Myeloid cell-targeted miR-146a mimic inhibits NF-κB-driven inflammation and leukemia progression in vivo.

NF-κB is a key regulator of inflammation and cancer progression, with an important role in leukemogenesis. Despite its therapeutic potential, targeting NF-κB using pharmacologic inhibitors has proven challenging. Here, we describe a myeloid cell-selective NF-κB inhibitor using an miR-146a mimic oligonucleotide conjugated to a scavenger receptor/Toll-like receptor 9 agonist (C-miR146a). Unlike an unconjugated miR146a, C-miR146a was rapidly internalized and delivered to the cytoplasm of target myeloid cells and leukemic cells. C-miR146a reduced expression of classic miR-146a targets (IRAK1 and TRAF6), thereby blocking activation of NF-κB in target cells. IV injections of C-miR146a mimic to miR-146a-deficient mice prevented excessive NF-κB activation in myeloid cells, and thus alleviated myeloproliferation and mice hypersensitivity to bacterial challenge. Importantly, C-miR146a showed efficacy in dampening severe inflammation in clinically relevant models of chimeric antigen receptor (CAR) T-cell-induced cytokine release syndrome. Systemic administration of C-miR146a oligonucleotide alleviated human monocyte-dependent release of IL-1 and IL-6 in a xenotransplanted B-cell lymphoma model without affecting CD19-specific CAR T-cell antitumor activity. Beyond anti-inflammatory functions, miR-146a is a known tumor suppressor commonly deleted or expressed at reduced levels in human myeloid leukemia. Using The Cancer Genome Atlas acute myeloid leukemia data set, we found an inverse correlation of miR-146a levels with NF-κB-related genes and with patient survival. Correspondingly, C-miR146a induced cytotoxic effects in human MDSL, HL-60, and MV4-11 leukemia cells in vitro. The repeated IV administration of C-miR146a inhibited expression of NF-κB target genes and thereby thwarted progression of disseminated HL-60 leukemia. Our results show the potential of using myeloid cell-targeted miR-146a mimics for the treatment of inflammatory and myeloproliferative disorders.

Baicalein ameliorates ulcerative colitis by improving intestinal epithelial barrier via AhR/IL-22 pathway in ILC3s.

Ulcerative colitis (UC) is a chronic inflammatory disease of the gastrointestinal tract, which is closely related to gut barrier dysfunction. Emerging evidence shows that interleukin-22 (IL-22) derived from group 3 innate lymphoid cells (ILC3s) confers benefits on intestinal barrier, and IL-22 expression is controlled by aryl hydrocarbon receptor (AhR). Previous studies show that baicalein protects the colon from inflammatory damage. In this study we elucidated the molecular mechanisms underlying the protective effect of baicalein on intestinal barrier function in colitis mice. Mice were administered baicalein (10, 20, 40 mg·kg-1·d-1, i.g.) for 10 days; the mice freely drank 3% dextran sulfate sodium (DSS) on D1-D7 to induce colitis. We showed that baicalein administration simultaneously ameliorated gut inflammation, decreased intestinal permeability, restored tight junctions of colons possibly via promoting AhR/IL-22 pathway. Co-administration of AhR antagonist CH223191 (10 mg/kg, i.p.) partially blocked the therapeutic effects of baicalein in colitis mice, whereas AhR agonist FICZ (1 µg, i.p.) ameliorated symptoms and gut barrier function in colitis mice. In a murine lymphocyte line MNK-3, baicalein (5-20 µM) dose-dependently increased the expression of AhR downstream target protein CYP1A1, and enhanced IL-22 production through facilitating AhR nuclear translocation, these effects were greatly diminished in shAhR-MNK3 cells, suggesting that baicalein induced IL-22 production in AhR-dependent manner. To further clarify that, we constructed an in vitro system consisting of MNK-3 and Caco-2 cells, in which MNK-3 cell supernatant treated with baicalein could decrease FITC-dextran permeability and promoted the expression of tight junction proteins ZO-1 and occluding in Caco-2 cells. In conclusion, this study demonstrates that baicalein ameliorates colitis by improving intestinal epithelial barrier via AhR/IL-22 pathway in ILC3s, thus providing a potential therapy for UC.

