Radioresponsive Delivery of Toll-like Receptor 7/8 Agonist for Tumor Radioimmunotherapy Enabled by Core-Cross-Linked Diselenide Nanoparticles.

The development of a radioresponsive delivery platform has led to an innovative combination radioimmunotherapy strategy for treating tumors. However, controlling the release of immunomodulators by local radiotherapy in vivo remains a significant challenge in order to minimize off-target toxicity, reduce radiation-induced immunosuppression, and maximize synergistic radioimmunotherapy efficacy. In this study, we report the development of core-cross-linked diselenide nanoparticles (dSeNPs) as carriers for radioresponsive delivery of the toll-like receptors 7/8 agonist through systemic administration to achieve combined radioimmunotherapy of tumors. The dSeNPs were fabricated from a ring-opening reaction between 2,2'-diselenidebis(ethylamine) and the ethylene oxide group of an amphiphilic block copolymer. The diselenide bonds were naturally protected in the core of the self-assembled nanostructure, making the dSeNPs extremely stable in the physiological environment. However, they exhibited dose- and time-dependent radiosensitivity, meaning that X-ray irradiation could spatiotemporally control the release of R848 from the dSeNPs. In vivo results showed that local radioresponsive R848 release from dSeNPs greatly improved the synergistic efficacy of combined radioimmunotherapy via the programmed cooperative immune system activation process. This process included macrophage polarization, dendritic cell maturation, and cytotoxic T cell activation. Our findings suggest that core-cross-linked dSeNPs are a promising platform for combined radiotherapy due to their spatiotemporal controllability of radioresponsive drug release.

Neutrophils-mediated bioinspired nanoagents for noninvasive monitoring of inflammatory recruitment dynamics and navigating phototherapy in rheumatoid arthritis.

Neutrophils play a crucial role in inflammatory immune responses, but their in vivo homing to inflammatory lesions remains unclear, hampering precise treatment options. In this study, we employed a biomineralization-inspired multimodal nanoagent to label neutrophils, enabling noninvasive monitoring of the dynamic process of inflammatory recruitment and guiding photothermal therapy in rheumatoid arthritis. Our nanoagents allowed visualization of neutrophil fate through magnetic resonance imaging, photoacoustic imaging, and fluorescence imaging in the first and second near-infrared windows. Histopathology and immunofluorescence analysis revealed pronounced inflammatory cell infiltration in rheumatoid arthritis compared to the normal limb. Furthermore, the recruitment quantity of neutrophils positively correlated with the inflammatory stage. Additionally, the inherent photothermal effect of the nanoagents efficiently ablated inflammatory cells during the optimal homing time and inflammatory phase. This neutrophil imaging-guided photothermal therapy precisely targeted inflammatory nuclei in rheumatoid arthritis and downregulated pro-inflammatory cytokines in serum. These results demonstrate that in vivo tracking of inflammatory immune response cells can significantly optimize the treatment of inflammatory diseases, including rheumatoid arthritis.

Clinical effects of homodigital dorsal branch of the proper digital artery flap on repairing soft tissue defects of fingers.

Homodigital dorsal branch of proper digital artery flaps (HDBPDAF) have been proved to be an excellent alternative for repairing distal soft tissue defects of fingertip. This study was to evaluate the clinical effects of HDBPDAF on repairing various soft tissue defects of fingers, including thumb and multi-fingers defects. A retrospective study was conducted in 40 patients with 44 finger defects treated with HDBPDAF from August of 2014 to December of 2021. The defects were located on fingertip and finger pulp (n = 28), finger pulp (n = 10), and dorsum of fingers (n = 6) with bone, tendon or nerve exposed. The average size of the flaps was 1.9 × 3.9 cm. Semmes Weinstein monofilament (SWM) test, Static two-point discrimination (2-PD), Total active motion (TAM) scores, the mean Disabilities of the Arm, Shoulder, and Hand (DASH) score were evaluated through long-term follow-up. Forty-two flaps survived completely and uneventfully. Partial flap necrosis was observed in two flaps because of the absence of dorsal branch of proper digital artery. No visible scar contracture and joint limitation were noticed. The mean SWM score of flaps was 4.11 ± 0.4 g. The average 2-PD of the flaps was 8.9 ± 0.9 mm. The mean TAM of injured fingers was 268.7 ± 5.2° (contralateral side: 283.2 ± 6.4°, p < 0.05). The mean DASH score value was 29.7 ± 7.9. The HDBPDAF was an optimal and reliable alternative to repair various distal soft tissue defects of fingers, despite a lower absent rate of dorsal branch.

