Plasminogen activator inhibitor-1 promotes immune evasion in tumors by facilitating the expression of programmed cell death-ligand 1.

Background: Increased levels of plasminogen activator inhibitor-1 (PAI-1) in tumors have been found to correlate with poor clinical outcomes in patients with cancer. Although abundant data support the involvement of PAI-1 in cancer progression, whether PAI-1 contributes to tumor immune surveillance remains unclear. The purposes of this study are to determine whether PAI-1 regulates the expression of immune checkpoint molecules to suppresses the immune response to cancer and demonstrate the potential of PAI-1 inhibition for cancer therapy. Methods: The effects of PAI-1 on the expression of the immune checkpoint molecule programmed cell death ligand 1 (PD-L1) were investigated in several human and murine tumor cell lines. In addition, we generated tumor-bearing mice and evaluated the effects of a PAI-1 inhibitor on tumor progression or on the tumor infiltration of cells involved in tumor immunity either alone or in combination with immune checkpoint inhibitors. Results: PAI-1 induces PD-L1 expression through the JAK/STAT signaling pathway in several types of tumor cells and surrounding cells. Blockade of PAI-1 impedes PD-L1 induction in tumor cells, significantly reducing the abundance of immunosuppressive cells at the tumor site and increasing cytotoxic T-cell infiltration, ultimately leading to tumor regression. The anti-tumor effect elicited by the PAI-1 inhibitor is abolished in immunodeficient mice, suggesting that PAI-1 blockade induces tumor regression by stimulating the immune system. Moreover, combining a PAI-1 inhibitor with an immune checkpoint inhibitor significantly increases tumor regression. Conclusions: PAI-1 protects tumors from immune surveillance by increasing PD-L1 expression; hence, therapeutic PAI-1 blockade may prove valuable in treating malignant tumors.

Remediation of cadmium-contaminated soil: GLDA-assisted extraction and sequential FeCl3-CaO-based post-stabilization.

Cadmium (Cd) contamination of farmland soils is a growing concern because of its highly toxic impact on ecosystems and human health. Chelator-assisted washing and chemical immobilization are effective remediation strategies for Cd-contaminated soils. Ethylenediaminetetraacetic acid (EDTA) has traditionally been used for soil washing, but its persistence in the environment and subsequent toxicity have raised significant ecological concerns. Consequently, biodegradable chelators have gained increasing attention as eco-friendly alternatives to the persistent chelator, EDTA. Therefore, this study evaluated the performance and efficacy of three biodegradable chelators: L-glutamate-N,N'-diacetic acid (GLDA), methylglycine-diacetic acid (MGDA), and 3-hydroxy-2,2'-iminodisuccinic acid (HIDS) in comparison to EDTA for remediating a real Cd-contaminated agricultural soil. The influence of treatment parameters, including chelator variants, washing time, chelator concentration, solution pH, and liquid-to-soil ratio (L/S) on Cd extraction was studied and optimized to attain the maximum removal rate. Following chelator-assisted washing, the efficacy of a stabilization preference combining FeCl3 and CaO in reducing the leaching potential of residual Cd in chelator-washed soil residues was also investigated. GLDA demonstrated comparable Cd extraction efficiency to EDTA, and the Cd extraction efficiency was found to be positively correlated with the soil washing parameters. However, under the optimized conditions (chelator concentration: 10 mmol L-1; washing time: 3 h; solution pH: 3; L/S ratio: 10:1), GLDA exhibited a higher Cd extraction rate than EDTA or the other chelators. Furthermore, a post-treatment process incorporating FeCl3 and CaO substantially diminished the water-leachable Cd content in the resultant soil residues. The proposed remediation strategy, which combines chemically assisted washing and stabilization, could be a practical option for extracting bulk Cd from soil and reducing the leaching potential of residual Cd.

Cell-compatible isotonic freezing media enabled by thermo-responsive osmolyte-adsorption/exclusion polymer matrices.

