Effects In Tumor-bearing Mice Research Articles

Membrane-destabilizing ionizable lipid empowered imaging-guided siRNA delivery and cancer treatment.

One of the imperative medical requirements for cancer treatment is how to establish an imaging-guided nanocarrier that combines therapeutic and imaging agents into one system. siRNA therapeutics have shown promising prospects in controlling life-threatening diseases. However, it is still challenging to develop siRNA formulations with excellent cellular entry capability, efficient endosomal escape, and simultaneous visualization. Herein, we fabricated multifunctional ionizable lipid nanoparticles (iLNPs) for targeted delivery of siRNA and MRI contrast agent. The iLNPs comprises DSPC, cholesterol, PEGylated lipid, contrast agent DTPA-BSA (Gd), and ionizable lipid termed iBL0104. siRNA-loaded iLNPs (iLNPs/siRNA) could be decorated with a tumor targeting cyclic peptide (c(GRGDSPKC)) (termed GARP), or without targeting modification (termed GAP). Data revealed that GARP/siRNA iLNPs exhibited significantly higher cellular entry efficiency than GAP/siRNA iLNPs. GARP/siRNA iLNPs rapidly and effectively escaped from endosome and lysosome after internalization. Compared with GAP/siPLK1, GARP/siPLK1 exhibited better tumor inhibition efficacy in both cell-line derived xenograft and liver cancer patient derived xenograft murine models. In addition, GARP formulation displayed ideal MRI effect in tumor-bearing mice, and was well tolerated by testing animals. Therefore, this study provides an excellent example for achieving imaging-guided and tumor-targeted siRNA delivery and cancer treatment, highlighting its promising potential for translational medicine application.

Effective Antitumor of Orally Intestinal Targeting Penetrating Peptide-Loaded Tyroserleutide/PLGA Nanoparticles in Hepatocellular Carcinoma.

PurposeHepatocellular carcinoma (HCC) is a common malignant tumor that seriously threatens human life and health. Currently, the majority of antitumor drugs are administered in an injectable manner, which can cause pain and side effects to patients. Objective of this study is to establish an effective oral drug delivery system for anti hepatoma drugs.MethodsIn this study, intestinal targeting cell penetrating peptide (R6LRVG) was obtained by binding cell penetrating peptide (R6) with the polypeptide of LRVG (targeting intestinal epithelial cells). Next, R6LRVG-modified tyroserleutide-poly(lactic-co-glycolic acid) (PLGA) nanoparticles (YSL-PLGA/R6LRVG NPs) were prepared. After that, the nanoparticles were characterized and their stability was evaluated. The cellular uptake, in vitro bioactivity and in vivo antitumor activity of the nanoparticles were investigated. In addition, the mechanism, including the endocytic pathway and respiratory rate detection of mitochondria, was further investigated.ResultsYSL-PLGA/R6LRVG NPs were successfully prepared. Characterization revealed YSL-PLGA/R6LRVG NPs to be globular particles with smooth surfaces and an average diameter of 222.6 nm. The entrapment efficiency and drug loading of tyroserleutide were 70.27% and 19.69%, respectively. Furthermore, the YSL-PLGA/R6LRVG NPs group exhibited the largest amount of YSL uptake. We also found that cell uptake of YSL-PLGA/R6LRVG NPs could be related to the endocytosis pathways mediated by reticulin and caveolae/lipid rafts. Additionally, the YSL-PLGA/R6LRVG NPs could interfere with mitochondrial function. In vivo experiments revealed that orally administered YSL-PLGA/R6LRVG NPs exerted excellent anticancer effects in tumor-bearing mice. Hematoxylin-eosin staining did not show any histological changes in the major organs.ConclusionTo summarize, YSL-PLGA/R6LRVG NPs could be a useful oral delivery system of YSL and may provide a new platform for the oral delivery of anticancer drugs.

