The hemostasis system is a complex structure that includes the fibrinolysis system, and Yes this is correct coagulation and anticoagulation parts. Due to the multicomponent nature, it becomes relevant to study the key changes in the functioning of signaling pathways, and develop new diagnostic methods and modern drugs with high selectivity. One of the ways to solve this problem is the development of molecular recognition elements capable of blocking one of the hemostasis systems and/or activating another. Aptamers can serve as ligands for targeting specific clinical needs, promising anticoagulants with minor side effects and significant biological activity. Aptamers with several clotting factors and platelet proteins are used for the treatment of thrombosis. This review is focused on the aptamers used for the correction of the hemostasis system, and their structural and functional features. G-rich nucleic acid aptamers, mostly versatile G-quadruplexes, recognize different components of the hemostasis system and are capable of correcting the functioning.

One of the promising novel methods for radical tumor resection at a single-cell level is magneto-mechanical microsurgery (MMM) with magnetic nano- or microdisks modified with cancer-recognizing molecules. A low-frequency alternating magnetic field (AMF) remotely drives and controls the procedure. Here, we present characterization and application of magnetic nanodisks (MNDs) as a surgical instrument (“smart nanoscalpel”) at a single-cell level. MNDs with a quasi-dipole three-layer structure (Au/Ni/Au) and DNA aptamer AS42 (AS42-MNDs) on the surface converted magnetic moment into mechanical and destroyed tumor cells. The effectiveness of MMM was analyzed on Ehrlich ascites carcinoma (EAC) cells in vitro and in vivo using sine and square-shaped AMF with frequencies from 1 to 50 Hz with 0.1 to 1 duty-cycle parameters. MMM with the “Nanoscalpel” in a sine-shaped 20 Hz AMF, a rectangular-shaped 10 Hz AMF, and a 0.5 duty cycle was the most effective. A sine-shaped field caused apoptosis, whereas a rectangular-shaped field caused necrosis. Four sessions of MMM with AS42-MNDs significantly reduced the number of cells in the tumor. In contrast, ascites tumors continued to grow in groups of mice and mice treated with MNDs with nonspecific oligonucleotide NO-MND. Thus, applying a “smart nanoscalpel” is practical for the microsurgery of malignant neoplasms.

Background: Oncological diseases are a major focus in medicine, with millions diagnosed each year, leading researchers to seek new diagnostic and treatment methods. One promising avenue is the development of targeted therapies and rapid diagnostic tests using recognition molecules. The pharmaceutical industry is increasingly exploring nucleic acid-based therapeutics. However, producing long oligonucleotides, especially aptamers, poses significant production challenges. Objectives: This study aims to demonstrate the efficacy of using molecular modeling, supported by experimental procedures, for altering aptamer nucleotide sequences while maintaining their binding capabilities. The focus is on reducing production costs and enhancing binding dynamics by removing nonfunctional regions and minimizing nonspecific binding. Methods: A molecular modeling approach was employed to elucidate the structure of a DNA aptamer, Gli-55, facilitating the truncation of nonessential regions in the Gli-55 aptamer, which selectively binds to glioblastoma (GBM). This process aimed to produce a truncated aptamer, Gli-35, capable of forming similar structural elements to the original sequence with reduced nonspecific binding. The efficiency of the truncation was proved by flow cytometry, fluorescence polarization (FP), and confocal microscopy. Results: The molecular design indicated that the new truncated Gli-35 aptamer retained the structural integrity of Gli-55. In vitro studies showed that Gli-35 had a binding affinity comparable to the initial long aptamer while the selectivity increased. Gli-35 internalized inside the cell faster than Gli-55 and crossed the blood–brain barrier (BBB), as demonstrated in an in vitro model. Conclusions: The success of this truncation approach suggests its potential applicability in scenarios where molecular target information is limited. The study highlights a strategic and resource-efficient methodology for aptamer development. By employing molecular modeling and truncation, researchers can reduce production costs and avoid trial and error in sequence selection. This approach is promising for enhancing the efficiency of therapeutic agent development, particularly in cases lacking detailed molecular target insights.