During the long-term storage of cells, it is necessary to inhibit ice crystal formation by adding cryoprotectants. Non-cell-permeable cryoprotectants have high osmotic pressure which dehydrates cells, indirectly suppressing intracellular ice crystal formation. However, the high osmotic pressure and dehydration often damage cells. Emerging polymer-type non-cell-permeable cryoprotectants form matrices surrounding cells. These matrices inhibit the influx of extracellular ice nuclei that trigger intracellular ice crystal formation. However, these polymer-type cryoprotectants also require high osmotic pressure to exert an effective cryoprotecting effect. In this study, we designed a poly(zwitterion) (polyZI) that forms firm matrices around cells based on their high affinity to cell membranes. The polyZI successfully cryopreserved freeze-vulnerable cells under isotonic conditions. These matrices also controlled osmotic pressure by adsorbing and desorbing NaCl depending on the temperature, which is a suitable feature for isotonic cryopreservation. Although cell proliferation was delayed by the cellular matrices, washing with a sucrose solution improved proliferation.

Evaluation of newly designed flushing techniques for on-site remediation of arsenic-contaminated excavated debris.

Excavated debris (soil and rock) contaminated with geogenic arsenic (As) is an increasing concern for regulatory organizations and construction stakeholders. Chelator-assisted soil flushing is a promising method for practical on-site remediation of As-contaminated soil, offering technical, economic, and environmental benefits. Ethylenediaminetetraacetic acid (EDTA) is the most prevalent chelator used for remediating As-contaminated soil. However, the extensive environmental persistence and potential toxicity of EDTA necessitate the exploration of eco-compliant alternatives. In this study, the feasibility of the conventional flushing method pump-and-treat and two newly designed immersion and sprinkling techniques were evaluated at the laboratory scale (small-scale laboratory experiments) for the on-site treatment of As-contaminated excavated debris. Two biodegradable chelators, L-glutamic acid-N,N'-diacetic acid (GLDA) and 3-hydroxy-2,2'-iminodisuccinic acid (HIDS), were examined as eco-friendly substitutes for EDTA. Additionally, this study highlights a useful post-treatment measure to ensure minimal mobility of residual As in the chelator-treated debris residues. The pump-and-treat method displayed rapid As-remediation (t, 3 h), but it required a substantial volume of washing solution (100 mL g-1). Conversely, the immersion technique demonstrated an excellent As-extraction rate using a relatively smaller washing solution (0.33 mL g-1) and shorter immersion time (t, 3 h). In contrast, the sprinkling technique showed an increased As-extraction rate over an extended period (t, 48 h). Among the chelators employed, the biodegradable chelator HIDS (10 mmol L-1; pH, 3) exhibited the highest As-extraction efficiency. Furthermore, the post-treatment of chelator-treated debris with FeCl3 and CaO successfully reduced the leachable As content below the permissible limit.

Anti-PD-1 antibodies recognizing the membrane-proximal region are PD-1 agonists that can down-regulate inflammatory diseases.

The PD-1 receptor triggers a negative immunoregulatory mechanism that prevents overactivation of immune cells and subsequent inflammatory diseases. Because of its biological significance, PD-1 has been a drug target for modulating immune responses. Immunoenhancing anti-PD-1 blocking antibodies have become a widely used cancer treatment; however, little is known about the required characteristics for anti-PD-1 antibodies to be capable of stimulating immunosuppressive activity. Here, we show that PD-1 agonists exist in the group of anti-PD-1 antibodies recognizing the membrane-proximal extracellular region in sharp contrast to the binding of the membrane-distal region by blocking antibodies. This trend was consistent in an analysis of 81 anti-human PD-1 monoclonal antibodies. Because PD-1 agonist antibodies trigger immunosuppressive signaling by cross-linking PD-1 molecules, Fc engineering to enhance FcγRIIB binding of PD-1 agonist antibodies notably improved human T cell inhibition. A PD-1 agonist antibody suppressed inflammation in murine disease models, indicating its clinical potential for treatment of various inflammatory disorders, including autoimmune diseases.

Enhanced remediation of arsenic-contaminated excavated soil using a binary blend of biodegradable surfactant and chelator.