Lappaconitine hydrochloride inhibits proliferation and induces apoptosis in human colon cancer HCT-116 cells via mitochondrial and MAPK pathway

Lappaconitine hydrochloride (LH), as a new synthetic alkaloid, exhibits antitumor activity, whereas its antitumor effect on colorectal cancer (CRC) has not been investigated. In this study, the effect of LH on HCT-116 cell proliferation and apoptosis in vivo and in vitro and underlying molecular mechanism were explored. The Cell Counting Kit-8 (CCK-8) was used to assess cell viability. Morphological change was observed by Hoechst 33342 staining. Cell cycle and apoptosis were performed using a flow cytometer. The western blot method was used to screen for related protein expression. The mitochondrial membrane potential (MMP) was confirmed using the 5, 5, 6, 6′-tetrachloro-1, 1′, 3, 3′-tetraethylbenzimi-dazolyl carbo cyanine iodide (JC-1) staining assay. Reactive oxygen species (ROS) was evaluated by a 20−70-dichlorofluorescein diacetate (DCFH-DA) staining assay. The antitumor effect was evaluated in vivo by the xenograft HCT-116 model. The results showed that LH significantly inhibited cell viability in a time- and concentration-dependent manner. LH induced apoptosis and S phase cell cycle arrest. LH promoted the reduction of MMP and ROS accumulation. Moreover, LH activated the mitochondrial and MAPK pathway. The experiments in vivo showed that LH had significant antitumor effect in tumor-bearing mice, and had virtually no effect on the weight and internal organs of the mice. In conclusion, LH could induce apoptosis in HCT-116 cells through mitochondrial and MAPK signaling pathways. LH may be a promising treatment for CRC.

Combined Effect of Neutron and Proton Radiations on the Growth of Solid Ehrlich Ascites Carcinoma and Remote Effects in Mice.

The combined effect of the irradiation with a proton pencil scanning beam (PBS) at a total dose of 80 Gy and neutron radiation at a dose of 5 Gy on the growth of solid Ehrlich ascites carcinoma (EAC) and the remote effects in tumor-bearing mice was studied. Combined irradiation of mice with neutrons before and after irradiation with PBS, as well as irradiation only with PBS, effectively suppressed the growth of solid EAC within 1 month. In terms of the frequency and severity of radiation-induced skin reactions of mice observed 15-40 days after therapy, neutron irradiation after the irradiation with PBS showed better values of these parameters as compared to only PBS; however, exposure to neutrons before PBS was more damaging as compared to the other two options. It was also shown that the tumor relapse rate in the groups of animals with combined irradiation was higher, and the total lifespan was lower than the group of mice irradiated with PBS alone.

Boosting tumor treatment by dredging the hurdles of chemodynamic therapy synergistic ion therapy

Although chemodynamic therapy (CDT) holds the advantage of tumor treatment by generating hydroxyl radicals (·OH) without the participation of O2, insufficient H2O2 and the hindered Fe ion circulation remain major hurdles for its clinical application. Herein, we developed a hybrid nanoparticle with H2O2 self-supply and iron ion self-circulation properties for enhanced CDT. Through encapsulation of CaO2 with ZIF-8 and in situ assembled Fe3+/TA (tannic acid) on the surface, the hybrid nanoparticles (CaO2@ZIF8@MPN) were prepared. The Fe3+/TA coating could dissociate in response to lysosomal acidity, and release Fe3+ and TA subsequently. Significantly, the unprotected CaO2 can be degraded to H2O2 and Ca2+, providing sufficient materials for CDT and resulting in a Ca2+ “storm” for ion therapy. Meanwhile, TA could convert Fe3+ to Fe2+, realizing a self-circulation for promoting the sustained generation of ·OH. Interestingly, Fe3+/TA coating endows the nanoparticles with photothermal property, which accelerated the degradation and further improved the efficiency of Fenton reaction, obtaining 77.7% of tumor growth inhibited rate without obvious adverse effect in tumor-bearing mice. Our findings showed CaO2@ZIF8@MPN had potential to be applied for enhanced CDT synergistic antitumor ion therapy.

