Enhancing adoptive T cell therapy for solid tumor with cell-surface anchored immune checkpoint inhibitor nanogels.

The efficacy of Adoptive Cell Therapy (ACT) for solid tumor is still mediocre. This is mainly because tumor cells can hijack ACT T cells' immune checkpoint pathways to exert immunosuppression in the tumor microenvironment. Immune Checkpoint Inhibitors such as anti-PD-1 (aPD1) can counter the immunosuppression, but the synergizing effects of aPD1 to ACT was still not satisfactory. Here we demonstrate an approach to safely anchor aPD1-formed nanogels onto T cell surface via bio-orthogonal click chemistry before adoptive transfer. The spatial-temporal co-existence of aPD1 with ACT T cells and the responsive drug release significantly improved the treatment outcome of ACT in murine solid tumor model. The average tumor weight of the group treated by cell-surface anchored aPD1 was only 18 % of the group treated by equivalent dose of free aPD1 and T cells. The technology can be broadly applicable in ACTs employing natural or Chimeric Antigen Receptor (CAR) T cells.

Scale up of Transmembrane NADH Oxidation in Synthetic Giant Vesicles.

The transfer of electrons across and along biological membranes drives the cellular energetics. In the context of artificial cells, it can be mimicked by minimal means, while using synthetic alternatives of the phospholipid bilayer and the electron-transducing proteins. Furthermore, the scaling up to biologically relevant and optically accessible dimensions may provide further insight and allow assessment of individual events but has been rarely attempted so far. Here, we visualized the mediated transmembrane oxidation of encapsulated NADH in giant unilamellar vesicles via confocal laser scanning and time-correlated single photon counting wide-field microscopy. To this end, we first augmented phospholipid membranes with an amphiphilic copolymer in order to check its influence on the oxidation kinetics spectrophotometrically. Then, we scaled up the compartments and followed the process microscopically.

Characterization and properties of phenolic resin doped modified lignin.

Phenolic resins occupy an important position in industrial applications, but phenol, one of the raw materials for synthesis, is a non-renewable resource. Lignin, as a natural polymer containing phenolic hydroxyl groups, alcohol hydroxyl groups and other reactive groups, can replace some of the phenol in the synthesis of phenolic resins, which can reduce the amount of phenol, thus reducing the cost of phenolic resins, while effectively promoting the high value-added use of renewable biomass resources. Due to its low reactivity, alkaline lignin is usually discharged as production waste, unaware that lignin macromolecules can be modified. In this paper, the phenolic monomers were obtained by acid-catalyzed depolymerization of DES (choline chloride/p-toluenesulfonic acid or choline chloride/lactic acid) from waste alkaline lignin, and the recovery rate of the DES solution during the catalytic treatment was more than 85 %, in which the main monomer was 2-methoxy-4-(1-propyl) phenol. The degradation of alkaline lignin is still favorable after five times of DES solvent recovery. The depolymerized lignin monomer replaced phenol by 50 wt% and then ternary co-polymerized with phenol and formaldehyde to form a biomass phenol-based phenolic resin, providing a green route for phenolic resin production. The cost of resin preparation was economically calculated, and it was found that the cost of resin after accumulating 4 cycles of DES treatment was only 51.1 % of that of pure phenolic resin. The density functional theory (DFT) was used to simulate the possible radical reactions in the intermediate process of phenolic resin reaction, to explore the microscopic mechanism and competition, to provide theoretical reference for further experimental realization of resin structure control and optimization, and to improve the theoretical system of resin synthesis.

Anisotropic chitosan/tunicate cellulose nanocrystals hydrogel with tunable interference color and acid-responsiveness.

A robust chitosan/tunicate cellulose nanocrystals (TCNCs) anisotropic hydrogel with bright interference colors was fabricated via combining the prestretching orientation method and chemically-physically dual cross-linking. The oriented regenerated chitosan nanofibrous network enabled the TCNCs alignment by covalent interaction and hydrogen bonding. The stretching alignment endows the chitosan/TCNCs hydrogel with enhanced tensile strength, from 0.63 MPa (draw ratio 1.0) to 2.06 MPa (draw ratio 3.5). Moreover, the orientation of chitosan nanofibers led to birefringence appearance, which could be regulated with the TCNCs introduction or draw ratios. The hydrogel swelled completely in 2 min in pH = 3 solution and the interference color disappeared. The oriented chitosan/TCNCs hydrogels showed distinct color change under acid stimulation, which could be quantitatively measured or directly observed under crossed polarizers. This work demonstrated a strategy for fabricating the interference color regulatable hydrogels with acid-response property for sensors and environmental monitoring.