The reclamation of geogenic As-contaminated excavated soils as construction additives can reduce the post-disposal impact on the ecosystem and space. Although retaining soil characteristics while reducing contaminant load is a challenging task, washing remediation with biodegradable surfactants or chelators is a promising alternative to non-biodegradable counterparts. In this study, newly synthesized biodegradable surfactants (SDG: sodium N-dodecanoyl-glycinate, SDBA: sodium N-dodecanoyl-ß-alaninate, SDGBH: sodium N-dodecanoyl-α,γ-glutamyl-bis-hydroxyprolinate, SDT: sodium N-dodecanoyl-taurinate, and DCPC: N-dodecyl-3-carbamoyl-pyridinium-chloride) and biodegradable chelators (EDDS: ethylenediamine N,N'-disuccinic acid, GLDA: L-glutamate-N, N'-diacetic acid, and HIDS: 3-hydroxy-2,2'-imino disuccinic acid) are evaluated for the remediation of As-contaminated soil. The operating variables, such as washing duration, solution pH, and surfactant or chelator concentration, are optimized for maximum As extraction. SDT shows the highest As-extraction efficiency irrespective of solution pH and surfactant variants, while HIDS is the superior chelator under acidic or alkaline conditions. A binary blend of SDT and HIDS is evaluated for As extraction under varying operating conditions. The SDT-HIDS binary blend demonstrates 6.9 and 1.6-times higher As-extraction rates than the SDT and HIDS-only washing, respectively, under acidic conditions. The proposed approach with a binary blend of a biodegradable surfactant and chelator is a green solution for recycling As-contaminated excavated soils for geotechnical applications.

Effect of Hydrophobic Chain Length on the Antioxidation Properties of Alanyl Tyrosine Dipeptide-type Surfactants.

The antioxidant (AOX) activities of alanyl tyrosine dipeptide-type surfactants with several chain lengths were investigated. The critical micelle concentration decreased exponentially with the carbon number of the hydrophobic chain of the surfactant. The antioxidative property was investigated using the 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid ammonium salt) (ABTS) assay and the oxygen radical absorbance capacity (ORAC) assay. The AOX activity was found to be strongly dependent on the chain length in the monomer solution. Therefore, an increase in the molecular size negatively influenced the AOX ability of the tyrosine residue. However, it was almost independent of the chain length of the surfactant in the micellar solution. The micelle particles acted as a catalyst for the reduction of the radicals in the ORAC assay.

Local Cross-Coupling Activity of Azide-Hexa(ethylene glycol)-Terminated Self-Assembled Monolayers Investigated by Atomic Force Microscopy.

Azide-oligo(ethylene glycol)-terminated self-assembled monolayers (N3-OEG-SAMs) are promising interfacial structures for surface functionalization. Its many potential applications include chemical/bio-sensing and construction of surface models owing to its cross-coupling activity that originates from the azide group and oligo(ethylene glycol) (OEG) units for non-specific adsorption resistance. However, there are only a few studies and limited information, particularly on the molecular-scale structures and local cross-coupling activities of N3-OEG-SAMs, which are vital to understanding its surface properties and interfacial molecular design. In this study, molecular-scale surface structures and cross-coupling activity of azide-hexa(ethylene glycol)-terminated SAMs (N3-EG6-SAMs) were investigated using frequency modulation atomic force microscopy (FM-AFM) in liquid. The N3-EG6-SAMs were prepared on Au(111) substrates through the self-assembly of 11-azido-hexa(ethylene glycol)-undecane-1-thiol (N3-EG6-C11-HS) molecules obtained from a liquid phase. Subnanometer-resolution surface structures were visualized in an aqueous solution using a laboratory-built FM-AFM instrument. The results show a well-ordered molecular arrangement in the N3-EG6-SAM and its clean surfaces originating from the adsorption resistance property of the terminal EG6 units. Surface functionalization by the cross-coupling reaction of copper(I)-catalyzed azide-alkyne cycloaddition was observed, indicating a structural change in the form of fluctuating structures and island-shaped structures depending on the concentration of the alkyne molecules. The FM-AFM imaging enabled to provide information on the relationship between the surface structures and cross-coupling activity. These findings provide molecular-scale information on the functionalization of the N3-EG6-SAMs, which is helpful for the interfacial molecular design based on alkanethiol SAMs in many applications.

Selective Separation of Radiocesium from Complex Aqueous Matrices Using Dual Solid-Phase Extraction Systems.

