Low dose of IL-2 combined with rapamycin restores and maintains the long-term balance of Th17/Treg cells in refractory SLE patients.

BACKGROUND: The development of Systemic lupus erythematosus (SLE) has been associated with the balance of Th17 and Treg cells. IL-2 and rapamycin can influence the populations of both Th17 and Treg cells. However, it is unclear whether low dose of IL-2 and rapamycin can relieve the symptoms of SLE patients and what is the mechanisms. In this study, we aim to analyze the effect of low dose of IL-2 plus rapamycin on the number of Tregs, Th17 cells and the ratio of Th17/Treg cells, as well as to evaluate its therapeutic efficacy in refractory SLE patients. RESULT: Fifty refractory SLE patients and 70 healthy controls were enrolled and followed up for 24 weeks. We found that compared with HC, the refractory SLE patients had a lower number of Tregs, a similar number of Th17 cells, but an increased ratio of Th17/Treg. After the treatment, the number of Tregs of the patients at 12th and 24th week was significantly increased. While the number of Th17 cells was unchanged, the ratio of Th17/Treg was significantly decreased at both 6 weeks and 24 weeks. After 6, 12 and 24 weeks of treatment, the SLEDAI score was significantly reduced. The prednison dosage at 6th,12th and 24th week post treatment was significantly decreased. CONCLUSION: Our results support that the reduction of Tregs and the imbalance of Th17/Treg cells were correlated with the occurrence and development of refractory SLE. Low dose of IL-2 combined with rapamycin was able to restore the number of Tregs and the balance of Th17/Treg cells. As a result, this approach was able to induce immune tolerance and promote disease remission, allowing for the reduction in prednisone dosage. TRIAL REGISTRATION: ChiCTR-IPR-16009451 Registration date: 2016/10/16.

Delivery of siRNA Complexed with Palmitoylated α-Peptide/ß-Peptoid Cell-Penetrating Peptidomimetics: Membrane Interaction and Structural Characterization of a Lipid-Based Nanocarrier System.

Proteolytically stable α-peptide/ß-peptoid peptidomimetics constitute promising cell-penetrating carrier candidates exhibiting superior cellular uptake as compared to commonly used cell-penetrating peptides (CPPs). The aim of the present study was to explore the potential of these peptidomimetics for delivery of small interfering RNA (siRNA) to the cytosol by incorporation of a palmitoylated peptidomimetic construct into a cationic lipid-based nanocarrier system. The optimal construct was selected on the basis of the effect of palmitoylation and the influence of the length of the peptidomimetic on the interaction with model membranes and the cellular uptake. Palmitoylation enhanced the peptidomimetic adsorption to supported lipid bilayers as studied by ellipsometry. However, both palmitoylation and increased peptidomimetic chain length were found to be beneficial in the cellular uptake studies using fluorophore-labeled analogues. Thus, the longer palmitoylated peptidomimetic was chosen for further formulation of siRNA in a dioleoylphosphatidylethanolamine/cholesteryl hemisuccinate (DOPE/CHEMS) nanocarrier system, and the resulting nanoparticles were found to mediate efficient gene silencing in vitro. Cryo-transmission electron microscopy (cryo-TEM) revealed multilamellar, onion-like spherical vesicles, and small-angle X-ray scattering (SAXS) analysis confirmed that the majority of the lipids in the nanocarriers were organized in lamellar structures, yet coexisted with a hexagonal phase, which is important for efficient nanocarrier-mediated endosomal escape of siRNA ensuring cytosolic delivery. The present work is a proof-of-concept for the use of α-peptides/ß-peptoid peptidomimetics in an efficient delivery system that may be more generally exploited for the intracellular delivery of biomacromolecular drugs.

Membrane adsorption and binding, cellular uptake and cytotoxicity of cell-penetrating peptidomimetics with α-peptide/ß-peptoid backbone: effects of hydrogen bonding and α-chirality in the ß-peptoid residues.

