an innate interferon (IFN) response (Figure 1B). In this study, a cocktail of these four aptamer-siRNA chimeras inhibited tumor growth in the orthotopic murine 4T1 breast adenocarcinoma mouse model and in the genetically engineered ErbB2ΔEx16 transgenic mice. Furthermore, the authors provided mechanistic evidence that the cocktail therapy led to a significant increase in infiltrating CD8+ T cells and a more functional immune response.17Zhang Y. Xie X. Yeganeh P.N. Lee D.-J. Valle-Garcia D. Meza-Sosa K.F. Junqueira C. Su J. Luo H.R. Hide W. Lieberman J. Immunotherapy for breast cancer using EpCAM aptamer tumor-targeted gene knockdown.Proc. Natl. Acad. Sci. USA. 2021; 118 (e2022830118)Google Scholar There are a number of factors that make cancers hard to treat. These include the location of the tumor, such as those cancers arising in or spread to the brain, and tumor resistance mechanisms, such as overexpression of drug efflux pumps. Targeted drug therapies can, by the way that they typically enter cells, abrogate the issues around drug efflux pumps. Targeted therapies bind to cell-surface receptors and are then internalized via receptor-mediated endocytosis.18Yoon S. Rossi J.J. Aptamers: Uptake mechanisms and intracellular applications.Adv. Drug Deliv. Rev. 2018; 134: 22-35Crossref PubMed Scopus (45) Google Scholar If these targeted therapies carry drugs, then these drugs are internalized alongside the targeting ligand.19Zhu G. Niu G. Chen X. Aptamer-Drug Conjugates.Bioconjug. Chem. 2015; 26: 2186-2197Crossref PubMed Scopus (115) Google Scholar By far, the most well-characterized aptamer-drug conjugate is the anthracycline, doxorubicin-HCl (Dox). Dox has been used in the treatment of cancers for nearly 50 years and is still on the World Health Organization's (WHO's) list of essential medicines (WHO 2019).20Tacar O. Sriamornsak P. Dass C.R. Doxorubicin: an update on anticancer molecular action, toxicity and novel drug delivery systems.J. Pharm. Pharmacol. 2013; 65: 157-170Crossref PubMed Scopus (1322) Google Scholar It has a known mechanism of action and can be effective in some patients, although it has the typical side effects of a non-targeted therapeutic, as well as dose-limiting cardiotoxicity. Dox works by intercalating into double-stranded (ds)DNA, typically in the nucleus, following successful cellular uptake. This mechanism of action can be very simply applied to the double-stranded stem region of an aptamer as long as the correct base pairs are present (G-C).21Xiang D. Shigdar S. Bean A.G. Bruce M. Yang W. Mathesh M. Wang T. Yin W. Tran P.H.-L. Al Shamaileh H. et al.Transforming doxorubicin into a cancer stem cell killer via EpCAM aptamer-mediated delivery.Theranostics. 2017; 7: 4071-4086Crossref PubMed Scopus (33) Google Scholar,22Bagalkot V. Farokhzad O.C. Langer R. Jon S. An aptamer-doxorubicin physical conjugate as a novel targeted drug-delivery platform.Angew. Chem. Int. Ed. Engl. 2006; 45: 8149-8152Crossref PubMed Scopus (461) Google Scholar This presents a very simplified and quick-to-assemble targeted therapeutic. The complex is very stable at physiological pH, making this an ideal aptamer drug conjugate, but the complex dissociates at acidic pH, similar to the pH observed in lysosomes.21Xiang D. Shigdar S. Bean A.G. Bruce M. Yang W. Mathesh M. Wang T. Yin W. Tran P.H.-L. Al Shamaileh H. et al.Transforming doxorubicin into a cancer stem cell killer via EpCAM aptamer-mediated delivery.Theranostics. 2017; 7: 4071-4086Crossref PubMed Scopus (33) Google Scholar,23Moses J.E. Moorhouse A.D. The growing applications of click chemistry.Chem. Soc. Rev. 2007; 36: 1249-1262Crossref PubMed Scopus (1926) Google Scholar This latter property is, by far, the most important characteristic of this conjugate because if the drug is not released from the aptamer once inside the cell, then the drug remains inert and will have no cytotoxic effect. Dox is not the only drug that intercalates into dsDNA. Care must be taken to ensure that the drug chosen is able to detach from the aptamer to intercalate into the cellular DNA. Intercalation is not the only method for utilizing aptamers for drug delivery but is