The release of radiocesium (r-Cs) into natural aqueous systems is of concern because of its extended solubility as an alkaline metal ion and its facile incorporation into living beings. A technique for the selective separation of Cs from an aqueous matrix using dual solid-phase extraction (SPE) systems in a series is proposed in this paper. The SPEs equipped with chelates (Nobias Chelate-PA1 and Nobias Chelate-PB1), an ion-exchange resin (Nobias Ion SC-1), or macrocycles (MetaSEP AnaLig Cs-01 and MetaSEP AnaLig Cs-02) were evaluated in terms of selectivity and retention/recovery behavior toward Cs and other potentially competing ions (Li, Na, K, Rb, Ba, Ca, Mg, and Sr). The simulated solution of 133Cs, a chemical analog of r-Cs, was used to optimize the separation process. Operating parameters such as pH (3-13), flow rate (0.2-5.0 mL min-1), and elution behavior (HCl, 0.1-5.0 mol L-1) were optimized to ensure maximum removal of Cs from the aqueous matrices. The dual SPE system comprised Nobias Chelate-PB1 that minimized the competing impact of ions, while selective Cs retention was attained with MetaSEP AnaLig Cs-02. The proposed process was verified using real r-Cs-contaminated water from Fukushima, Japan, to observe the quantitative separation and preconcentration of r-Cs from the complex matrices.

STAT5 interferes with PD-1 transcriptional activation and affects CD8+ T-cell sensitivity to PD-1-dependent immunoregulation.

Programmed cell death-1 (PD-1) is a co-inhibitory receptor that dampens immune responses upon interaction with PD-L1 and PD-L2. Although PD-1 expression on T cells is known to be activation-dependent, how cytokines modify its regulation is not fully resolved. Using polyclonal T-cell activation to study cytokine-dependent PD-1 regulation, we found that IL-2 inhibited transcriptional up-regulation of PD-1 despite the promotion of T-cell activation. The IL-2-mediated reduction in PD-1 expression augmented CD8+ T-cell activities against PD-L1-expressing target cells. To study the mechanism of PD-1 reduction, we focused on STAT5 activation in the IL-2 signaling pathway. Bioinformatic analysis suggested a novel conserved PD-1 promoter domain where NFAT and STAT5 can potentially compete with each other for binding. NFAT1 interaction with this domain revealed substantial potency in PD-1 transcription compared to STAT5A, and STAT5A overexpression could quench NFAT1-dependent PD-1 up-regulation in a sequence-specific manner. Chromatin immunoprecipitation analysis of activated T cells showed that IL-2 treatment significantly diminished the binding of NFAT1 and NFAT2 in the hypothesized competition site, while STAT5 binding to the same region was increased. These results raise the possibility that the competition of transcriptional factors might be involved in the fine-tuning of PD-1 expression by cytokines such as IL-2.

Effect of optokinetic stimulation on weight-bearing shift in standing and sitting positions in stroke patients.

BACKGROUND: Patients with hemiplegia after stroke tend to bear weight on the non-paretic side and exhibit large postural sway during static standing and walking, which may increase their risk of falls. Improvement of the sitting posture balance in the early phase of rehabilitation by adjusting weight-bearing would minimize the risk of falls as early rehabilitation reportedly improves walking ability and prevents falls in later phases of rehabilitation or at discharge. AIM: This study aimed to evaluate the effect of optokinetic stimulation (OKS) on shift of the weight-bearing (displacement of the center of pressure [CoP]) in patients with hemiplegia who are incapable of independent standing. DESIGN: Quasi-experimental, cross-sectional study. SETTING: Rehabilitation hospital. POPULATION: Patients with hemiplegia in the subacute phase after stroke (N.=37). METHODS: Standing and sitting balance tests were performed during OKS projected onto a screen. For OKS, a pattern of random dots was presented, which continuously moved in horizontal or torsional directions during both static standing and sitting conditions. Postural sway was assessed during standing and sitting by measuring the sway path, sway area, sway velocity, and mean displacement of CoP. The magnitude of the lateral change in CoP as an indicator of the weight-bearing shift was evaluated by subtraction of the mean CoP of the right-left axis component in the stationary condition from the mean CoP sway during OKS. RESULTS: OKS induced a unilateral change of the mean CoP position in patients during both, sitting and static standing, indicating that OKS can shift the weight-bearing in patients after stroke, irrespective of the posture condition. Moreover, the same OKS approach evoked an analogous shift in patients with more severe symptoms, with impairment in independent standing. CONCLUSIONS: OKS could induce a significant shift in weight balance in patients with hemiplegia after stroke who are incapable of independent standing, suggesting that the OKS approach can be applied to a broader spectrum of patients, including those with more severe symptoms. CLINICAL REHABILITATION IMPACT: OKS approach would improve exercise training in the early phase of rehabilitation of patients with hemiplegia after stroke.