Targeted In Vivo Delivery of NF-κB Decoy Inhibitor Augments Sensitivity of B Cell Lymphoma to Therapy.

Despite recent advances, non-Hodgkin's B cell lymphoma patients often relapse or remain refractory to therapy. Therapeutic resistance is often associated with survival signaling via nuclear factor κB (NF-κB) transcription factor, an attractive but undruggable molecular target. In this study, we describe a bipartite inhibitor comprising a NF-κB-specific decoy DNA tethered to a CpG oligodeoxynucleotide (ODN) targeting Toll-like receptor-9-expressing B cell lymphoma cells. The Bc-NFκBdODN showed efficient uptake by human diffuse large B cell (U2932, OCI-Ly3), Burkitt (RaJi), and mantle cell (Jeko1) lymphomas, respectively. We confirmed that Bc-NFκBdODN inhibited NF-κB nuclear translocation and DNA binding, resulting in CCND2 and MYC downregulation. Bc-NFκBdODN enhanced radiosensitivity of lymphoma cells in vitro. In xenotransplanted human lymphoma, local injections of Bc-NFκBdODN reduced NF-κB activity in whole tumors. When combined with a local 3-Gy dose of radiation, Bc-NFκBdODN effectively arrested OCI-Ly3 lymphoma progression. In immunocompetent mice, intratumoral injections of Bc-NFκBdODN suppressed growth of directly treated and distant A20 lymphomas, as a result of systemic CD8 T cell-dependent immune responses. Finally, systemic administration of Bc-NFκBdODN to mice bearing disseminated A20 lymphoma induced complete regression and extended survival of most of the treated mice. Our results underscore clinical relevance of this strategy as monotherapy and in support of radiation therapy to benefit patients with resistant or relapsed B cell lymphoma.

Graphene Quantum Dots-Mediated Theranostic Penetrative Delivery of Drug and Photolytics in Deep Tumors by Targeted Biomimetic Nanosponges.

Dual-targeted delivery of drugs and energy by nanohybrids can potentially alleviate side effects and improve the unique features required for precision medicine. To realize this aim, however, the hybrids which are often rapidly removed from circulation and the piled up tumors periphery near the blood vessels must address the difficulties in low blood half-lives and tumor penetration. In this study, a sponge-inspired carbon composites-supported red blood cell (RBC) membrane that doubles as a stealth agent and photolytic carrier that transports tumor-penetrative agents (graphene quantum dots and docetaxel (GQD-D)) and heat with irradiation was developed. The RBC-membrane enveloped nanosponge (RBC@NS) integrated to a targeted protein that accumulates in tumor spheroids via high lateral bilayer fluidity exhibits an 8-fold increase in accumulation compared to the NS. Penetrative delivery of GQDs to tumor sites is actuated by near-infrared irradiation through a one-atom-thick structure, facilitating penetration and drug delivery deep into the tumor tissue. The synergy of chemotherapy and photolytic effects was delivered by the theranostic GQDs deep into tumors, which effectively damaged and inhibited the tumor in 21 days when treated with a single irradiation. This targeted RBC@GQD-D/NS with the capabilities of enhanced tumor targeting, NIR-induced drug penetration into tumors, and thermal ablation for photolytic therapy promotes tumor suppression and exhibits potential for other biomedical applications.

Ultrasound-Vortex-Assisted Dispersive Liquid-Liquid Microextraction Combined with High Performance Liquid Chromatography-Diode Array Detection for Determining UV Filters in Cosmetics and the Human Stratum Corneum.