possibly the simplest. As chemistries have developed over time, more and more "linkers" have been developed to allow for drugs to be attached to the aptamer.23Moses J.E. Moorhouse A.D. The growing applications of click chemistry.Chem. Soc. Rev. 2007; 36: 1249-1262Crossref PubMed Scopus (1926) Google Scholar These linkers often require a simple chemical reaction to be coupled to other molecules, and with companies such as Baseclick licensing their technology, it is becoming simpler for researchers to attach cytotoxic payloads to aptamers. Indeed, with Dox intercalation having an effect on the tertiary structure of the aptamer and potentially affecting its specificity and sensitivity, the functionalization of an aptamer through click chemistry would likely have minor effects on these parameters.24Macdonald J. Denoyer D. Henri J. Jamieson A. Burvenich I.J.G. Pouliot N. Shigdar S. Bifunctional Aptamer-Doxorubicin Conjugate Crosses the Blood-Brain Barrier and Selectively Delivers Its Payload to Epithelial Cell Adhesion Molecule-Positive Tumor Cells.Nucleic Acid Ther. 2020; 30: 117-128Crossref PubMed Scopus (11) Google Scholar,25Huang Y.F. Shangguan D. Liu H. Phillips J.A. Zhang X. Chen Y. Tan W. Molecular assembly of an aptamer-drug conjugate for targeted drug delivery to tumor cells.ChemBioChem. 2009; 10: 862-868Crossref PubMed Scopus (293) Google Scholar The choice of the biomarker is also an important consideration for targeting hard-to-treat cancers. Although it is often said that there are no perfect cell-surface receptors, because there are no unique biomarkers to cancer cells, some cell-surface receptors are overexpressed on cancer cells but have a low expression profile on normal cells. It is possible that tuning the binding affinity of the aptamer to its target could prevent binding to normal cells. This was quite nicely exemplified by Münz et al.26Münz M. Murr A. Kvesic M. Rau D. Mangold S. Pflanz S. Lumsden J. Volkland J. Fagerberg J. Riethmüller G. et al.Side-by-side analysis of five clinically tested anti-EpCAM monoclonal antibodies.Cancer Cell Int. 2010; 10: 44Crossref PubMed Scopus (73) Google Scholar who performed a side-by-side analysis of EpCAM antibodies. EpCAM was one of the first cancer cell biomarkers described and consequently, was the target of the first monoclonal antibody characterized for therapeutic applications. However, clinical trials demonstrated that patients had a poor tolerability due to pancreatitis. Further investigations into the antibody confirmed that it had a picomolar binding affinity and was able to bind to cells with a fairly low expression of EpCAM such as normal pancreatic cells. Next-generation antibodies with nanomolar binding affinity did not have the same poor tolerability. Due to the unique nature of aptamers, modifying binding regions so that they will only bind to the cancer cells with a higher expression rather than to all cells that express a particular biomarker is a fairly simple process.27Shigdar S. Macdonald J. O'Connor M. Wang T. Xiang D. Al Shamaileh H. Qiao L. Wei M. Zhou S.-F. Zhu Y. et al.Aptamers as theranostic agents: modifications, serum stability and functionalisation.Sensors (Basel). 2013; 13: 13624-13637Crossref PubMed Scopus (0) Google Scholar If the sensitivity can be adjusted while maintaining the specificity through changing some nucleotides in the loop or stem region of the aptamer, then fewer side effects will be associated with the treatment, and bioavailability at the cancer cells might be increased. With these considerations in mind, how may aptamers progress to enhance survivability with treatment-retardant cancers? Although there has been a great deal of success in treating leukemias with small-molecule drugs and monoclonal antibodies, prognosis still remains poor for a number of patients. It is often thought that leukemias, being described as liquid tumors, are easier to treat than solid tumors due to a less complex environment. However, if that were the case, then the long-term survival rates would be much improved. The issues surrounding drug resistance in leukemic cells relate to the interplay among the cancerous cells, the bone marrow niche, and the extracellular matrix. This interaction leads to changes in cell signaling, which favors drug resistance and provides what has been termed a "safe haven" for the stem cells.28Cartledge Wolf D.M. Langhans S.A. Moving Myeloid Leukemia Drug Discovery Into the Third Dimension.Front Pediatr. 