An L-ribonucleic acid aptamer (L-RNA aptamer, trade name Spiegelmer) is an

RNA Ribonucleic acid (RNA) is a polymeric molecule that is essential for most biological functions, either by performing the function itself (non-coding RNA) or by forming a template for the production of proteins (messenger RNA). RNA and deoxyrib ...

-like molecule built from L-

ribose Ribose is a simple sugar and carbohydrate with molecular formula C5H10O5 and the linear-form composition H−(C=O)−(CHOH)4−H. The naturally occurring form, , is a component of the ribonucleotides from which RNA is built, and so this comp ...

units. It is an artificial

oligonucleotide Oligonucleotides are short DNA or RNA molecules, oligomers, that have a wide range of applications in genetic testing, Recombinant DNA, research, and Forensic DNA, forensics. Commonly made in the laboratory by Oligonucleotide synthesis, solid-phase ...

named for being a

mirror image A mirror image (in a plane mirror) is a reflection (physics), reflected duplication of an object that appears almost identical, but is reversed in the direction perpendicular to the mirror surface. As an optical phenomenon, optical effect, it r ...

of natural oligonucleotides. L-RNA aptamers are a form of

aptamer Aptamers are oligomers of artificial ssDNA, RNA, Xeno nucleic acid, XNA, or peptide that ligand, bind a specific target molecule, or family of target molecules. They exhibit a range of affinities (Dissociation constant, KD in the pM to μM rang ...

s. Due to their L-nucleotides, they are highly resistant to degradation by

nucleases In biochemistry, a nuclease (also archaically known as nucleodepolymerase or polynucleotidase) is an enzyme capable of cleaving the phosphodiester bonds that link nucleotides together to form nucleic acids. Nucleases variously affect single and ...

. L-RNA aptamers are considered potential drugs and are currently being tested in clinical trials.

Features

Chemical properties

L-RNA aptamers, built using L-ribose, are the enantiomers of natural oligonucleotides, which are made with D-ribose. Nucleic acid aptamers, including L-RNA aptamers, contain

adenosine monophosphate Adenosine monophosphate (AMP), also known as 5'-adenylic acid, is a nucleotide. AMP consists of a phosphate group, the sugar ribose, and the nucleobase adenine. It is an ester of phosphoric acid and the nucleoside adenosine. As a substituent it t ...

guanosine monophosphate Guanosine monophosphate (GMP), also known as 5′-guanidylic acid or guanylic acid (conjugate base guanylate), is a nucleotide that is used as a monomer in RNA. It is an ester of phosphoric acid with the nucleoside guanosine. GMP consists of the p ...

cytidine monophosphate Cytidine monophosphate, also known as 5'-cytidylic acid or simply cytidylate, and abbreviated CMP, is a nucleotide that is used as a monomer in RNA. It is an ester of phosphoric acid with the nucleoside cytidine. CMP consists of the phosphate grou ...

uridine monophosphate Uridine monophosphate (UMP), also known as 5′-uridylic acid ( conjugate base uridylate), is a nucleotide that is used as a monomer in RNA. It is an ester of phosphoric acid with the nucleoside uridine. UMP consists of the phosphate group, th ...

, a

phosphate group Phosphates are the naturally occurring form of the element phosphorus. In chemistry, a phosphate is an anion, salt, functional group or ester derived from a phosphoric acid. It most commonly means orthophosphate, a derivative of orthophosp ...

, a

nucleobase Nucleotide bases (also nucleobases, nitrogenous bases) are nitrogen-containing biological compounds that form nucleosides, which, in turn, are components of nucleotides, with all of these monomers constituting the basic building blocks of nuc ...

and a ribose sugar.

Biological characteristics

Like other aptamers, L-RNA aptamers are able to bind molecules such as

peptides Peptides are short chains of amino acids linked by peptide bonds. A polypeptide is a longer, continuous, unbranched peptide chain. Polypeptides that have a molecular mass of 10,000 Dalton (unit), Da or more are called proteins. Chains of fewer t ...

proteins Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, re ...

, and substances of low molecular weight. The affinity of L-RNA aptamers to their target molecules often lies in the pico to nanomolar range and is thus comparable to

antibodies An antibody (Ab) or immunoglobulin (Ig) is a large, Y-shaped protein belonging to the immunoglobulin superfamily which is used by the immune system to identify and neutralize antigens such as bacteria and viruses, including those that caus ...