This study explores the amounts of common chemical ultraviolet (UV) filters (i.e., avobenzone, bemotrizinol, ethylhexyl triazone, octocrylene, and octyl methoxycinnamate) in cosmetics and the human stratum corneum. An ultrasound-vortex-assisted dispersive liquid-liquid microextraction (US-VA-DLLME) method with a high-performance liquid chromatography-diode array detector was used to analyze UV filters. A bio-derived solvent (i.e., anisole) was used as the extractant in the US-VA-DLLME procedure, along with methanol as the dispersant, a vortexing time of 4 min, and ultrasonication for 3 min. The mass-transfer rate of the extraction process was enhanced due to vortex-ultrasound combination. Various C18 end-capped columns were used to investigate the separation characteristics of the UV filters, with XBridge BEH or CORTECS selected as the separation column. Calibration curves were constructed in the 0.05-5 µg/mL (all filters except octocrylene) and 0.1-10 µg/mL (octocrylene) ranges, and excellent analytical linearities with coefficients of determination (r2) above 0.998. The developed method was successfully used to analyze sunscreen. Moreover, experiments were designed to simulate the sunscreen-usage habits of consumers, and the cup method was used to extract UV filters from the human stratum corneum. The results suggest that a makeup remover should be employed to remove water-in-oil sunscreens from skin.

B Cell Lymphoma Immunotherapy Using TLR9-Targeted Oligonucleotide STAT3 Inhibitors.

Growing evidence links the aggressiveness of non-Hodgkin's lymphoma, especially the activated B cell-like type diffuse large B cell lymphomas (ABC-DLBCLs) to Toll-like receptor 9 (TLR9)/MyD88 and STAT3 transcription factor signaling. Here, we describe a dual-function molecule consisting of a clinically relevant TLR9 agonist (CpG7909) and a STAT3 inhibitor in the form of a high-affinity decoy oligodeoxynucleotide (dODN). The CpG-STAT3dODN blocked STAT3 DNA binding and activity, thus reducing expression of downstream target genes, such as MYC and BCL2L1, in human and mouse lymphoma cells. We further demonstrated that injections (i.v.) of CpG-STAT3dODN inhibited growth of human OCI-Ly3 lymphoma in immunodeficient mice. Moreover, systemic CpG-STAT3dODN administration induced complete regression of the syngeneic A20 lymphoma, resulting in long-term survival of immunocompetent mice. Both TLR9 stimulation and concurrent STAT3 inhibition were critical for immune-mediated therapeutic effects, since neither CpG7909 alone nor CpG7909 co-injected with unconjugated STAT3dODN extended mouse survival. The CpG-STAT3dODN induced expression of genes critical to antigen-processing/presentation and Th1 cell activation while suppressing survival signaling. These effects resulted in the generation of lymphoma cell-specific CD8/CD4-dependent T cell immunity protecting mice from tumor rechallenge. Our results suggest that CpG-STAT3dODN as a systemic/local monotherapy or in combination with PD1 blockade can provide an opportunity for treating patients with B cell NHL.

STAT3 in Tumor-Associated Myeloid Cells: Multitasking to Disrupt Immunity.

Myeloid immune cells, such as dendritic cells, monocytes, and macrophages, play a central role in the generation of immune responses and thus are often either disabled or even hijacked by tumors. These new tolerogenic activities of tumor-associated myeloid cells are controlled by an oncogenic transcription factor, signal transducer and activator of transcription 3 (STAT3). STAT3 multitasks to ensure tumors escape immune detection by impairing antigen presentation and reducing production of immunostimulatory molecules while augmenting the release of tolerogenic mediators, thereby reducing innate and adaptive antitumor immunity. Tumor-associated myeloid cells and STAT3 signaling in this compartment are now commonly recognized as an attractive cellular target for improving efficacy of standard therapies and immunotherapies. Hereby, we review the importance and functional complexity of STAT3 signaling in this immune cell compartment as well as potential strategies for cancer therapy.

The C-terminal disulfide bonds of Helicobacter pylori GroES are critical for IL-8 secretion via the TLR4-dependent pathway in gastric epithelial cells.