Cell-penetrating peptides (CPPs) provide a promising approach for enhancing intracellular delivery of therapeutic biomacromolecules by increasing transport through membrane barriers. Here, proteolytically stable cell-penetrating peptidomimetics with α-peptide/ß-peptoid backbone were studied to evaluate the effect of α-chirality in the ß-peptoid residues and the presence of guanidinium groups in the α-amino acid residues on membrane interaction. The molecular properties of the peptidomimetics in solution (surface and intramolecular hydrogen bonding, aqueous diffusion rate and molecular size) were studied along with their adsorption to lipid bilayers, cellular uptake, and toxicity. The surface hydrogen bonding ability of the peptidomimetics reflected their adsorbed amounts onto lipid bilayers as well as with their cellular uptake, indicating the importance of hydrogen bonding for their membrane interaction and cellular uptake. Ellipsometry studies further demonstrated that the presence of chiral centers in the ß-peptoid residues promotes a higher adsorption to anionic lipid bilayers, whereas circular dichroism results showed that α-chirality influences their overall mean residue ellipticity. The presence of guanidinium groups and α-chiral ß-peptoid residues was also found to have a significant positive effect on uptake in living cells. Together, the findings provide an improved understanding on the behavior of cell-penetrating peptidomimetics in the presence of lipid bilayers and live cells.

Interaction of peptidomimetics with bilayer membranes: biophysical characterization and cellular uptake.

Enzymatically stable cell-penetrating α-peptide/ß-peptoid peptidomimetics constitute promising drug delivery vehicles for the transport of therapeutic biomacromolecules across membrane barriers. The aim of the present study was to elucidate the mechanism of peptidomimetic-lipid bilayer interactions. A series of peptidomimetics consisting of alternating cationic and hydrophobic residues displaying variation in length and N-terminal end group were applied to fluid-phase, anionic lipid bilayers, and their interaction was investigated using isothermal titration calorimetry (ITC) and ellipsometry. Titration of lipid vesicles into solutions of peptidomimetics resulted in exothermic adsorption processes, and the interaction of all studied peptidomimetics with anionic lipid membranes was found to be enthalpy-driven. The enthalpy and Gibbs free energy (ΔG) proved more favorable with increasing chain length. However, not all charges contribute equally to the interaction, as evidenced by the charge-normalized ΔG being inversely correlated to the sequence length. Ellipsometry data suggested that the hydrophobic residues also played an important role in the interaction process. Furthermore, ΔG extracted from ellipsometry data showed good agreement with that obtained with ITC. To further elucidate their interaction with biological membranes, quantitative uptake and cellular distribution were studied in proliferating HeLa cells by flow cytometry and confocal microscopy. The cellular uptake of carboxyfluorescein-labeled peptidomimetics showed a similar ranking as that obtained from the adsorbed amount, and binding energy to model membranes demonstrated that the initial interaction with the membrane is of key importance for the cellular uptake.

99mTc-3PRGD2 for integrin receptor imaging of lung cancer: a multicenter study.

UNLABELLED: (99m)Tc-3PRGD2 is a new SPECT tracer targeting integrin α(V)ß(3) receptor for detecting tumors, imaging angiogenesis, and evaluating tumor response to therapy. A multicenter study was designed to investigate the efficacy of (99m)Tc-3PRGD2 for the evaluation of patients with lung cancer. METHODS: Seventy patients (51 men, 19 women; mean age ± SD, 63 ± 9 y) with a suspected lung lesion and for whom definite pathologic diagnosis was finally obtained (malignant, n = 58; benign, n = 12) were recruited from 6 centers. Whole-body planar scanning and chest SPECT were performed at 1 and 4 h, respectively, after intravenous injection of 11.1 MBq/kg (0.3 mCi/kg) of (99m)Tc-3PRGD2. The images were read in consensus by 6 experienced nuclear medicine physicians masked to the source, history, and pathologic diagnosis. The tumor-to-background (T/B) ratios were calculated for semiquantitative analysis. A Student t test was used for statistical analysis, and a P value less than 0.05 was considered significant. RESULTS: With low (99m)Tc-3PRGD2 background in the lungs and mediastinum, most lung malignancies were prominent on the 1-h images (T/B ratio, 1.65 ± 0.47 for the planar imaging and 2.78 ± 1.52 for SPECT). The T/B ratios were significantly lower in the benign lesions (P < 0.05). The sensitivity was 88% for semiquantitative analysis and could reach 93%-97% in visual analysis when considering the volume effect, necrosis, and metastasis. However, the specificity was only 58%-67%. Most lymph node and bone metastases could also be detected. CONCLUSION: (99m)Tc-3PRGD2 imaging at 1 h is sensitive for the detection of lung cancer, meriting further investigation of (99m)Tc-3PRGD2 as a novel clinical tracer for integrin receptor imaging.