Grape Seed Proanthocyanidins play the roles of radioprotection on Normal Lung and radiosensitization on Lung Cancer via differential regulation of the MAPK Signaling Pathway.

Radiation-induced lung injury (RILI) is a common serious complication and dose-limiting factor caused by radiotherapy for lung cancer. This study was to investigate radioprotective effects of grape seed proanthocyanidins (GSP) on normal lung as well as radiosensitizing effects on lung cancer. In vitro, we demonstrated radioprotective effects of GSP on normal alveolar epithelial cells (MLE-12 and BEAS/2B) and radiosensitizing effects on lung cancer cells (LLC and A549). In vivo, we confirmed these two-way effects in tumor-bearing mice. The results showed that GSP inhibited tumor growth, and played a synergistic killing effect with radiotherapy on lung cancer. Meanwhile, GSP reduced radiation damage to normal lung tissues. The two-way effects related to the differential regulation of the MAPK signaling pathway by GSP on normal lung and lung cancer. Moreover, GSP regulated secretion of cytokines IL-6 and IFN-γ and expression of p53 and Ki67 on normal lung and lung cancer. Our findings suggest that GSP is expected to be an ideal radioprotective drug for lung cancer patients who are treated with radiotherapy.

Dual-Targeting Nanoparticle-Mediated Gene Therapy Strategy for Hepatocellular Carcinoma by Delivering Small Interfering RNA.

As a gene therapy strategy, RNA interference (RNAi) offers tremendous tumor therapy potential. However, its therapeutic efficacy is restricted by its inferior ability for targeted delivery and cellular uptake of small interfering RNA (siRNA). This study sought to develop a dual-ligand nanoparticle (NP) system loaded with siRNA to promote targeted delivery and therapeutic efficacy. We synthesized a dual receptor-targeted chitosan nanosystem (GCGA), whose target function was controlled by the ligands of galactose of lactobionic acid (LA) and glycyrrhetinic acid (GA). By loading siPAK1, an siRNA targeting P21-activated kinase 1 (PAK1), a molecular-targeted therapeutic dual-ligand NP (GCGA–siPAK1) was established. We investigated the synergistic effect of these two targeting units in hepatocellular carcinoma (HCC). In particular, GCGA–siPAK1 enhanced the NP targeting ability and promoted siPAK1 cell uptake. Subsequently, dramatic decreases in cell proliferation, invasion, and migration, with an apparent increase in cell apoptosis, were observed in treated cells. Furthermore, this dual-ligand NP gene delivery system demonstrated significant anti-tumor effects in tumor-bearing mice. Finally, we illuminated the molecular mechanism, whereby GCGA–siPAK1 promotes endogenous cell apoptosis through the PAK1/MEK/ERK pathway. Thus, the dual-target property effectively promotes the HCC therapeutic effect and provides a promising gene therapy strategy for clinical applications.

Antitumor immunity by small extracellular vesicles collected from activated dendritic cells through effective induction of cellular and humoral immune responses

Dendritic cell-derived small extracellular vesicles (DC-sEVs) are proposed as a novel candidate for tumor antigen-based cancer immunotherapy. In order to improve the DC-sEV-induced antitumor immunity, production of DC-sEVs capable of inducing potent antigen-specific humoral and cellular immune responses is necessary. Here, we collected sEVs from DCs and added ovalbumin (OVA), which was used as model antigen, as well as LPS and IFN-γ, to prepare DC-sEVs with high immune activity. After confirming that the collected sEVs, named activated-DCOVA-sEVs, contained OVA and possessed immunologically relevant components (MHC class I molecule displaying antigen epitopes and co-stimulatory molecules, as well as sEV marker proteins), we found that activated-DCOVA-sEV stimulated macrophages and DCs through Toll-like receptor 4 signaling and boosted innate immunity in the tumor microenvironment. Moreover, activated-DCOVA-sEVs induced potent antigen-specific humoral and cellular immune responses both in vitro and in vivo. Finally, immunization with activated-DCOVA-sEVs exhibited stronger in vivo antitumor effects in tumor-bearing mice induced by inoculation with OVA-expressing tumor cells.

