DNA Thioaptamer with Homing Specificity to Lymphoma Bone Marrow Involvement.

Selective drug accumulation in the malignant tissue is a prerequisite for effective cancer treatment. However, most drug molecules and their formulated particles are blocked en route to the destiny tissue due to the existence of multiple biological and physical barriers including the tumor microvessel endothelium. Since the endothelial cells on the surface of the microvessel wall can be modulated by inflammatory cytokines and chemokines secreted by the tumor or stromal cells, an effective drug delivery approach is to enhance interaction between the drug particles and the unique spectrum of surface proteins on the tumor endothelium. In this study, we performed in vivo screening for thioaptamers that bind to the bone marrow endothelium with specificity in a murine model of lymphoma with bone marrow involvement (BMI). The R1 thioaptamer was isolated based on its high homing potency to bones with BMI, and 40-60% less efficiency in accumulation to healthy bones. In cell culture, R1 binds to human umbilical vein endothelial cells (HUVEC) with a high affinity ( Kd ≈ 3 nM), and the binding affinity can be further enhanced when cells were treated with a mixture of lymphoma cell and bone marrow cell conditioned media. Cellular uptake of R1 is through clathrin-mediated endocytosis. Conjugating R1 on to the surface of liposomal doxorubicin nanoparticles resulted in 2-3-fold increase in drug accumulation in lymphoma BMI. Taking together, we have successfully identified a thioaptamer that preferentially binds to the endothelium of lymphoma BMI. It can serve as an affinity moiety for targeted delivery of drug particles to the disease organ.

A Novel DNA Aptamer for Dual Targeting of Polymorphonuclear Myeloid-derived Suppressor Cells and Tumor Cells.

Aptamers have the potential to be used as targeting ligands for cancer treatment as they form unique spatial structures. Methods: In this study, a DNA aptamer (T1) that accumulates in the tumor microenvironment was identified through in vivo selection and validation in breast cancer models. The use of T1 as a targeting ligand was evaluated by conjugating the aptamer to liposomal doxorubicin. Results: T1 exhibited a high affinity for both tumor cells and polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs). Treatment with T1 targeted doxorubicin liposomes triggered apoptosis of breast cancer cells and PMN-MDSCs. Suppression of PMN-MDSCs, which serve an immunosuppressive function, leads to increased intratumoral infiltration of cytotoxic T cells. Conclusion: The cytotoxic and immunomodulatory effects of T1-liposomes resulted in superior therapeutic efficacy compared to treatment with untargeted liposomes, highlighting the promise of T1 as a targeting ligand in cancer therapy.

NMR structure of the viral peptide linked to the genome (VPg) of poliovirus.

VPgs are essential for replication of picornaviruses, which cause diseases such as poliomyelitis, foot and mouth disease, and the common cold. VPg in infected cells is covalently linked to the 5' end of the viral RNA, or, in a uridylylated form, free in the cytoplasm. We show here the first solution structure for a picornaviral VPg, that of the 22-residue peptide from poliovirus serotype 1. VPg in buffer is inherently flexible, but a single conformer was obtained by adding trimethylamine N-oxide (TMAO). TMAO had only minor effects on the TOCSY spectrum. However, it increased the amount of structured peptide, as indicated by more peaks in the NOESY spectrum and an up to 300% increase in the ratio of normalized NOE cross peak intensities to that in buffer. The data for VPg in TMAO yielded a well defined structure bundle with 0.6 A RMSD (versus 6.6 A in buffer alone), with 10-30 unambiguous constraints per residue. The structure consists of a large loop region from residues 1 to 14, from which the reactive tyrosinate projects outward, and a C-terminal helix from residues 18 to 21 that aligns the sidechains of conserved residues on one face. The structure has a stable docking position at an area on the poliovirus polymerase crystal structure identified as a VPg binding site by mutagenesis studies. Further, UTP and ATP dock in a base-specific manner to the reactive face of VPg, held in place by residues conserved in all picornavirus VPgs.

Thioaptamer conjugated liposomes for tumor vasculature targeting.

Recent developments in multi-functional nanoparticles offer a great potential for targeted delivery of therapeutic compounds and imaging contrast agents to specific cell types, in turn, enhancing therapeutic effect and minimizing side effects. Despite the promise, site specific delivery carriers have not been translated into clinical reality. In this study, we have developed long circulating liposomes with the outer surface decorated with thioated oligonucleotide aptamer (thioaptamer) against E-selectin (ESTA) and evaluated the targeting efficacy and PK parameters. In vitro targeting studies using Human Umbilical Cord Vein Endothelial Cell (HUVEC) demonstrated efficient and rapid uptake of the ESTA conjugated liposomes (ESTA-lip). In vivo, the intravenous administration of ESTA-lip resulted in their accumulation at the tumor vasculature of breast tumor xenografts without shortening the circulation half-life. The study presented here represents an exemplary use of thioaptamer and liposome and opens the door to testing various combinations of thioaptamer and nanocarriers that can be constructed to target multiple cancer types and tumor components for delivery of both therapeutics and imaging agent.