Common coupling methods include joints, oligo nucleic acid coupling, and liposome coupling. We fused a single aptamer with a 23bp double-stranded RNA adaptor protein and deliberately left 2-3 unpaired bases between the adaptor and the aptamer to provide space for each individual aptamer. Adaptors can be composed of various biosynthetic structural units, such as nucleotides, peptides, organic chains, and inorganic materials. For studies involving immunomodulatory bsApt, aptamer modules are often linked by oligonucleotide adaptors. Almost every bsApt construct has used a different sequence. The DNA aptamer with affinity for CD62L (aptamer LD201t1) present on naive T cells was coupled to other aptamers that recognize PTK 7 (aptamer sgc 8), PD-L1 (aptamer PD-L1) or whole Ramos cell (B cell lymphoma; aptamer TE02). These aptamer conjugates were joined together by the 5′and 3′ends of the called circular bsApts because the 5′and 3′ends were joined together using T4 DNA ligase after crossing the 13-nt flanking complementary sequence. But this coupling is because, without free ends, c-bsApts would be more resistant to degradation by serum exonuclease. In addition to the above nucleic acid-based ligation mode, a strategy using liposomes co-modified with aptamers exists as a ligation mode.

The bispecific aptamer can recognize two or more different cell surface receptors simultaneously and target two molecules simultaneously, showing better therapeutic effects and less drug resistance than traditional monoclonal antibodies. The dual-specificity molecule adopts a two-in-one form, has low administration complexity, and has a more favorable regulatory approval than the proposed combination of two single molecules. However, complex techniques such as high immunogenicity and production difficulties also affect antibody-based bispecific molecules. We developed an RNA-based dual-specificity aptamer capable of blocking both targets simultaneously using a single target aptamer to a double-stranded RNA adaptor. Bispecific aptamers were found to be more effective than individual aptamers in their ability to inhibit cell growth and induce apoptosis. When these aptamers were tested, the bispecific aptamer significantly inhibited intraperitoneal tumor growth more than a single aptamer alone or in combination. The enhanced efficacy of the bispecific aptamer is most likely attributed to its increased cycling time over the individual aptamers. Furthermore, we show that the bispecific aptamer is not toxic to the host and is unable to trigger innate immunogenicity.TEK Biotech has rich experience in screening aptamers, which can help customers better complete the screening and connection of dual-specific aptamers.In addition, a variety of small molecules, protein nucleic acid aptamer screening services.

Most immunomodulatory bsApt use ssDNA or double-stranded oligonucleotide adaptors of 15-25 nt (105-175 A). The researchers found that the joint length of 7-22 nt (49-152 A) mediated a significant cytotoxic effect, but not over 29 nt (200 A). The joint composition also plays a role in determining its rigidity, which affects the conformational entropy and 3D presentation of the binding domain. Most studies have used double-stranded adaptors and thus constitute a more rigorous design. Only two groups used single-stranded joints with lower stiffness. It was also shown that flexible joints containing both ssDNA and double-stranded DNA (dsDNA) parts (versus dsDNA alone) were less effective in forming intercellular junctions, which they attributed to poor thermal stability. Other studies suggest that two or three unpaired nucleotides flanking its 23-nt adaptor sequence are essential for function. When bsApt lost their specific and cytotoxic effects, using fully paired adaptors or those with only one unpaired nucleotide. These unpaired nucleotides provide each aptamer with the spatial degree of freedom to form its 3D structure. The adaptors used in each study show that various features are acceptable for a proof-of-concept design as long as the adaptors do not affect the folding and stability of the appropriate gamete.

Immunomodulatory bsApts can be classified according to the valence and the specificity of the two aptamer modules, where [m] corresponds to the valence of the tumor-targeting aptamer module and [n] corresponds to the valence of the immune cell-targeting gamete module. For example, the [1 + 2] name describes a construct consisting of an aptamer that binds the target tumor cell and two aptamers that bind the immune cells. Immune cell-targeting aptamers for most agonistic co-stimulatory receptors do require a bivalent design ([n] = 2) because crosslinking of the receptor through its dimeric form is required to induce signaling and T cell activation. Other aptamers that function as monomers (e.g., localization to TAA, natural killer [NK] cell activation, antagonism) can be designed using [1 + 1]. The primary goal of some bsApt with increased transparency is to increase the affinity of their targets, both on tumors and immune cells. Although no triple-specific immunomodulatory aptamers are currently available in the literature, their design can still be classified using this nomenclature.[1 + 1] Design can effectively bring immune cells in close proximity to tumor cells to generate silent intercellular junctions or cytotoxic immune synapses, as illustrated by multiple measures.[1 + 2] design increases the valence of aptamers targeting immune cells. To date, all immunomodulatory bsApts described using this design contain the dimer aptamer, a costimulatory receptor on agonistic immune cells.

The researchers found that the dual-specific ring aptamer of CIL-6A6-1, which binds IL-6 and sIL-6R with high affinity, is stable in serum for more than 48 hours. It can effectively block the IL-6 / s IL-6R interaction and significantly inhibit cellular inflammation. Most importantly, this bispecific aptamer is far more effective than combining the aptamer alone as well as the commercially available tocilizumab, highlighting the advantage of choosing the bi-specific ring aptamer as a molecular tool for anti-inflammatory therapy. Researchers have developed a unique chemical signature of the TME as a pathway for the de-specific detection of tumor cells and their elevated protein expression. This was achieved by designing a bispecific aptamer with one arm to detect the highly expressed tumor-associated metabolites ATP and to detect cell surface markers expressed on other T cell lymphomas. Bispecific aptamers (bsApts) have been shown to induce artificial immune synapses that promote T cell activation and subsequent tumor cell lysis in various in vitro and in vivo preclinical models. Bispecific aptamers may also serve as a novel strategy for targeting the enhanced antitumor immune response against malignancies expressing MUC 1. TEK Biotech can prepare bispecific aptamers to support researchers in overcoming problems in cancer and immune diseases.