. L-RNA aptamers themselves have low antigenicity. In contrast to other aptamers, L-RNA aptamers have high stability in blood serum, since they are less susceptible to be cleaved hydrolytically by enzymes. They are excreted by the

kidneys In humans, the kidneys are two reddish-brown bean-shaped blood-filtering organs that are a multilobar, multipapillary form of mammalian kidneys, usually without signs of external lobulation. They are located on the left and right in the retro ...

in a short time due to their low molar mass (which is below the renal threshold). L-RNA aptamers modified with a higher molar mass, such as

PEGylated PEGylation (or pegylation) is the process of both covalent and non-covalent attachment or amalgamation of polyethylene glycol (PEG, in pharmacy called macrogol) polymer chains to molecules and macrostructures, such as a drug, therapeutic protein ...

L-RNA aptamers, show a prolonged plasma half-life.

Production

Unlike other aptamers, L-RNA aptamers are not directly made using systematic evolution of ligands by exponential enrichment (SELEX), as L-nucleic acids are not amenable to enzymatic methods, such as

polymerase chain reaction The polymerase chain reaction (PCR) is a method widely used to make millions to billions of copies of a specific DNA sample rapidly, allowing scientists to amplify a very small sample of DNA (or a part of it) sufficiently to enable detailed st ...

(PCR), used in SELEX. Therefore, the selection is done with mirrored target molecules.

Reflection of the target molecule

The first step is the production of the target's

enantiomer In chemistry, an enantiomer (Help:IPA/English, /ɪˈnænti.əmər, ɛ-, -oʊ-/ Help:Pronunciation respelling key, ''ih-NAN-tee-ə-mər''), also known as an optical isomer, antipode, or optical antipode, is one of a pair of molecular entities whi ...

. In the case of peptides and small proteins that are produced synthetically, an enantiomer is made using synthetic D-

amino acid Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although over 500 amino acids exist in nature, by far the most important are the 22 α-amino acids incorporated into proteins. Only these 22 a ...

s. If the target is a larger protein molecule, beyond synthetic abilities, the enantiomer of an

epitope An epitope, also known as antigenic determinant, is the part of an antigen that is recognized by the immune system, specifically by antibodies, B cells, or T cells. The part of an antibody that binds to the epitope is called a paratope. Although e ...

is produced.

SELEX

Conventional (up to 10¹⁶ different oligonucleotides) existing molecule library serves as a starting point for the subsequent SELEX process. Selection, separation, and amplification using the mirror image of the target molecule is performed.

Sequencing and synthesis

The sequence of the oligonucleotide selected using SELEX is determined with the help of

DNA sequencing DNA sequencing is the process of determining the nucleic acid sequence – the order of nucleotides in DNA. It includes any method or technology that is used to determine the order of the four bases: adenine, thymine, cytosine, and guanine. The ...

. This information is used for the synthesis of the oligonucleotide's enantiomer, the L-RNA aptamer, using L-nucleotides.

Use

L-RNA aptamers have been obtained for the

chemokines Chemokines (), or chemotactic cytokines, are a family of small cytokines or signaling proteins secreted by cells that induce directional movement of leukocytes, as well as other cell types, including endothelial and epithelial cells. In addit ...

CCL2 The chemokine (C-C motif) ligand 2 (CCL2) is also referred to as monocyte chemoattractant protein 1 (MCP1) and small inducible cytokine A2. CCL2 is a small cytokine that belongs to the CC chemokine family. CCL2 tightly regulates cellular mecha ...

and

CXCL12 The stromal cell-derived factor 1 (SDF-1), also known as C-X-C motif chemokine 12 (CXCL12), is a chemokine protein that in humans is encoded by the ''CXCL12'' gene on chromosome 10. It is ubiquitously expressed in many tissues and cell types. St ...

, the complement components C5a and

ghrelin Ghrelin (; or lenomorelin, INN) is a hormone primarily produced by enteroendocrine cells of the gastrointestinal tract, especially the stomach, and is often called a "hunger hormone" because it increases the drive to eat. Blood levels of ghrel ...

. They are currently in preclinical or clinical development. Proof-of-concept for an anti-CCL2/MCP-1 L-RNA aptamers has recently been demonstrated in diabetic nephropathy patients. They can also be used as diagnostic agents.

References

{{reflist Nucleic acids Peptides Biotechnology