Dimethyl Fumarate Combined With Vemurafenib Enhances Anti-Melanoma Efficacy via Inhibiting the Hippo/YAP, NRF2-ARE, and AKT/mTOR/ERK Pathways in A375 Melanoma Cells.

Melanoma is a deadly form of skin cancer with high rates of resistance to traditional chemotherapy and radiotherapy. BRAF inhibitors (BRAFi) can achieve initial efficacy when used to treat melanoma patients, but drug resistance and relapse are common, emphasizing the need for new therapeutic strategies. Herein, we reported that combination of dimethyl fumarate (DMF) and vemurafenib (Vem) inhibited melanoma cell proliferation more significantly and induced more cell death than single agent did both in vitro and in vivo. DMF/Vem treatment induced cell death through inhibiting the expression and transcriptional activity of NRF2 thereby resulting in more reactive oxygen species (ROS) and via inhibiting the expression of YAP, a key downstream effector of Hippo pathway. DMF/Vem treatment also reduced phosphorylation of AKT, 4EBP1, P70S6K and ERK in AKT/mTOR/ERK signaling pathways. RNA-seq analysis revealed that DMF/Vem treatment specifically suppressed 4561 genes which belong to dozens of cell signaling pathways. These results indicated that DMF/Vem treatment manifested an enhanced antitumor efficacy through inhibiting multiple cell signaling pathways, and thus would be a novel promising therapeutic approach targeted for melanoma.

Isoform-specific and cell/tissue-dependent effects of p38 MAPKs in regulating inflammation and inflammation-associated oncogenesis.

p38 MAPK (mitogen-activated protein kinases) family proteins (α, ß, γ and δ) are key inflammatory kinases and play an important role in relaying and processing intrinsic and extrinsic signals in response to inflammation, stress, and oncogene to regulate cell growth, cell death and cell transformation. Recent studies in genetic mouse models revealed that p38α in epithelial cells mostly suppresses whereas in immune cells it promotes inflammation and inflammation-associated oncogenesis. On the contrary, p38γ and p38δ signaling in immune and epithelial cells is both pro-inflammatory and oncogenic. This review summarizes recent discoveries in this field, discusses possible associated mechanisms, and highlights potentials of systemically targeting isoform-specific p38 MAPKs. Understanding of p38 MAPK isoform-specific and cell/tissue- and perhaps stage-dependent effects and their integrated regulated activity in inflammation and in inflammation-associated oncogenesis is essential for effectively targeting this group of kinases for therapeutic intervention.

Robust Nanofiber Mats Exfoliated From Tussah Silk for Potential Biomedical Applications.

Nanofibers as elements for bioscaffolds are pushing the development of tissue engineering. In this study, tussah silk was mechanically disintegrated into nanofibers dispersed in aqueous solution which was cast to generate tussah silk fibroin (TSF) nanofiber mats. The effect of treatment time on the morphology, structure, and mechanical properties of nanofiber mats was examined. SEM indicated decreasing diameter of the nanofiber with shearing time, and the diameter of the nanofiber was 139.7 nm after 30 min treatment. These nanofiber mats exhibited excellent mechanical properties; the breaking strength increased from 26.31 to 72.68 MPa with the decrease of fiber diameter from 196.5 to 139.7 nm. The particulate debris was observed on protease XIV degraded nanofiber mats, and the weight loss was greater than 10% after 30 days in vitro degradation. The cell compatibility experiment confirmed adhesion and spreading of NIH-3T3 cells and enhanced cell proliferation on TSF nanofiber mats compared to that on Bombyx mori silk nanofiber mats. In conclusion, results indicate that TSF nanofiber mats prepared in this study are mechanically robust, slow biodegradable, and biocompatible materials, and have promising application in regenerative medicine.

Implantation of nanofibrous silk scaffolds seeded with bone marrow stromal cells promotes spinal cord regeneration (6686 words).