Inactivation of the PD-1-Dependent Immunoregulation in Mice Exacerbates Contact Hypersensitivity Resembling Immune-Related Adverse Events.

Blockade of PD-1, an indispensable physiological immunoregulatory mechanism, enhances immune activities and is widely used in the immunotherapy of cancer. This treatment often accompanies inflammatory complication called immune-related adverse events (irAE), most frequently in the skin. To analyze how skin inflammation develops by the blockade of PD-1-dependent immunoregulation, we studied the exacerbation of oxazolone-induced contact hypersensitivity by PD-L1 blockade. The inactivation of PD-1 signaling enhanced swelling of the skin with massive CD8+ T cell infiltration. Among PD-1-expressing cells, T cells were the predominant targets of anti-PD-L1 mAb treatment since PD-L1 blockade did not affect skin inflammation in RAG2-/- mice. PD-L1 blockade during immunization with oxazolone significantly promoted the development of hapten-reactive T cells in the draining lymph nodes. The enhancement of local CD8+ T cell-dominant immune responses by PD-L1 blockade was correlated with the upregulation of CXCL9 and CXCL10. Challenges with a low dose of oxazolone did not demonstrate any significant dermatitis; however, the influence of PD-L1 blockade on T cell immunity was strong enough to cause the emergence of notable dermatitis in this suboptimal dosing, suggesting its relevance to dermal irAE development. In the low-dose setting, the blockade of CXCR3, receptor of CXCL9/10, prevented the induction of T cell-dominant inflammation by anti-PD-L1 mAb. This experimental approach reproduced CD8+ T cell-dominant form of cutaneous inflammation by the blockade of PD-L1 that has been observed in dermal irAE in human patients.

Enhanced Aggregation of Stimuli Responsive Surfactants by Esterolytic Reactions.

Thioester surfactants, [C12H25N(CH3)2(CH2)mSCOCH3] Br (C12mSAc, m = 4, 11, 12), yielded thiol surfactants via thiol-thioester exchange upon addition of dithiothreitol in aqueous solution. The thiolthioester exchange reaction was enhanced in a micellar system owing to the concentration effect. The enhanced aggregation upon deprotection of the thioacetate group was observed by dynamic light scattering measurements. The thiol surfactants yielded disulfide-linked gemini surfactants upon air oxidation and incubation with hydrogen peroxide. In contrast, the thiol surfactants yielded thioester double-tailed products by esterolytic reactions with p-nitrophenyl hexanoate. The introduction of an alkyl chain to the second side chain significantly decreased the critical micelle concentration and induced the micellar growth.

Differing Effect of the Sodium-Glucose Cotransporter 2 Inhibitor Ipragliflozin on the Decrease of Fat Mass vs. Lean Mass in Patients With or Without Metformin Therapy.

BACKGROUND: We previously reported changes of body composition determined by dual-energy X-ray absorptiometry after treatment with ipragliflozin, a sodium-glucose cotransporter 2 (SGLT2) inhibitor. In that study, mean body weight was decreased by 3.5 kg (4.3% of the baseline value) after ipragliflozin treatment at 50 mg/day, with fat mass and lean mass showing similar reductions of 1.7 and 1.8 kg, respectively. A long-term decrease of lean mass in patients treated with SGLT2 inhibitors may be associated with loss of skeletal muscle, which could potentially have an impact on quality of life. METHODS: In this post hoc analysis, we investigated whether changes of body composition were influenced by other medications for diabetes in 20 patients (11 men and nine women) who received ipragliflozin for 24 weeks. RESULTS: When we divided the patients into two subgroups with or without metformin treatment, fat mass showed a significant decrease in the ipragliflozin + metformin subgroup and a significantly greater decrease compared to the ipragliflozin subgroup (2.0 kg; 95% confidence interval (CI): 0.1 - 3.9; P = 0.038). Lean mass was significantly decreased in the ipragliflozin subgroup, but the decrease showed no significant difference from that in the ipragliflozin + metformin subgroup (1.9 kg; 95% CI: -4.1 - 0.3; P = 0.087). No significant differences of body composition changes were observed with other antidiabetic agents. CONCLUSIONS: More desirable weight reduction due to preferential fat loss and less muscle loss may be achieved by combining an SGLT2 inhibitor with metformin.

Solubilization of Genistein in Phospholipid Vesicles and Their Atioxidant Capacity.

