Discovery of targeted nucleic acid small molecules: Cell-SELEX technology-CAR-T Molecule Discovery Topic-Tekbiotech-Yeast and Phage Display CRO, Expert in Nano-body and Antibody Drug Development

Discovery of targeted nucleic acid small molecules: Cell-SELEX technology

Background

We all understand the concept of a key fitting a lock. Developing targeted drugs aims to create a "key" that can precisely open or lock a disease. But what if we don't even know what the "lock" looks like? The surfaces of many disease targets, especially cancer cells, display numerous unknown markers. Conventional methods involve purifying a single antigen and then designing experiments to screen for drugs against that purified protein. This entire process is time-consuming and labor-intensive. Furthermore, it can fail because the purified antigen may have lost its native structure.

Cell-SELEX technology was developed to address this. The core of this technology is the nucleic acid aptamer. An aptamer is a short strand of DNA or RNA. Because aptamers can fold into complex three-dimensional structures, they can "grab" target proteins with high affinity and specificity. They are often referred to as "chemical antibodies". Compared to traditional antibodies, aptamers offer several advantages, including smaller size, better tissue penetration, higher thermal stability, lower immunogenicity, easier production, lower synthesis costs, and facilitated conjugation or modification with different functional moieties ^[1]. They can be selected against a wide range of target molecules, including proteins, cells, microorganisms, and compounds. Therefore, aptamers are powerful alternatives to traditional antibodies in the fields of biomarker discovery, diagnostics, and imaging.

Principle

The technology consists of three main steps, as illustrated^[2]. In the initial rounds, to sufficiently enrich for positive aptamers, the starting nucleic acid aptamer library is first incubated with target cells under specific conditions and for a defined time. Next, after several washes, unbound aptamers are removed, while bound aptamers are retained. Finally, these bound aptamers are amplified by PCR to generate the new starting library for the next round. In addition to this forward selection, to ensure accuracy and specificity, a counter-selection step is introduced in later rounds. This involves incubating the eluted aptamers with non-target (negative) cells. Non-specific aptamers that bind to the negative cells are removed. The unbound sequences are then amplified by PCR, and the resulting library serves as the starting point for the next round of selection. Typically, 8-15 rounds are performed.

Figure 1: Schematic diagram of the Cell-SELEX principle ^[2]

Advantages and Applications

1.The cell surface contains a variety of molecules with complex structures, allowing for aptamer selection against multiple targets simultaneously.2.Molecules on the cell surface retain their native conformation, including post-translational modifications and correct folding.3.The aptamer library may identify novel, unknown membrane proteins.

Based on this technology, applications include the development of targeted drugs using aptamers to precisely attack cancer cells, the creation of diagnostic kits for cancer detection, and the utilization of aptamers as novel molecular probes for real-time observation of membrane protein dynamics.

Challenges and Prospects

Cell-SELEX itself still presents several issues. For example, the entire screening process is relatively lengthy and labor-intensive due to the high number of cycles. Furthermore, because of the complexity of cell surface molecules, it is difficult to standardize the selection process across different targets. Therefore, optimizing the aptamer screening workflow is necessary. In the future, with further technological development, it is hoped that this technology can be combined with artificial intelligence. We could leverage AI's powerful computational and analytical capabilities to predict aptamer structures, potentially reducing the number of selection cycles, shortening development time, and lowering costs.

TekBiotech has established a comprehensive nucleic acid aptamer selection platform. We are dedicated to providing high-quality aptamer discovery services to scientists worldwide, including one-stop solutions encompassing library design, SELEX screening, candidate sequencing and characterization, affinity validation, and downstream functional validation. Aligned with the evolving needs of life sciences, we develop innovative targeted therapeutic tools for challenging disease targets, providing robust support for our clients' research projects.

References

[1] Bayat P, Nosrati R, Alibolandi M, et al. SELEX methods on the road to protein targeting with nucleic acid aptamers. Biochimie. 2018.

[2] Chen C, Zhou S, Cai Y, et al. Nucleic acid aptamer application in diagnosis and therapy of colorectal cancer based on cell-SELEX technology. NPJ Precis Oncol. 2017;1(1):37.

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