Helicobacter pylori GroES (HpGroES), a potent immunogen, is a secreted virulence factor that stimulates production of proinflammatory cytokines and may contribute to gastric carcinogenesis. HpGroES is larger than other bacterial orthologs because of an additional C-terminal region, known as domain B. We found that the HpGroES-induced IL-8 release by human gastric epithelial cells was dependent on activation of the MAPK and NF-κB pathways. HpGroES lacking domain B was unable to induce IL-8 release. Additionally, a TLR4 inhibitor significantly inhibited IL-8 secretion and reduced HpGroES-induced activation of MAPKs. Furthermore, HpGroES-induced IL-8 release by primary gastric epithelial cells from TLR4(-/-) mice was significantly lower than from wild-type mice. We also found that HpGroES bound to TLR4 in cell lysates and colocalized with TLR4 on the cell membrane only when domain B was present. We then constructed two deletion mutants lacking C-terminal regions and mutants with point mutations of two of the four cysteine residues, C111 and C112, in domain B and found that the deletion mutants and a double mutant lacking the C94-C111 and C95-C112 disulfide bonds were unable to interact with TLR4 or induce IL-8 release. We conclude that HpGroES, in which a unique conformational structure, domain B, is generated by these two disulfide bonds, induces IL-8 secretion via a TLR4-dependent mechanism.

Jejunal Cavernous Hemangioma Mimicking Malignancy With Increased Activity on 18 F-FDG PET/CT.

ABSTRACT: A 50-year-old woman underwent 18 F-FDG PET/CT to evaluate possible abdominal malignancy, which was revealed by CT. The images showed a large cystic-solid lesion with peripherally increased FDG activity in the left mid-abdomen. Histopathology of the excised lesion confirmed a jejunal cavernous hemangioma. We reported a rare case of jejunal cavernous hemangioma with FDG accumulation on PET/CT, mimicking malignancy.

Rabies Virus Glycoprotein-Mediated Transportation and T Cell Infiltration to Brain Tumor by Magnetoelectric Gold Yarnballs.

T lymphocyte infiltration with immunotherapy potentially suppresses most devastating brain tumors. However, local immune privilege and tumor heterogeneity usually limit the penetration of immune cells and therapeutic agents into brain tumors, leading to tumor recurrence after treatment. Here, a rabies virus glycoprotein (RVG)-camouflaged gold yarnball (RVG@GY) that can boost the targeting efficiency at a brain tumor via dual hierarchy- and RVG-mediated spinal cord transportation, facilitating the decrease of tumor heterogeneity for T cell infiltration, is developed. Upon magnetoelectric irradiation, the electron current generated on the GYs activates the electrolytic penetration of palbociclib-loaded dendrimer (Den[Pb]) deep into tumors. In addition, the high-density GYs at brain tumors also induces the disruption of cell-cell interactions and T cell infiltration. The integration of the electrolytic effects and T cell infiltration promoted by drug-loaded RVG@GYs deep in the brain tumor elicits sufficient T cell numbers and effectively prolongs the survival rate of mice with orthotopic brain tumors.

Optimization and Development of Selective Histone Deacetylase Inhibitor (MPT0B291)-Loaded Albumin Nanoparticles for Anticancer Therapy.

Histone deacetylase (HDAC) inhibitors have emerged as a new class of antitumor agent for various types of tumors. MPT0B291, a novel selective inhibitor of HDAC6, demonstrated significant antiproliferative activity in various human cancer cell types. However, MPT0B291 has very low water solubility, which limits its clinical use for cancer therapy. In the current study, MPT0B291 was encapsulated in human serum albumin (HSA), and its anticancer activities were investigated. Nanoparticles (NPs) were prepared using two-stage emulsification resulting in 100~200-nm NPs with a fine size distribution (polydispersity index of <0.3). The in vitro drug release profiles of MPT0B291-loaded HSA NPs presented sustained-release properties. The cytotoxic effect on MIA PaCa-2 human pancreatic carcinoma cells was found to be similar to MPT0B291-loaded HSA NPs and the free-drug group. The albumin-based formulation provided a higher maximum tolerated dose than that of a drug solution with reduced toxicity toward normal cells. Furthermore, in vivo pharmacokinetic studies demonstrated an effective increase (5~8-fold) in the bioavailability of NPs containing MPT0B291 loaded in HSA compared to the free-drug solution with an extended circulation time (t1/2) leading to significantly enhanced efficacy of anticancer treatment.