Water-insoluble genistein was solubilized in aqueous medium by using phospholipid vesicles composed of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dioleoyl-sn-glycerophosphocholine (DOPC) with 0-30% cholesterol. For each vesicle, the maximum solubilization amount of genistein was investigated by X-ray scattering measurement. In addition, the antioxidant capacity of the solubilized genistein was evaluated by the ABTS assay. Genistein was found to be solubilized by 10-20% and 40-50% of the vesicle concentrations of pure DPPC and DOPC respectively. The maximum solubilization amount of genistein decreased to 0-10% and 20-30% when 30% of cholesterol is present in the respective vesicles. Cholesterol is solubilized in a hydrophobic core whereas genistein is solubilized in the polar head region or in the polar-apolar interface. The overlapping of solubilizing sites affected the solubilization of genistein when cholesterol was present in the vesicles. Moreover, the lamellar interval was largely affected by cholesterol in compared to the little impact of genistein because the later can indirectly affect the acyl chains. Genistein solubilized in DOPC showed the same degree of antioxidant capacity as that of vesicle-free genistein system. On the other hand, genistein solubilized in DPPC had lower antioxidant activity than the former systems. The distinction of antioxidant activity at different systems probably related to the difference of accessibility of ABTS radical cation to solubilized genistein through different vesicles. Finally, cholesterol-free DOPC vesicles were found to be the best solubilizer for genistein among the investigated systems.

Germanene Epitaxial Growth by Segregation through Ag(111) Thin Films on Ge(111).

Large-scale two-dimensional sheets of graphene-like germanium, namely, germanene, have been epitaxially prepared on Ag(111) thin films grown on Ge(111), using a segregation method, differing from molecular beam epitaxy used in previous reports. From the scanning tunneling microscopy (STM) images, the surface is completely covered with an atom-thin layer showing a highly ordered long-range superstructure in wide scale. Two types of protrusions, named hexagon and line, form a (7√7 × 7√7) R19.1° supercell with respect to Ag(111), with a very large periodicity of 5.35 nm. Auger electron spectroscopy and high-resolution synchrotron radiation photoemission spectroscopy demonstrate that Ge atoms are segregated on the Ag(111) surface as an overlayer. Low-energy electron diffraction clearly shows incommensurate "(1.35 × 1.35)" R30° spots, corresponding to a lattice constant of 0.39 nm, in perfect accord with close-up STM images, which clearly reveal an internal honeycomb arrangement with corresponding parameter and low buckling within 0.01 nm. As this 0.39 nm value is in good agreement with the theoretical lattice constant of free-standing germanene, conclusively, the segregated Ge atoms with trivalent bonding in honeycomb configuration form a characteristic two-dimensional germanene-like structure.

Aggregation Behavior and Thiol-Thioester Exchange for Cationic Surfactants with Propylthioacetate Side Chain.

A series of cationic surfactants containing the thioacetate group, [CnH2n+1N(CH3)2(CH2)3SCOCH3] Cl (Cn3SAc, n = 12, 14, 16), were prepared and their properties in aqueous solution were investigated by conductivity, fluorescence, and dynamic light scattering measurements. The critical micelle concentrations (CMCs) of Cn3SAc decreased to about half the value of the corresponding alkyltrimethylammonium chloride. Thioacetate was eliminated with the addition of dithiothreitol (DTT) as well as NaOH. HPLC (high performance liquid chromatography) analysis and NMR (nuclear magnetic resonance) spectroscopy showed that thiol surfactants, [CnH2n+1N(CH3)2(CH2)3SH]Cl (Cn3SH), were generated upon the addition of DTT in aqueous solution via thiol-thioester exchange, whereas gemini surfactants, [CnH2n+1N(CH3)2(CH2)3SS(CH2)3N(CH3)2CnH2n+1]2Cl (2Cn3SS), were generated upon incubation in alkaline solution via hydrolysis and air oxidation.

Oxygen-dependent regulation of immune checkpoint mechanisms.