2019; 7: 314Crossref PubMed Scopus (5) Google Scholar An additional reason for failure with classical cytotoxic drugs is that they typically target rapidly dividing cells, and it is thought that once leukemic cells take up residence in the bone marrow niche, they become fairly quiescent. This suggests that targeting biomarkers on the cell surface for drug delivery would be highly effective. Although some leukemias have very well defined cell-surface markers, such as CD19 in B cell acute lymphoblastic leukemia (ALL), not all leukemias have such well-characterized markers,29Haubner S. Perna F. Köhnke T. Schmidt C. Berman S. Augsberger C. Schnorfeil F.M. Krupka C. Lichtenegger F.S. Liu X. et al.Coexpression profile of leukemic stem cell markers for combinatorial targeted therapy in AML.Leukemia. 2019; 33: 64-74Crossref PubMed Scopus (85) Google Scholar suggesting that a multipronged or synergistic targeting approach will be required in the future.17Zhang Y. Xie X. Yeganeh P.N. Lee D.-J. Valle-Garcia D. Meza-Sosa K.F. Junqueira C. Su J. Luo H.R. Hide W. Lieberman J. Immunotherapy for breast cancer using EpCAM aptamer tumor-targeted gene knockdown.Proc. Natl. Acad. Sci. USA. 2021; 118 (e2022830118)Google Scholar Probably the most well-characterized aptamers to leukemia cells are those developed by the Tan group30Shangguan D. Li Y. Tang Z. Cao Z.C. Chen H.W. Mallikaratchy P. Sefah K. Yang C.J. Tan W. Aptamers evolved from live cells as effective molecular probes for cancer study.Proc. Natl. Acad. Sci. USA. 2006; 103: 11838-11843Crossref PubMed Scopus (1078) Google Scholar using cell-SELEX (systematic evolution of ligands by exponential enrichment). The sgc8 aptamer was originally selected against the T cell ALL cell line, CCRF-CEM, and shown to have a binding affinity of 0.8 nM. Further testing confirmed that sgc8 could also bind to other T cell ALL cell lines, suggesting a common biomarker to these cell lines that was not present on acute myeloid leukemia (AML) cell lines. Further studies suggested that sgc8 binds to the protein tyrosine kinase 7 (PTK7) protein31Shangguan D. Cao Z. Meng L. Mallikaratchy P. Sefah K. Wang H. Li Y. Tan W. Cell-specific aptamer probes for membrane protein elucidation in cancer cells.J. Proteome Res. 2008; 7: 2133-2139Crossref PubMed Scopus (336) Google Scholar,32Xiao Z. Shangguan D. Cao Z. Fang X. Tan W. Cell-specific internalization study of an aptamer from whole cell selection.Chemistry. 2008; 14: 1769-1775Crossref PubMed Scopus (188) Google Scholar and that it could deliver daunorubicin into a T cell ALL line and demonstrate specific cytotoxicity.33Taghdisi S.M. Abnous K. Mosaffa F. Behravan J. Targeted delivery of daunorubicin to T-cell acute lymphoblastic leukemia by aptamer.J. Drug Target. 2010; 18: 277-281Crossref PubMed Scopus (0) Google Scholar Finally, this aptamer was truncated from 88 nucleotides to 41 nucleotides with no loss of binding affinity.34Shangguan D. Tang Z. Mallikaratchy P. Xiao Z. Tan W. Optimization and modifications of aptamers selected from live cancer cell lines.ChemBioChem. 2007; 8: 603-606Crossref PubMed Scopus (179) Google Scholar Dox was then intercalated into this shortened aptamer, sgc8c, which reduced the binding affinity from 0.78 nM to 2 nM. However, this did not affect cytotoxicity, with a similar effect observed between free Dox and aptamer-delivered Dox to the CCRF-CEM cell line.25Huang Y.F. Shangguan D. Liu H. Phillips J.A. Zhang X. Chen Y. Tan W. Molecular assembly of an aptamer-drug conjugate for targeted drug delivery to tumor cells.ChemBioChem. 2009; 10: 862-868Crossref PubMed Scopus (293) Google Scholar Recently, through click chemistry, this aptamer has also been functionalized with radionuclides for imaging, although this could just as easily be used for therapeutic or theranostic applications.35Sicco E. Baez J. Ibarra M. Fernández M. Cabral P. Moreno M. Cerecetto H. Calzada V. Sgc8-c Aptamer as a Potential Theranostic Agent for Hemato-Oncological Malignancies.Cancer Biother. Radiopharm. 