A NAG-Guided Nano-Delivery System for Redox- and pH-Triggered Intracellularly Sequential Drug Release in Cancer Cells.

AimSequential treatment with paclitaxel (PTXL) and gemcitabine (GEM) is considered clinically beneficial for non-small-cell lung cancer. This study aimed to investigate the effectiveness of a nano-system capable of sequential release of PTXL and GEM within cancer cells.MethodsPTXL-ss-poly(6-O-methacryloyl-d-galactopyranose)-GEM (PTXL-ss-PMAGP-GEM) was designed by conjugating PMAGP with PTXL via disulfide bonds (-ss-), while GEM via succinic anhydride (PTXL:GEM=1:3). An amphiphilic block copolymer N-acetyl-d-glucosamine(NAG)-poly(styrene-alt-maleic anhydride)58-b-polystyrene130 acted as a targeting moiety and emulsifier in formation of nanostructures (NLCs).ResultsThe PTXL-ss-PMAGP-GEM/NAG NLCs (119.6 nm) provided a sequential in vitro release of, first PTXL (redox-triggered), then GEM (pH-triggered). The redox- and pH-sensitive NLCs readily distributed homogenously in the cytoplasm. NAG augmented the uptake of NLCs by the cancer cells and tumor accumulation. PTXL-ss-PMAGP-GEM/NAG NLCs exhibited synergistic cytotoxicity in vitro and strongest antitumor effects in tumor-bearing mice compared to NLCs lacking pH/redox sensitivities or free drug combination.ConclusionThis study demonstrated the abilities of PTXL-ss-PMAGP-GEM/NAG NLCs to achieve synergistic antitumor effect by targeted intracellularly sequential drug release.

First-in-human phase 1 study of IT1208, a defucosylated humanized anti-CD4 depleting antibody, in patients with advanced solid tumors

BackgroundTransient CD4+ T cell depletion led to the proliferation of tumor-specific CD8+ T cells in the draining lymph node and increased infiltration of PD-1+CD8+ T cells into the tumor, which resulted in strong anti-tumor effects in tumor-bearing mice. This is a first-in-human study of IT1208, a defucosylated humanized anti-CD4 monoclonal antibody, engineered to exert potent antibody-dependent cellular cytotoxicity.MethodsPatients with advanced solid tumors were treated with intravenous IT1208 at doses of 0.1 or 1.0 mg/kg. The first patient in each cohort received a single administration, and the other patients received two administrations of IT1208 on days 1 and 8.ResultsEleven patients were enrolled in the 0.1 mg/kg (n = 4) and 1.0 mg/kg cohorts (n = 7). Grade 1 or 2 infusion-related reactions was observed in all patients. Decreased CD4+ T cells in peripheral blood due to IT1208 were observed in all patients and especially in those receiving two administrations of 1.0 mg/kg. CD8+ T cells increased on day 29 compared with baseline in most patients, resulting in remarkably decreased CD4/8 ratios. One microsatellite-stable colon cancer patient achieved durable partial response showing increased infiltration of both CD4+ and CD8+ T cells into tumors after IT1208 administration. Moreover, transcriptomic profiling of the liver metastasis of the patient revealed upregulation of the expression of interferon-stimulated genes, T cell activation-related genes, and antigen presentation-related genes after IT1208 administration. Two additional patients with gastric or esophageal cancer achieved stable disease lasting at least 3 months.ConclusionsIT1208 monotherapy successfully depleted CD4+ T cells with a manageable safety profile and encouraging preliminary efficacy signals, which warrants further investigations, especially in combination with immune checkpoint inhibitors.

Inhibition of tumor metastasis by targeted daunorubicin and dioscin codelivery liposomes modified with PFV for the treatment of non-small-cell lung cancer.