Spinal cord injury (SCI) is a common pathology often resulting in permanent loss of sensory, motor, and autonomic function. Numerous studies in which stem cells have been transplanted in biomaterial scaffolds into animals have demonstrated their considerable potential for recovery from SCI. In the present study, a three-dimensional porous silk fibroin (SF) scaffold with a mean pore size of approximately 383 µm and nanofibrous structure was fabricated, the silk scaffold enabling the enhanced attachment and proliferation of bone marrow stromal cells (BMSCs). Investigation of its therapeutic potential was conducted by implantation of the nanofibrous SF scaffold seeded with BMSCs into a transected spinal cord model. Recovery of the damaged spinal cord was significantly improved after 2 months, compared with a non-nanofibrous scaffold, in combination with decreased glial fibrillary acidic protein (GFAP) expression and improved axonal regeneration at the site of injury. Furthermore, elevated Basso-Beattie-Bresnahan (BBB) scores indicated greatly improved hindlimb movement. Together, these results demonstrate that transplantation of neural scaffolds consisting of nanofibrous SF and BMSCs is an attractive strategy for the promotion of functional recovery following SCI.

Facile preparation of a strong chitosan-silk biocomposite film.

Chitosan-silk biocomposite films with nanofibrous structures have been prepared by facile solution casting of chitosan and silk co-dissolved in formic acid. The morphology, structure and mechanical properties of the chitosan-silk biocomposite were characterized by SEM, FTIR, TG-DSC, and mechanical testing. The results demonstrate that the prepared biocomposite films with a chitosan-silk ratio of 3:1 shows a high tensile strength of 97.8 MPa, a strain at break of 10.8% and a Young's modulus of 3.5 GPa, indicating its high strength and elasticity. Also, the preliminary cell culture experiment demonstrated the ideal biocompatibility of chitosan-silk composite films. As a result the superior mechanical properties of this composite film can be attributed to the silk nanofibrils and chitosan self-assembled nanofibers, and the strong hydrogen bonding interaction between the silk nanofibril and chitosan nanofibers. The specific nanostructure, enhanced mechanical properties, and biocompatibility make the biocomposite films a promising material for applications in biomedical devices.

Conservative tibiotalocalcaneal fusion for partial talar avascular necrosis in conjunction with ankle and subtalar joint osteoarthritis in Kashin-Beck disease: A case report.

RATIONALE: Kashin-Beck disease (KBD) is known for some typical characters like finger joint enlargement, shortened fingers, and dwarfism. However, Avascular necrosis (AVN) of the talus in KBD has rarely been reported in the literature. Here, we reported on a KBD patient presented with partial AVN of the talus in conjunction with ankle and subtalar arthritis. PATIENT CONCERNS: A 50-year-old woman presented with severe pain and limited range of motion in her left ankle and subtalar joint while walking for 2 years. She had been walking with the aid of crutches for many years. Conservative treatment with rigid orthosis and activity restriction could not help reduce the pain in the left foot. DIAGNOSES: Radiographs demonstrated that partial AVN was developed in the body of the talus and arthritis was viewed in the left ankle and subtalar joint. Hence, we established the diagnosis of partial talar AVN in conjunction with ankle and subtalar arthritis. INTERVENTIONS: A conservative tibiotalocalcaneal fusion attempting to preserve as much viable talar body as possible was performed using a humeral locking plate and 2 cannulated compression screws. OUTCOMES: Bone union proved by CT scan and a good alignment of the left limb were achieved at 4-month follow-up postoperatively. LESSONS: Partial AVN of the talus along with ankle and subtalar arthritis in KBD patients has rarely been reported as it is not a common characteristic of KBD in clinical practice. Conservative tibiotalocalcaneal fusion could help preserving much more viable talar body, maintaining most structural integrity of the ankle joint, and achieving a stable and plantigrade foot postoperatively.

Gentamicin-loaded silk/nanosilver composite scaffolds for MRSA-induced chronic osteomyelitis.