Cationic polymer synergizing with chemotherapeutics and re-purposing antibiotics against cancer cells.

Chemotherapy is one of the most effective treatments for cancer. However, toxicity and the development of drug resistance have become the major hurdles to the commonly used chemotherapeutics such as doxorubicin and paclitaxel. Antibiotics have also been used as anti-cancer drugs due to their anti-proliferative and cytotoxic effects. However, these anti-tumor antibiotics like ciprofloxacin face the similar resistance and toxicity issues. In this study, we used a quaternary ammonium-functionalized cationic polycarbonate to synergize with the existing chemotherapeutics and re-purpose antibiotics to address the resistance and toxicity issues. When used in combination with the drugs, the cationic polymer induced 2-3 fold more damage in the cancer cell membrane within 2 hours, thus enhancing the uptake of chemotherapeutics up to 2.5 fold more into the breast, liver and even chemotherapeutics-resistant cancer cells. On the other hand, the chemotherapeutics increased the cellular uptake of polymer. The combined effects resulted in 3-10 fold reduction in IC50 of chemotherapy drugs and yielded therapeutic synergy at a clinically-relevant concentration range of drugs when treating multiple types of cancer cells, while the use of guanidinium-functionalized polymer capable of membrane translocation did not lead to a synergistic effect. Thus, the quaternary ammonium-functionalized cationic polymer can increase the therapeutic efficacies of existing drugs, mitigating toxicities by lowering required dosage and circumventing drug resistance via its membrane disruption mechanism. The findings of this study provide insights into designing future anticancer therapy.

Redox-responsive interleukin-2 nanogel specifically and safely promotes the proliferation and memory precursor differentiation of tumor-reactive T-cells.

Interleukin-2 (IL-2) is a potent T-cell mitogen that can adjuvant anti-cancer adoptive T-cell transfer (ACT) immunotherapy by promoting T-cell engraftment. However, the clinical applications of IL-2 in combination with ACT are greatly hindered by the severe adverse effects such as vascular leak syndrome (VLS). Here, we developed a synthetic delivery strategy for IL-2 via backpacking redox-responsive IL-2/Fc nanogels (NGs) to the plasma membrane of adoptively transferred T-cells. The NGs prepared by traceless chemical cross-linking of cytokine proteins selectively released the cargos in response to T-cell receptor activation upon antigen recognition in tumors. We found that IL-2/Fc delivered by T-cell surface-bound NGs expanded transferred tumor-reactive T-cells 80-fold more than the free IL-2/Fc of an equivalent dose administered systemically and showed no effects on tumor-infiltrating regulatory T-cell expansion. Intriguingly, IL-2/Fc NG backpacks that facilitated a sustained and slow release of IL-2/Fc also promoted the CD8+ memory precursor differentiation and induced less T-cell exhaustion in vitro compared to free IL-2/Fc. The controlled responsive delivery of IL-2/Fc enabled the safe administration of repeated doses of the stimulant cytokine with no overt toxicity and improved efficacy against melanoma metastases in a mice model.

Polymers with distinctive anticancer mechanism that kills MDR cancer cells and inhibits tumor metastasis.