Background: Chemotherapy for non-small-cell lung cancer (NSCLC) still leads to unsatisfactory clinical prognosis because of poor active targeting and tumor metastasis.Purpose: The objective of this study was to construct a kind of PFV peptide modified targeted daunorubicin and dioscin codelivery liposomes, which could enhance tumor targeting and inhibit tumor cell metastasis.Methods and results: Targeted daunorubicin and dioscin codelivery liposomes were prepared by film dispersion and the ammonium sulfate gradient method. With the ideal physicochemical properties, targeted daunorubicin and dioscin codelivery liposomes exhibited enhanced cellular uptake and showed strong cytotoxicity to tumor cells. The encapsulation of dioscin increased the inhibitory effects of daunorubicin on A549 cells, vasculogenic mimicry (VM) channels and tumor metastasis. The enhanced antimetastatic mechanism of the targeted liposomes was attributed to the downregulation of matrix metalloproteinase-2 (MMP-2), vascular endothelial cadherin (VE-Cad), transforming growth factor-β1 (TGF-β1) and hypoxia inducible factor-1α (HIF-1α). Meanwhile, the targeted daunorubicin and dioscin codelivery liposomes exhibited significant antitumor effects in tumor-bearing mice. H&E staining, immunohistochemistry with Ki-67 and TUNEL assay also showed the promoted antitumor activity of the targeted liposomes.Conclusion: Targeted daunorubicin and dioscin codelivery liposomes may provide an effective strategy for the treatment of NSCLC.

A comprehensive study of epigenetic alterations in hepatocellular carcinoma identifies potential therapeutic targets

A causal link has recently been established between epigenetic alterations and hepatocarcinogenesis, indicating that epigenetic inhibition may have therapeutic potential. We aimed to identify and target epigenetic modifiers that show molecular alterations in hepatocellular carcinoma (HCC). We studied the molecular-clinical correlations of epigenetic modifiers including bromodomains, histone acetyltransferases, lysine methyltransferases and lysine demethylases in HCC using The Cancer Genome Atlas (TCGA) data of 365 patients with HCC. The therapeutic potential of epigenetic inhibitors was evaluated in vitro and in vivo. RNA sequencing analysis and its correlation with expression and clinical data in the TCGA dataset were used to identify expression programs normalized by Jumonji lysine demethylase (JmjC) inhibitors. Genetic alterations, aberrant expression, and correlation between tumor expression and poor patient prognosis of epigenetic enzymes are common events in HCC. Epigenetic inhibitors that target bromodomain (JQ-1), lysine methyltransferases (BIX-1294 and LLY-507) and JmjC lysine demethylases (JIB-04, GSK-J4 and SD-70) reduce HCC aggressiveness. The pan-JmjC inhibitor JIB-04 had a potent antitumor effect in tumor bearing mice. HCC cells treated with JmjC inhibitors showed overlapping changes in expression programs related with inhibition of cell proliferation and induction of cell death. JmjC inhibition reverses an aggressive HCC gene expression program that is also altered in patients with HCC. Several genes downregulated by JmjC inhibitors are highly expressed in tumor vs. non-tumor parenchyma, and their high expression correlates with a poor prognosis. We identified and validated a 4-gene expression prognostic signature consisting of CENPA, KIF20A, PLK1, and NCAPG. The epigenetic alterations identified in HCC can be used to predict prognosis and to define a subgroup of high-risk patients that would potentially benefit from JmjC inhibitor therapy. In this study, we found that mutations and changes in expression of epigenetic modifiers are common events in human hepatocellular carcinoma, leading to an aggressive gene expression program and poor clinical prognosis. The transcriptional program can be reversed by pharmacological inhibition of Jumonji enzymes. This inhibition blocks hepatocellular carcinoma progression, providing a novel potential therapeutic strategy.

Effects of immunization against PCSK9 in an experimental model of breast cancer.