Methicillin-resistant Staphylococcus aureus (MRSA) often induces chronic osteomyelitis and then bone defects. Here, gentamicin-loaded silk/nanosilver composite scaffolds were developed to treat MRSA-induced chronic osteomyelitis. AgNO3 was reduced with silk as a reducing agent in formic acid, forming silver nanoparticles in situ that were distributed uniformly in the composite scaffolds. Superior antibacterial properties against MRSA were achieved for the composite scaffolds, without the compromise of osteogenesis capacity. Then gentamicin was loaded on the scaffolds for better treatment of osteomyelitis. In vivo results showed effective inhibition of the growth of MRSA bacteria, confirming the promising future in the treatment of chronic osteomyelitis.

Effective cancer immunotherapy by Ganoderma lucidum polysaccharide-gold nanocomposites through dendritic cell activation and memory T cell response.

Polysaccharides purified from natural herbs possess immunoregulatory functions, while the efficacy of natural polysaccharides on cancer treatment remains unreliable, likely due to their low prescribed doses and fast clearances in clinical settings. In this study, gold nanocomposites containing Ganoderma lucidum polysaccharide (GLP-Au) efficiently induced dendritic cell (DC) activation, evident by the increase of CD80/CD86/CD40/MHCII, decrease of phagocytic ability and acid phosphatase activity, and increased cytokine transcription. GLP-Au significantly promoted the proliferation of CD4+ and CD8+ T cells in splenocytes. DC/T cell co-culture study proved that GLP-Au activation on DC directly resulted in T cell proliferation. GLP-Au exhibited strong inhibitory effects on 4T1 tumor growth and pulmonary metastasis when combined with doxorubicin. GLP-Au recovered body weight loss by doxorubicin and increased the percentage of CD4+/CD44+ memory T cells. This work suggests that polysaccharides from natural herbs can be incorporated into nanocomposites with immunoregulatory characteristics for enhanced efficacy on tumor therapy.

Reliability of distal tibio-fibular syndesmotic instability measurements using weightbearing and non-weightbearing cone-beam CT.

BACKGROUND: To investigate the reliability and reproducibility of syndesmosis measurements on weightbearing (WB) cone-beam computed tomography (CBCT) images and compare them with measurements obtained using non-weightbearing (NWB) images. METHODS: In this IRB-approved, retrospective study of 5 men and 9 women with prior ankle injuries, simultaneous WB and NWB CBCT scans were taken. A set of 21 syndesmosis measurements using WB and NWB images were performed by 3 independent observers. Pearson/Spearman correlation and intra-class correlation (ICC) were used to assess intra- and inter-observer reliability, respectively. RESULTS: We observed substantial to perfect intra-observer reliability (ICC=0.72-0.99) in 20 measurements. Moderate to perfect agreement (ICC=0.45-0.97) between observers was noted in 19 measurements. CONCLUSION: Measurements evaluating the distance between tibia and fibula in the axial plane 10mm above the plafond had high intra- and inter-observer reliability. Mean posterior tibio-fibular distance, diastasis, and angular measurement were significantly different between WB and NWB images.

Vaccination With Mitoxantrone-Treated Primary Colon Cancer Cells Enhances Tumor-Infiltrating Lymphocytes and Clinical Responses in Colorectal Liver Metastases.

BACKGROUND: Colorectal cancer remains a leading cause of cancer-related mortality worldwide. Metastases to the liver are often present at initial presentation and will form in most patients during their course of disease. We have previously demonstrated that enhanced trafficking and activation of tumor-infiltrating lymphocytes in colorectal liver metastases (CRLM) may improve antitumor immune responses. Thus, development of novel mechanisms to increase lymphocyte infiltration and activation are needed to improve patient outcomes. METHODS: CT26 murine colorectal cancer cells were treated with physiologic levels of the potent inducer of immunogenic cell death mitoxantrone (MTX). An in situ vaccine was created with treated cells in an established model of CRLM. Cells were evaluated by flow cytometry for cell cycle evaluation and calreticulin expression. Splenic and tumor-infiltrating lymphocytes were isolated for phenotypic studies. RESULTS: MTX-treatment of colon cancer cells resulted in a sub-G1 peak, inhibition of G1 cell cycle progression, and increased G2/M cell fractions while simultaneously increasing dynamic exposure of calreticulin on the cell surface (P < 0.05). Vaccination with MTX-treated cells resulted in significant decreases in CRLM formation associated with increased tumor-infiltrating leukocytes that displayed increased expression of the T cell surface activation marker CD69. CONCLUSIONS: Vaccination with MTX-treated primary colon cancer cells enhances tumor-infiltrating lymphocytes and clinical responses in CRLM.