2020; 35: 262-270Crossref PubMed Scopus (3) Google Scholar Although this study tested the aptamer in an in vivo B cell lymphoma model, it did show biodistribution in the bone, with sustained accumulation over 3 days, suggesting potential for future therapeutic delivery to tackle the leukemic cells in the bone marrow niche. There are several more aptamers that have demonstrated promise for targeting leukemic cells that have been reviewed elsewhere.36Yang S. Li H. Xu L. Deng Z. Han W. Liu Y. Jiang W. Zu Y. Oligonucleotide Aptamer-Mediated Precision Therapy of Hematological Malignancies.Mol. Ther. Nucleic Acids. 2018; 13: 164-175Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar However, one worth highlighting in this review is the CD117 aptamer.37Zhao N. Pei S.-N. Qi J. Zeng Z. Iyer S.P. Lin P. Tung C.-H. Zu Y. Oligonucleotide aptamer-drug conjugates for targeted therapy of acute myeloid leukemia.Biomaterials. 2015; 67: 42-51Crossref PubMed Scopus (53) Google Scholar CD117 is a biomarker that appears to correlate with patient prognosis, whereby a higher level of expression is associated with a poorer prognosis. This is similar to PTK7, which is involved in resistance mechanisms to chemotherapy.38Goto H. Grantham R.W. Mizuuchi M. Yamaguchi T. Teshima T. Reed J.C. Matsuzawa S.-i. PTK7 Is Involved in the Chemotherapy Resistance of T-Cell Acute Lymphoblastic Leukemia through Interaction with FADD-Death Domain.Blood. 2015; 126: 3701Crossref Google Scholar This would suggest that the sgc8 aptamer and this CD117 aptamer have the potential to treat "hard-to-treat" leukemias. Another reason to highlight this aptamer is that with the use of click chemistry to attach methotrexate (MTX) to the aptamer, Zhao et al.37Zhao N. Pei S.-N. Qi J. Zeng Z. Iyer S.P. Lin P. Tung C.-H. Zu Y. Oligonucleotide aptamer-drug conjugates for targeted therapy of acute myeloid leukemia.Biomaterials. 2015; 67: 42-51Crossref PubMed Scopus (53) Google Scholar demonstrated a favorable result in patient samples. They demonstrated that, when incubated with primary bone marrow patient samples, the aptamer-MTX was able to kill the leukemic cells, whereas it had no effect on the healthy bone marrow cells. Both Dox and MTX have been used in targeted delivery as a means to reduce the toxicity of free drug dosing and not necessarily for overcoming drug resistance outside of drug efflux pumps. Although a low cellular accumulation of long-chain polyglutamates has been associated with poor therapeutic outcome, enzyme mutations and cytogenetic aberrations also contribute to MTX resistance.39Wojtuszkiewicz A. Peters G.J. van Woerden N.L. Dubbelman B. Escherich G. Schmiegelow K. Sonneveld E. Pieters R. van de Ven P.M. Jansen G. et al.Methotrexate resistance in relation to treatment outcome in childhood acute lymphoblastic leukemia.J. Hematol. Oncol. 2015; 8: 61Crossref PubMed Scopus (33) Google Scholar Although MTX has been used for decades and has known toxicities, simply having a higher accumulation in specific cell populations does not fix the inherent issue of drug resistance associated with enzyme mutations. One of the issues with hard-to-treat cancers is that the patient may appear in remission for an extended period but is actually relapsing. However, the current methods of detection are not sensitive enough to detect minimal residual disease (MRD). One very recent publication has detailed the generation of an aptamer specific to a patient diagnosed with multiple myeloma. Their aptamer isolation process is automatable and only requires a small amount of patient sera. Importantly, this aptamer was 2000-fold more sensitive than immunofixation electrophoresis and was able to detect MRD at 6 months, whereas the conventional methods failed to diagnose MRD for 25 months. Although only a proof-of-principle study, it suggests that aptamers could play a much larger role in personalized medicine in the future.40Tapia-Alveal C. Olsen T.R. Worgall T.S. Personalized immunoglobulin apt