IntroductionInhibition of proprotein convertase subtilisin/kexin 9 (PCSK9) is an efficient strategy for lowering low-density lipoprotein cholesterol (LDL-C). There are, however, scant data on the efficacy and safety of PCSK9 inhibitors in non-cardiovascular diseases, particularly cancer. The present study aimed to evaluate the effect of PCSK9 inhibition using a nanoliposomal antiPCSK9 vaccine on cancer behavior and endpoints in mice bearing breast tumor.Material and methodsTo induce antiPCSK9 antibody in vivo, a nanoliposomal antiPCSK9 vaccine absorbed on 0.4% alum adjuvant was used. To induce tumor, BALB/c mice were subcutaneously inoculated with 4T1 breast carcinoma cells. After the tumor mass was palpable (approximately 10 mm3), the mice were randomly divided into four groups and subjected to different treatment protocols: (1) PBS (untreated control), (2) vaccine group, (3) combination of vaccine and Doxil, and (4) Doxil (positive control) group. Vaccine was subcutaneously administered to mice four times at 2-week intervals. Two weeks after the last administration, the vaccinated and non-vaccinated mice were subcutaneously inoculated with 4T1 breast carcinoma cells. To evaluate therapeutic efficacy, mouse body weight, tumor size, and survival were monitored every other day for 60 days.ResultsThe nanoliposomal antiPCSK9 vaccine was found to efficiently induce specific antibodies against PCSK9 in BALB/c mice, thereby decreasing plasma levels of PCSK9 and inhibiting its function. Tumor size analysis showed that time to reach endpoint (TTE) of the vaccine, combination, Doxil, and control groups was 47 ±10, 57 ±4, 60 ±4 and 39 ±9 days, respectively. Rate of tumor growth in vaccine, combination and Doxil groups was decreased by 21%, 48% and 53%, respectively, compared to the control group. Lifespan was increased by 4.2% in the vaccine group, compared with the control group. Additionally, the survival in the combination and Doxil groups was significantly higher than the vaccine and control groups.ConclusionsOur results revealed that PCSK9 inhibition may moderately improve breast cancer outcomes while having no harmful effects in tumor-bearing mice.

OK-432 Administration Inhibits Murine Allergic Rhinitis at the Induction Phase, through the Macrophage Activation with TLR2 Signaling Pathway.

OK-432, a preparation of a low-virulence strain (Su) of Streptococcus pyogenes (Group A) killed by a penicillin and lyophilized, is a stiff inducer of Th1 cytokines, and exerts anti-cancer effects in tumor-bearing mice. OK-432 has been reported to consist of many bacterial components, such as peptidoglycan, M-protein, etc. However, it is yet to be ascertained which bacterial component induces T helper 1 (Th1) responses. For the last decade, Toll-like receptor (TLR) family proteins are well elucidated to play a role in recognizing bacterial components and inducing interleukin (IL)-12 from macrophages. Above all, peptidoglycan seems to be the agonist of TLR2 rather than the obverse. In our present study, the role of TLR2 for the recognition of OK-432 by macrophages and the effects of OK-432 are examined on murine allergic rhinitis model. Interestingly, results show IL-12 production by macrophages derived from TLR2 knock-out (ko) mice was significantly decreased, in comparison with that of macrophages derived from wild-type mice. Moreover, in TLR2 ko mice, no regulatory effect of OK-432 was observed on an allergic rhinitis model. These data indicate that TLR2 signaling is involved in regulating OK-432-induced anti-T helper 2 (Th2) immunity, and may offer a new prophylactic and therapeutic approach using OK-432 to downregulate allergic disorders, such as allergic rhinitis.

120 - ROS-targeting of melanoma cells provides immuno-protection against tumor growth