Recombinant Domain V of beta (2)-Glycoprotein I Inhibits OxLDL-induced TF Expression in Macrophages.

OBJECTIVE: To investigate if lectin-like oxidised low density lipoprotein receptor is implicated in oxidised low density lipoprotein induced up regulation of tissue factor and whether recombinant domain V of beta (2)-Glycoprotein I expressed in Pichia pastoris inhibits the binding of oxidised and lectin-like low density lipoprotein. METHODS: The expression of tissue factor and lectin-like oxidised low density lipoprotein receptor was detected using Western blot methods. Small interference ribonucleic acid of lectin-like oxidised low density lipoprotein receptor was used to block lectin-like oxidised low density lipoprotein receptor expression. Flow cytometry was used to test the effect of beta (2)-Glycoprotein I expressed in Pichia pastoris on the binding of oxidised low density lipoprotein with lectin-like oxidised low density lipoprotein receptor by using the lectin-like oxidised low density lipoprotein receptor-expressing 293T cells. RESULTS: Oxidised low density lipoprotein at 5-10 g/mL increased tissue factor and lectin-like oxidised low density lipoprotein receptor expression, whereas 20-50 g/mL oxidised low density lipoprotein attenuated tissue factor expression. Inhibiting lectin-like oxidised low density lipoprotein receptor expression by small interference ribonucleic acid of lectin-like oxidised low density lipoprotein receptor impaired oxidised low density lipoprotein-induced tissue factor over expression in macrophages. Pretreatment with beta (2)-Glycoprotein I expressed in Pichia pastoris led to a strong inhibition of tissue factor and lectin-like oxidised low density lipoprotein receptor expression in a dose-dependent manner in macrophages. Flow cytometry analysis showed that beta (2)-Glycoprotein I expressed in Pichia pastoris attenuated the interaction of oxidised low density lipoprotein with lectin-like oxidised low density lipoprotein receptor in lectin-like oxidised low density lipoprotein receptor-expressing 293T cells. CONCLUSIONS: Lectin-like oxidised low density lipoprotein receptor was implicated in the expression of tissue factor induced by oxidised low density lipoprotein, and beta (2)-Glycoprotein I expressed in Pichia pastoris inhibited oxidised low density lipoprotein-induced tissue factor and lectin-like oxidised low density lipoprotein receptor expression, at least in part, via inhibition of the interaction between oxidised low density lipoprotein and lectin-like oxidised low density lipoprotein receptor.

The ω-carboxyl group of 7-ketocholesteryl-9-carboxynonanoate mediates the binding of oxLDL to CD36 receptor and enhances caveolin-1 expression in macrophages.

CD36 signal transduction modulates the uptake of oxidized low-density lipoprotein (oxLDL) and foam cell formation. We previously observed that 7-ketocholesteryl-9-carboxynonanoate (oxLig-1), the lipid moiety of oxLDL, activates the CD36-Src-JNK/ERK1/2 signalling pathway. In this study, we assessed the role of the ω-carboxyl group in the binding of oxLig-1 to CD36 and investigated whether the binding of the ω-carboxyl group to CD36 triggers CD36-mediated signalling, thereby resulting in the upregulation of caveolin-1 expression. Our results showed that oxLig-1 bound to CD36 and that the ω-carboxyl group was critical for this binding. Furthermore, immunoprecipitation and Western blot analyses showed that interaction between the ω-carboxyl group of oxLig-1 and CD36 triggered intracellular Src-JNK/ERK1/2 signal transduction. Moreover, the binding of the ω-carboxyl group to CD36 induced caveolin-1 expression and translocation to the membrane in macrophages. Additionally, inhibitors of Src, JNK and ERK and siRNA targeting CD36 and NF-κB significantly suppressed the enhanced caveolin-1 expression induced by oxLig-1. In conclusion, these observations suggest that oxLig-1 is a critical epitope of oxLDL that mediates the binding of oxLDL to CD36 and activates downstream Src-JNK/ERK1/2-NF-κB signal transduction, resulting in upregulation of caveolin-1 expression in macrophages.

