**1. Introduction**

Aptamers are small single-stranded RNA or DNA oligonucleotides that specifically bind to their target due to their unique 3-dimensional structure. They were first independently developed by the groups of Gold and Szostak in 1990 [1, 2]. Aptamers are selected from a pool of random oligonucleotide library (>1015 random sequences) by iterative rounds of selection and amplification by a process called Systematic Evolution of Ligands by EXponential Enrichment (SELEX). Different intermolecular interactions facilitate the interaction of aptamers with their target including van der Waal's forces, electrostatic interactions between charged groups, three-dimensional shape, stacking and hydrogen bonds.

Functionally aptamers are same as antibodies; however, their advantages over antibodies include small size, less immunogenicity, ease of synthesis, easy chemical modification that allows conjugation with a variety of molecules and stability at a higher temperature. Aptamer is comparably smaller (~3 nm) than an antibody (10–15 nm) [3], thus, allowing higher penetration in tissues and the ability to bind more dense cellular epitopes in living cells. Unlike antibodies, aptamers can be selected against non-immunogenic molecules, including proteins or peptides as well as toxins. Aptamers have been successfully developed against proteins, peptides, dyes, metal ions, viruses, bacteria, toxins, and whole cells. They have high specificity and affinity often comparable with antibodies. Unlike antibodies, they can be selected at non-physiological conditions such as extremely high or low temperature, or pH. Synthetic manufacturing of aptamers allows minimal batch-to-batch variation, which is a tedious task to maintain in antibody development. All these properties make aptamers as suitable candidates for therapeutic use, particularly targeted nano-delivery. Several aptamers have already been selected against cell surface targets for targeted delivery of therapeutic payload for different diseases, e.g., cancer and human immunodeficiency virus (HIV) infected cells [4]. **Table 1** lists the common differences between antibodies and aptamers.

In this book chapter, we discuss the challenges that hinder the clinical use of aptamers as targeted delivery molecules. For example, the small size of aptamers is like a double-edged sword. Though it helps in higher tissue penetration, but also results in high renal filtration that causes loss of its therapeutic efficiency. Using examples we show that how challenges in serum stability, renal filtration, selection


**3**

**Figure 1.**

*molecules.*

*Aptamers for Targeted Delivery: Current Challenges and Future Opportunities*

methodology and endocytic escape have been overcome. We explain the importance of proper controls and validation methods in SELEX methodology, which would otherwise result in the development of poor quality aptamers and, this might ultimately diminish the confidence in this technology. Lastly, we discuss about the development of automation in SELEX technology, an important step toward

Currently, many aptamers are in different stages of clinical trials for various diseases [5]. Most of the drugs are administered systemically, i.e., intravenously. This method poses a challenge of overcoming serum stability, which involves overcoming nuclease activity and coagulation. Here in we discuss these two topics in detail:

Aptamers have been used to functionalize liposomes to increase their bioavailability through targeted delivery (**Figure 1**) [4]. Since aptamers are negatively charged it is easier to pack them with cationic liposomes. However, serum proteins being charged molecules bind to cationic lipids, thus, lowering their targeting efficiency due to change in structure, aggregation or dissociation of the aptamercationic lipid complex [6, 7–9]. Polycationic lipids have also shown cyto(toxic) effects [10]. The problem of aggregation incurred by the charge of cationic lipids can be overcome by using neutral helper lipids [11, 12]. But another challenge is that lipid conjugated aptamers may also bind to erythrocytes, and filtered out through liver and spleen. For example, 1,2-dioleoyl phosphatidylethanolamine (DOPE), a neutral helper lipid, binds to erythrocytes. As an alternative, another neutral helper lipid, cholesterol, may be used which shows significantly lower binding to erythro-

Cholesterol conjugated RNA aptamer against Hepatitis C virus (HCV) NS5B protein has been used to inhibit HCV replication in human liver cells without induction of cytotoxicity *in vitro*. Next, in this study, the same molecule was injected in a mouse model but found no induction of innate immunity. The half-life of the cholesterol-conjugated aptamer was longer than that of the

*Illustration showing (a) non-functionalized liposome and (b) aptamer-functionalized liposome carrying drug* 

*DOI: http://dx.doi.org/10.5772/intechopen.84217*

industrializing the aptamer selection process.

**2. Serum stability**

**2.1 Aggregation**

cytes as compared to DOPE [10, 13].

**Table 1.**

*This table lists the common differences between an antibody (IgG) and an aptamer.*

*Aptamers for Targeted Delivery: Current Challenges and Future Opportunities DOI: http://dx.doi.org/10.5772/intechopen.84217*

methodology and endocytic escape have been overcome. We explain the importance of proper controls and validation methods in SELEX methodology, which would otherwise result in the development of poor quality aptamers and, this might ultimately diminish the confidence in this technology. Lastly, we discuss about the development of automation in SELEX technology, an important step toward industrializing the aptamer selection process.