Although mortality continues to decline over the past two decades, cancer is still a pervasive healthcare problem worldwide due to the increase in the number of cases, multidrug resistance (MDR) and metastasis. As a consequence of multidrug resistance, cancer treatment must rely on a host of chemotherapeutic agents and chemosensitizers to achieve remission. To overcome these problems, a series of biodegradable triblock copolymers of PEG, guanidinium-functionalized polycarbonate and polylactide (PEG-PGCx-PDLAy) is designed as chemotherapeutic agents. These copolymers self-assemble into micellar nanoparticles, and are highly effective against various cancer cell lines including human breast cancer (BCap37), liver cancer (HepG2), lung cancer (A549) and epidermoid carcinoma (A431) cell lines as well as MDR Bats-72 and Bads-200 cancer cells that were developed from BCap37. Multiple treatments with the polymers at sub-lethal doses do not induce resistance. The polymers kill cancer cells by a non-apoptotic mechanism with significant vacuolization and subsequent membrane disruption. In vivo antitumor efficacy is evaluated in a metastatic 4T1 subcutaneous tumor model. Treatment with stereocomplexes of PEG-PGC43-PLLA19 and PEG-PGC43-PDLA20 at a dose of 20 mg/kg of mouse body weight suppresses tumor growth and inhibits tumor metastasis in vivo. These polymers show promise in the treatment of cancer without the onset of resistance.

Development and Validation of a Small Animal Immobilizer and Positioning System for the Study of Delivery of Intracranial and Extracranial Radiotherapy Using the Gamma Knife System.

The purpose of this research is to establish a process of irradiating mice using the Gamma Knife as a versatile system for small animal irradiation and to validate accurate intracranial and extracranial dose delivery using this system. A stereotactic immobilization device was developed for small animals for the Gamma Knife head frame allowing for isocentric dose delivery. Intercranial positional reproducibility of a reference point from a primary reference animal was verified on an additional mouse. Extracranial positional reproducibility of the mouse aorta was verified using 3 mice. Accurate dose delivery was validated using film and thermoluminescent dosimeter measurements with a solid water phantom. Gamma Knife plans were developed to irradiate intracranial and extracranial targets. Mice were irradiated validating successful targeted radiation dose delivery. Intramouse positional variability of the right mandible reference point across 10 micro-computed tomography scans was 0.65 ± 0.48 mm. Intermouse positional reproducibility across 2 mice at the same reference point was 0.76 ± 0.46 mm. The accuracy of dose delivery was 0.67 ± 0.29 mm and 1.01 ± 0.43 mm in the coronal and sagittal planes, respectively. The planned dose delivered to a mouse phantom was 2 Gy at the 50% isodose with a measured thermoluminescent dosimeter dose of 2.9 ± 0.3 Gy. The phosphorylated form of member X of histone family H2A (γH2AX) staining of irradiated mouse brain and mouse aorta demonstrated adjacent tissue sparing. In conclusion, our system for preclinical studies of small animal irradiation using the Gamma Knife is able to accurately deliver intracranial and extracranial targeted focal radiation allowing for preclinical experiments studying focal radiation.

In vivo targeting of adoptively transferred T-cells with antibody- and cytokine-conjugated liposomes.

In adoptive cell therapy (ACT), autologous tumor-specific T-cells isolated from cancer patients are activated and expanded ex vivo, then infused back into the individual to eliminate metastatic tumors. A major limitation of this promising approach is the rapid loss of ACT T-cell effector function in vivo due to the highly immunosuppressive environment in tumors. Protection of T-cells from immunosuppressive signals can be achieved by systemic administration of supporting adjuvant drugs such as interleukins, chemotherapy, and other immunomodulators, but these adjuvant treatments are often accompanied by serious toxicities and may still fail to optimally stimulate lymphocytes in all tumor and lymphoid compartments. Here we propose a novel strategy to repeatedly stimulate or track ACT T-cells, using cytokines or ACT-cell-specific antibodies as ligands to target PEGylated liposomes to transferred T-cells in vivo. Using F(ab')2 fragments against a unique cell surface antigen on ACT cells (Thy1.1) or an engineered interleukin-2 (IL-2) molecule on an Fc framework as targeting ligands, we demonstrate that >95% of ACT cells can be conjugated with liposomes following a single injection in vivo. Further, we show that IL-2-conjugated liposomes both target ACT cells and are capable of inducing repeated waves of ACT T-cell proliferation in tumor-bearing mice. These results demonstrate the feasibility of repeated functional targeting of T-cells in vivo, which will enable delivery of imaging contrast agents, immunomodulators, or chemotherapy agents in adoptive cell therapy regimens.