Breaking barriers in science necessitates crossing of boundaries between disciplines. An intimate link is presented that connects the fields of redox research, cancer therapy, and immunology. Immunogenic cancer cell death (ICD) provides inflammatory stimuli to elicit anti-tumor adaptive immune responses. Release of adjuvant find-me (HMGB1, ATP) and eat-me (calreticulin) signals facilitates DC maturation and cross presentation of neoantigens to T cells. Key hallmarks of ICD in tumor cells are stress of the endoplasmic reticulum and the unfolded proteins response. Both are known to be subject to redox control. Potent ICD-inducing therapies are drugs such as anthracyclines, ionizing irradiation, or photodynamic therapies. Intriguingly, these antitumor approaches are reported to show concomitant generation of reactive species that can drive signaling responses through redox relays. Hence, the importance of reactive species and specifically hydrogen peroxide signaling are underappreciated in the onset of ICD in cancer cells. Recently, a novel technology capable of generating various oxidants has not only been shown to confer remarkable effects in tumor-bearing mice but is also a potent ICD inducer. This cold physical plasma, a partially ionized gas at about body temperature, can be applied to tumors topically, and first palliative cancer patients benefited from plasma therapy. Experimental evidence will be presented that i) physical plasma is an ICD inducer in various cell types including melanoma, ii) this is concomitant with a global HMOX1 signature, and iii) plasma-derived ROS evoke an antitumor T-cell response in tumor-bearing mice.

Combined Blockade of IL6 and PD-1/PD-L1 Signaling Abrogates Mutual Regulation of Their Immunosuppressive Effects in the Tumor Microenvironment.

Recently emerging cancer immunotherapies combine the applications of therapeutics to disrupt the immunosuppressive conditions in tumor-bearing hosts. In this study, we found that targeting the proinflammatory cytokine IL6 enhances tumor-specific Th1 responses and subsequent antitumor effects in tumor-bearing mice. IL6 blockade upregulated expression of the immune checkpoint molecule programmed death-ligand 1 (PD-L1) on melanoma cells. This PD-L1 induction was canceled in IFNγ-deficient mice or CD4+ T cell-depleted mice, suggesting that CD4+ T cell-derived IFNγ is important for PD-L1 induction in tumor-bearing hosts. In some patients with melanoma, however, treatment with the anti-PD-1 antibody nivolumab increased systemic levels of IL6, which was associated with poor clinical responses. This PD-L1 blockade-evoked induction of IL6 was reproducible in melanoma-bearing mice. We found that PD-1/PD-L1 blockade prompted PD-1+ macrophages to produce IL6 in the tumor microenvironment. Depletion of macrophages in melanoma-bearing mice reduced the levels of IL6 during PD-L1 blockade, suggesting macrophages are responsible for the IL6-mediated defective CD4+ Th1 response. Combined blockade of the mutually regulated immunosuppressive activities of IL6 and PD-1/PD-L1 signals enhanced expression of T cell-attracting chemokines and promoted infiltration of IFNγ-producing CD4+ T cells in tumor tissues, exerting a synergistic antitumor effect, whereas PD-L1 blockade alone did not promote Th1 response. Collectively, these findings suggest that IL6 is a rational immunosuppressive target for overcoming the narrow therapeutic window of anti-PD-1/PD-L1 therapy.Significance: These findings advance our understanding of IL6-PD1/PD-L1 cross-talk in the tumor microenvironment and provide clues for targeted interventional therapy that may prove more effective against cancer. Cancer Res; 78(17); 5011-22. ©2018 AACR.

Inhibition of Tumor Growth via Systemic siRNA Delivery Using Reducible Bile Acid-Conjugated Polyethylenimine.

RNA interference (RNAi), mediated by small interfering RNA (siRNA), has been considered as a potential therapeutic agent for cancer owing to its ability to suppress target genes in a sequence-specific manner. In this study, a conjugate of the low molecular weight (MW) polyethylenimine (PEI) (MW 1800) and deoxycholic acid (DA) was further modified with 4-fluorothiophenol (FTP) (TP-DA-PEI) to achieve systemic siRNA delivery. The thiophenol group would be involved with disulfide bonds between the polymer chains and siRNA modified with free thiols (thiol-siRNA) to form and π–π interactions between the pendent aromatic groups and coprostane ring of the bile acid. The TP-DA-PEI conjugates could generate stable nanoparticles with thiol-siRNA. The TP-DA-PEI conjugate not only achieved enhanced intracellular uptake, serum stability, and transfection efficiency, but also showed high accumulation of TP-DA-PEI/thiol-siRNA polyplexes and significant tumor growth inhibition effect in tumor-bearing mice after systemic administration.