The lipid moiety 7-ketocholesteryl-9-carboxynonanoate mediates binding interaction of oxLDL to LOX-1 and upregulates ABCA1 expression through PPARγ.

AIM: Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1), a specific membrane receptor for oxidized low-density lipoprotein (oxLDL), plays a crucial role in atherosclerosis progression. The aim of this study was to elucidate the role of 7-ketocholesteryl-9-carboxynonanoate (oxLig-1), a lipid component of oxLDL, in the binding of oxLDL to LOX-1 and to determine whether oxLig-1 binding to LOX-1 is involved in the upregulation of ABCA1 expression. MAIN METHODS: OxLig-1 binding to LOX-1 was analysed by AutoDock 4.2.6 and confirmed by fluorescence immunocytochemistry and enzyme-linked immunosorbent assays (ELISAs). LOX-1, LXRα and ABCA1 protein expression induced by oxLig-1 or methylated oxLig-1 was measured by western blotting. In addition, PPARγ activation was investigated using a dual-luciferase reporter system. Furthermore, the signalling cascade involved in oxLig-1-induced ABCA1 expression was assessed using inhibitors for PPARγ and LXRα and specific siRNAs against LOX-1, PPARγ and LXRα. KEY FINDINGS: Docking, fluorescence immunocytochemistry and ELISA analyses showed that oxLig-1 bound LOX-1 and that the ω-carboxyl group was critical for this binding. Moreover, oxLig-1, but not methylated oxLig-1, increased LOX-1, LXRα, and ABCA1 expression. Luciferase reporter assays indicated that oxLig-1 activated PPARγ in the presence of LOX-1. Additionally, the inhibitor and siRNA experiments showed that oxLig-1 triggered the PPARγ-LXRα signalling pathway, leading to upregulation of ABCA1, and that this process required the participation of LOX-1. SIGNIFICANCE: Our observations indicate that oxLig-1 is a critical epitope of oxLDL that mediates the binding of oxLDL to LOX-1 and initiates PPARγ signal transduction to upregulate the expression of ABCA1.

LIGHT Elevation Enhances Immune Eradication of Colon Cancer Metastases.

The majority of patients with colon cancer will develop advanced disease, with the liver being the most common site of metastatic disease. Patients with increased numbers of tumor-infiltrating lymphocytes in primary colon tumors and liver metastases have improved outcomes. However, the molecular factors that could empower antitumor immune responses in this setting remain to be elucidated. We reported that the immunostimulatory cytokine LIGHT (TNFSF14) in the microenvironment of colon cancer metastases associates with improved patient survival, and here we demonstrate in an immunocompetent murine model that colon tumors expressing LIGHT stimulate lymphocyte proliferation and tumor cell-specific antitumor immune responses. In this model, increasing LIGHT expression in the microenvironment of either primary tumors or liver metastases triggered regression of established tumors and slowed the growth of liver metastases, driven by cytotoxic T-lymphocyte-mediated antitumor immunity. These responses corresponded with significant increases in tumor-infiltrating lymphocytes and increased expression of lymphocyte-homing signals in the metastatic tumors. Furthermore, we demonstrated evidence of durable tumor-specific antitumor immunity. In conclusion, increasing LIGHT expression increased T-cell proliferation, activation, and infiltration, resulting in enhanced tumor-specific immune-mediated tumor regressions in primary tumors and colorectal liver metastases. Mechanisms to increase LIGHT in the colon cancer microenvironment warrant further investigation and hold promise as an immunotherapeutic strategy. Cancer Res; 77(8); 1880-91. ©2017 AACR.

Colon cancer cell treatment with rose bengal generates a protective immune response via immunogenic cell death.