Background: With the rising incidence of cancer, there is a growing need for improved preclinical models to test new therapies. While patient-derived xenografts (PDX) in immunodeficient mice are the gold standard, they are costly and result in a complete absence of a functional immune system, limiting their utility for studying tumor–immune interactions. This study characterizes a pharmacological partial immunosuppression protocol in immunocompetent mice as a promising alternative, evaluating its impact on the immune system and demonstrating its efficacy for growing human tumor xenografts. Methods: Mice received a regimen of cyclosporine (20 mg/kg, i.p., every 48 h for 12 days), cyclophosphamide (60 mg/kg, i.p., every 48 h for 8 days), and ketoconazole (10 mg/kg, p.o., for 12 days). The dynamics of CD3⁺, CD4⁺, CD8⁺, and CD19⁺ lymphocyte subpopulations and the CD4/CD8 index were monitored via flow cytometry on days 1, 5, 8, 12, 16, and 21. The protocol’s utility was tested by orthotopic transplantation of human glioma and lung cancer cells, and subcutaneous transplantation of breast cancer cells (MCF7). Tumor engraftment and growth were assessed using in vivo microscopy, MRI, and histology. Results: The immunosuppressive protocol induced a significant but partial reduction in CD3⁺ T-cells and CD19⁺ B-cells by day 8 (p = 0.0277). A profound and progressive decrease in the CD4/CD8 index was observed, indicating a shift towards immunosuppression. Crucially, CD8⁺ and CD4⁺ T-cells populations recovered rapidly post-therapy, demonstrating that the protocol creates a temporary and modifiable immune window rather than inducing complete ablation. The protocol enabled successful engraftment and growth of all three tested tumors in a residual immune microenvironment, confirmed by in vivo imaging and histopathological analysis. Conclusions: This drug-induced partial immunosuppression protocol effectively creates a reproducible state of transient immunodeficiency in outbred mice, suitable for various human tumor xenograft models. It represents a cost-effective and flexible alternative to genetic models, with the distinct advantage of preserving a residual immune microenvironment, making it particularly valuable for preclinical studies that require a partially intact host immune system.

BACKGROUND: Lung cancer (LC) is one of the most prevalent types of cancer, with mortality rates reaching 25% of all cancer-related deaths. The majority of patients are classified as stage IV at the time of diagnosis, a stage at which the probability of successful treatment is considerably low. Conventional diagnostic methods for lung cancer are expensive, labor-intensive, and highly invasive if a biopsy is performed. Consequently, liquid biopsy, which involves the extraction of circulating tumor cells from blood samples, has emerged as a pivotal area of research in oncology. AIM: To develop a method for the isolation and identification of circulating tumor cells in the peripheral blood of patients with lung cancer using DNA aptamers. MATERIALS AND METHODS: The study objects are lung cancer tissue, blood of patients with lung cancer and primary lung cancer cell culture. Gold-decorated magnetic nanoparticles and thiolated aptamers LC-17, LC-183 and LC-224 were used to isolate proteins. A hybrid of a thiol primer and a non-specific DNA sequence composed of two AG nucleotide repeats was used as a control. Mass spectrometry was performed using UltiMate 3000 nano-UHPLC system coupled with Orbitrap Fusion mass spectrometer (Thermo Scientific, USA). Circulating tumor cell counts were measured on CytoFLEX flow cytometer (Beckman Coulter, USA) after triple staining. Fluorescence microscopy was used for CSC visualization. RESULTS: The potential protein targets of the aptamers LC-17, LC-183, and LC-224 were identified. These aptamers were then used to isolate circulating tumor cells from the blood of patients with lung cancer. The identification of circulating tumor cells was performed using flow cytometry and fluorescence microscopy. CONCLUSIONS: The proposed method for the identification of circulating tumor cells through magnetic separation and flow cytometry provides a quantitative analysis of the target analyte, facilitated by the use of cell-specific lung cancer aptamers, including LC-17, LC-183, and LC-224, which exhibit high-affinity binding properties.

Background: Glioblastoma remains a challenging brain tumor to treat due to its infiltrative nature. Accurately identifying tumor boundaries during surgery is crucial for successful tumor resection. This study introduces an innovative intraoperative visualization method utilizing a surgical fluorescence microscope to precisely locate tumor cells. The focus of the study is on IR-Glint, a novel preparation comprising Cy7.5-labeled aptamers Gli-233nt and Gli-55_3L specific to human glioblastoma. Methods: The aptamers were modified using molecular modeling and quantum chemical techniques. The effectiveness of the preparation was assessed using flow cytometry and microscopy on primary cultures of human glioblastoma. In vivo studies were conducted on mouse and rabbit models, employing orthotopic xenotransplantation of human brain glioblastoma with various imaging techniques, including PET/CT, in vivo fluorescence visualization, confocal laser scanning, and surgical microscopy. Results: The experiments validated the potential of IR-Glint for intraoperative visualization of glioblastoma using infrared imaging. Surface application of the aptamer-based formulation on the brain allowed clear visualization of the tumor, aiding surgeons in tumor resection. This approach also reduces the dosage required and mitigates potential toxic effects on the patient. Conclusions: This study demonstrates the promising potential of using in-frared dye-labeled aptamers for intraoperative visualization of glioblastoma. By improving surgical treatment outcomes in neurosurgery, this novel approach may pave the way for future advancements in the field.