P-38 - Reactive species in immunogenic cancer cell death

Immunogenic cancer cell death (ICD) provides inflammatory stimuli to elicit anti-tumor immunity. Key hallmarks of ICD are ER-stress and the unfolded proteins response, both being redox controlled. ICD-inducing therapies are drugs such as anthracyclines, ionizing irradiation, or photodynamic therapies. Intriguingly, these antitumor approaches show concomitant generation of reactive species that can drive signaling responses through redox relays. Hence, the importance of reactive species and specifically hydrogen peroxide signaling are underappreciated in the onset of ICD in cancer cells. Recently, a novel technology capable of generating various oxidants has not only been shown to confer remarkable effects in tumor-bearing mice but is also a potent ICD inducer. This cold physical plasma, a partially ionized gas, can be applied to tumors topically, and first palliative cancer patients benefited from plasma therapy. Experimental evidence will be presented that i) physical plasma is an ICD inducer in various cell types including melanoma, ii) by this it provides immunomodulatory stimuli especially in macrophages, and iii) plasma treatment is of relevance in animal tumor models.

Self-Assembling Lipid-Peptide Hybrid Nanoparticles of Phospholipid-Nonaarginine Conjugates for Enhanced Delivery of Nucleic Acid Therapeutics.

Despite potential applications of nucleic acid therapeutics, the lack of effective delivery systems hinders their clinical application. To overcome the barriers to nucleic acid delivery, we previously reported nanoparticles using phospholipid-polyethylenimine conjugates. However, toxicity of polyethylenimine remains as a problematic issue. Herein, we proposed to substitute the polyethylenimine with arginine-rich peptide to obtain a less-toxic carrier system. Nonaarginine was conjugated to the distal end of phospholipid hydrocarbon chains leading to phospholipid-nonaarginine conjugates (PL9R) and then lipid-peptide hybrid nanoparticles carrying oligonucleotide therapeutics (hNP) were constructed by self-assembly process. The hNP were further modified with cell penetrating Tat peptide (T-hNP) to enhance cellular uptake. The PL9R was less cytotoxic, and the hNP showed high loading capacity and colloidal stability. The T-hNP showed higher cellular uptake and transfection efficiency and effective accumulation to tumor tissue and silencing effect in tumor bearing mice. Altogether, T-hNP could provide a promising nanocarrier for nucleic acid therapeutics.

Inhibition of mouse breast adenocarcinoma growth by ablation with intratumoral alpha-irradiation combined with inhibitors of immunosuppression and CpG.

It has been demonstrated that aggressive in situ tumor destruction (ablation) could lead to the release of tumor antigens, which can stimulate anti-tumor immune responses. We developed an innovative method of tumor ablation based on intratumoral alpha-irradiation, diffusing alpha-emitters radiation therapy (DaRT), which efficiently ablates local tumors and enhances anti-tumor immunity. In this study, we investigated the anti-tumor potency of a treatment strategy, which combines DaRT tumor ablation with two approaches for the enhancement of anti-tumor reactivity: (1) neutralization of immunosuppressive cells such as regulatory T cells (Tregs) and myeloid-derived suppressor cells (MDSCs) and (2) boost the immune response by the immunoadjuvant CpG. Mice bearing DA3 mammary adenocarcinoma with metastases were treated with DaRT wires in combination with a MDSC inhibitor (sildenafil), Treg inhibitor (cyclophosphamide at low dose), and the immunostimulant, CpG. Combination of all four therapies led to a complete rejection of primary tumors (in 3 out of 20 tumor-bearing mice) and to the elimination of lung metastases. The treatment with DaRT and Treg or MDSC inhibitors (without CpG) also resulted in a significant reduction in tumor size, reduced the lung metastatic burden, and extended survival compared to the corresponding controls. We suggest that the therapy with DaRT combined with the inhibition of immunosuppressive cells and CpG reinforced both local and systemic anti-tumor immune responses and displayed a significant anti-tumor effect in tumor-bearing mice.