Immunotherapeutic approaches to manage patients with advanced gastrointestinal malignancies are desired; however, mechanisms to incite tumor-specific immune responses remain to be elucidated. Rose bengal (RB) is toxic at low concentrations to malignant cells and may induce damage-associated molecular patterns; therefore, we investigated its potential as an immunomodulator in colon cancer. Murine and human colon cancer lines were treated with RB (10% in saline/PV-10) for cell cycle, cell death, and apoptosis assays. Damage-associated molecular patterns were assessed with western blot, ELISA, and flow cytometry. In an immunocompetent murine model of colon cancer, we demonstrate that tumors regress upon RB treatment, and that RB induces cell death in colon cancer cells through G2/M growth arrest and predominantly necrosis. RB-treated colon cancer cells expressed distinct hallmarks of immunogenic cell death (ICD), including enhanced expression of calreticulin and heat-shock protein 90 on the cell surface, a decrease in intracellular ATP, and the release of HMGB1. To confirm the ICD phenotype, we vaccinated immunocompetent animals with syngeneic colon cancer cells treated with RB. RB-treated tumors served as a vaccine against subsequent challenge with the same CT26 colon cancer tumor cells, and vaccination with in vitro RB-treated cells resulted in slower tumor growth following inoculation with colon cancer cells, but not with syngeneic non-CT26 cancer cells, suggesting a specific antitumor immune response. In conclusion, RB serves as an inducer of ICD that contributes to enhanced specific antitumor immunity in colorectal cancer.

Targeting the metabolic pathway of human colon cancer overcomes resistance to TRAIL-induced apoptosis.

Colon cancer is a leading cause of cancer-related mortality for which targeted therapy is needed; however, trials using apoptosis-inducing ligand monotherapy to overcome resistance to apoptosis have not shown clinical responses. Since colon cancer cells selectively uptake and rapidly metabolize glucose, a property utilized for clinical staging, we investigated mechanisms to alter glucose metabolism in order to selectively target the cancer cells and to overcome evasion of apoptosis. We demonstrate TRAIL (tumor necrosis factor-related apoptosis-inducing ligand) resistance in the majority of human colon cancers tested and utilize the glucose analog 2-deoxy-d-glucose to sensitize TRAIL-resistant gastrointestinal adenocarcinoma cells, and not normal gastrointestinal epithelial cells, to TRAIL-induced apoptosis through enhanced death receptor 5 expression, downstream modulation of MAPK signaling and subsequent miRNA expression modulation by increasing the expression of miR-494 via MEK activation. Further, established human colon cancer xenografts treated with this strategy experience anti-tumor responses. These findings in colon adenocarcinoma support further investigation of manipulation of cellular energetics to selectively overcome resistance to apoptosis and to impart tumor regressions in established colon cancer tumors.

Insulin-like growth factor-1 receptor-mediated inhibition of A-type K(+) current induces sensory neuronal hyperexcitability through the phosphatidylinositol 3-kinase and extracellular signal-regulated kinase 1/2 pathways, independently of Akt.

Although IGF-1 has been implicated in mediating hypersensitivity to pain, the underlying mechanisms remain unclear. We identified a novel functional of the IGF-1 receptor (IGF-1R) in regulating A-type K(+) currents (IA) as well as membrane excitability in small trigeminal ganglion neurons. Our results showed that IGF-1 reversibly decreased IA, whereas the sustained delayed rectifier K(+) current was unaffected. This IGF-1-induced IA decrease was associated with a hyperpolarizing shift in the voltage dependence of inactivation and was blocked by the IGF-1R antagonist PQ-401; an insulin receptor tyrosine kinase inhibitor had no such effect. An small interfering RNA targeting the IGF-1R, or pretreatment of neurons with specific phosphatidylinositol 3-kinase (PI3K) inhibitors abolished the IGF-1-induced IA decrease. Surprisingly, IGF-1-induced effects on IA were not regulated by Akt, a common downstream target of PI3K. The MAPK/ERK kinase inhibitor U0126, but not its inactive analog U0124, as well as the c-Raf-specific inhibitor GW5074, blocked the IGF-1-induced IA response. Analysis of phospho-ERK (p-ERK) showed that IGF-1 significantly activated ERK1/2 whereas p-JNK and p-p38 were unaffected. Moreover, the IGF-1-induced p-ERK1/2 increase was attenuated by PI3K and c-Raf inhibition, but not by Akt blockade. Functionally, we observed a significantly increased action potential firing rate induced by IGF-1; pretreatment with 4-aminopyridine abolished this effect. Taken together, our results indicate that IGF-1 attenuates IA through sequential activation of the PI3K- and c-Raf-dependent ERK1/2 signaling cascade. This occurred via the activation of IGF-1R and might contribute to neuronal hyperexcitability in small trigeminal ganglion neurons.