Immunotherapy of cancer has finally materialized following the success of immune checkpoint blockade. Since down-regulation of immune checkpoint mechanisms is beneficial in cancer treatment, it is important to ask why tumors are infamously filled with the immunosuppressive mechanisms. Indeed, immune checkpoints are physiological negative feedback mechanisms of immune activities, and the induction of such mechanisms is important in preventing excessive destruction of inflamed normal tissues. A condition commonly found in tumors and inflamed tissues is tissue hypoxia. Oxygen deprivation under hypoxic conditions by itself is immunosuppressive because proper oxygen supply could support bioenergetic demands of immune cells for optimal immune responses. However, importantly, hypoxia has been found to up-regulate a variety of immune checkpoints and to be able to drive a shift toward a more immunosuppressive environment. Moreover, extracellular adenosine, which accumulates due to tissue hypoxia, also contributes to the up-regulation of other immune checkpoints. Taken together, tissue oxygen is a key regulator of the immune response by directly affecting the energy status of immune effectors and by regulating the intensity of immunoregulatory activity in the environment. The regulators of various immune checkpoint mechanisms may represent the next focus to modulate the intensity of immune responses and to improve cancer immunotherapy.

A2A Adenosine Receptor Gene Deletion or Synthetic A2A Antagonist Liberate Tumor-Reactive CD8+ T Cells from Tumor-Induced Immunosuppression.

Tumor hypoxia-driven accumulation of extracellular adenosine was shown to facilitate tumor evasion by engaging the immunosuppressive, intracellular cAMP-elevating A2 adenosine receptors (A2R) on tumor-reactive effector T cells, but there remains a need for careful evaluation of the limiting factors and properties of A2R blockade-enabled antitumor immunity. In studies of A2AR and/or A2BR gene-deficient mice, we found that A2AR deletion-but not A2BR deletion-liberates endogenous CD8+ T cell antitumor immunity against weakly immunogenic MCA205 sarcomas. Studies of adoptively transferred A2AR-/-, A2BR-/-, or A2AR-/-/A2BR-/- tumor-reactive T cells confirmed that immunosuppression in the tumor microenvironment was mediated by A2AR on CD8+ T cells. Treatment with A2AR antagonist mimicked A2AR gene deletion in adoptive T cell immunotherapy. This therapeutic benefit of targeting A2AR was independent of the anatomical location of tumor growth. The enhanced antitumor reactivity also led to the eradication of established intracranial tumors, which was associated with mouse survival and the maintenance of long-lasting, tumor-specific immunological memory. The blockade of the A2AR on adoptively transferred T cells by synthetic A2AR antagonist led to higher levels of IFN-γ secretion by tumor-infiltrating CD8+ T cells. These data clarify the mechanism of hypoxia-driven immunosuppression in the tumor microenvironment by A2AR on tumor-reactive CD8+ T cells and show that selective A2AR antagonists can be effective in improving the outcomes of T cell-based immunotherapies. Demonstration of the T cell dose dependency of tumor rejection points to a major limitation of current cancer immunotherapies, in which the presence of sufficient numbers of tumor-reactive T cells in a patient is not known.

Aggregation and pH Responsive Behavior of Thioester Surfactants and Formation of Disulfide Linkages in Aqueous Solutions.

pH responsive surfactants, [C12H25N(CH3)2(CH2)nSCOCH3]Br (C12nSAc, n = 4, 11, 12), were prepared, and their properties in aqueous solution were examined. The critical micelle concentration (cmc) and critical vesicle concentration (cvc) were determined based on changes in conductivity, as well as by fluorescence measurements, and light scattering methods. A significant increase in the light scattering intensities of the C12nSAc (n=11, 12) systems suggested that the growth of aggregates was accompanied by considerable counterion binding with increasing surfactant concentration. The diameter of C1211SAc, recorded by the dynamic light scattering measurements, was about 9.6 ±1.0 nm, which was slightly smaller than that for didodecyldimethylammonium bromide (DDAB) vesicles. The thioester group was easily hydrolyzed upon the addition of NaOH, while it was hardly hydrolyzed with the addition of HCl. The time course of alkaline hydrolysis was examined by the conductivity measurements and high-performance liquid chromatography analysis. [C12H25N(CH3)2(CH2)11SS(CH2)11N(CH3)2C12H25]2Br (2C1211SS) was generated in the C1211SAc alkaline solution because of air oxidation. The C1211SAc alkaline solution gradually became an opaque blue color with increasing light scattering at 346 nm, indicating the remarkable growth of vesicles. The chemical structure of 2C1211SS was consistent with that of a disulfide linked double tailed surfactant, similar to DDAB. The disulfide linkage between the double tailed surfactants will contribute to the stabilization and growth of vesicles.