Myeloid cell-targeted STAT3 inhibition sensitizes head and neck cancers to radiotherapy and T cell-mediated immunity.

The tumor microenvironment affects the outcome of radiotherapy against head and neck squamous cell carcinoma (HNSCC). We recently found that tolerogenic myeloid cells accumulate in the circulation of HNSCC patients undergoing radiotherapy. Here, we analyzed tumor-containing lymph node biopsies collected from these patients. After 2 weeks of radiotherapy, we found an increase in tumor-associated macrophages (TAMs) with activated STAT3, while CD8+ T cells were reduced as detected using multiplex IHC. Gene expression profiling indicated upregulation of M2 macrophage-related genes (CD163, CD206), immunosuppressive mediators (ARG1, LIF, TGFB1), and Th2 cytokines (IL4, IL5) in irradiated tumors. We next validated STAT3 as a potential target in human HNSCC-associated TAMs, using UM-SCC1 xenotransplants in humanized mice. Local injections of myeloid cell-targeted STAT3 antisense oligonucleotide (CpG-STAT3ASO) activated human DCs/macrophages and promoted CD8+ T cell recruitment, thereby arresting UM-SCC1 tumor growth. Furthermore, CpG-STAT3ASO synergized with tumor irradiation against syngeneic HPV+ mEERL and HPV- MOC2 HNSCC tumors in mice, triggering tumor regression and/or extending animal survival. The antitumor immune responses were CD8+ and CD4+ T cell dependent and associated with the activation of antigen-presenting cells (DCs/M1 macrophages) and increased CD8+ to regulatory T cell ratio. Our observations suggest that targeted inhibition of STAT3 in tumor-associated myeloid cells augments the efficacy of radiotherapy against HNSCC.

Supramolecularly enabled pH- triggered drug action at tumor microenvironment potentiates nanomedicine efficacy against glioblastoma.

The crucial balance of stability in blood-circulation and tumor-specific delivery has been suggested as one of the challenges for effective bench-to-bedside translation of nanomedicines (NMs). Herein, we developed a supramolecularly enabled tumor-extracellular (Tex) pH-triggered NM that can maintain the micellar structure with the entrapped-drug during systemic circulation and progressively release drug in the tumor by rightly sensing heterogeneous tumor-pH. Desacetylvinblastine hydrazide (DAVBNH), a derivative of potent anticancer drug vinblastine, was conjugated to an aliphatic ketone-functionalized poly(ethylene glycol)-b-poly(amino acid) copolymer and the hydrolytic stability of the derived hydrazone bond was efficiently tailored by exploiting the compartmentalized structure of polymer micelle. We confirmed an effective and safe therapeutic application of Tex pH-sensitive DAVBNH-loaded micelle (Tex-micelle) in orthotopic glioblastoma (GBM) models, extending median survival to 1.4 times in GBM xenograft and 2.6 times in GBM syngeneic model, compared to that of the free DAVBNH. The work presented here offers novel chemical insights into the molecular design of smart NMs correctly sensing Tex-pH via programmed functionalities. The practical engineering strategy based on a clinically relevant NM platform, and the encouraging therapeutic application of Tex-micelle in GBM, one of the most lethal human cancers, thus suggests the potential clinical translation of this system against other types of common cancers, including GBM.

Cytoplasmic DROSHA and non-canonical mechanisms of MiR-155 biogenesis in FLT3-ITD acute myeloid leukemia.

We report here on a novel pro-leukemogenic role of FMS-like tyrosine kinase 3-internal tandem duplication (FLT3-ITD) that interferes with microRNAs (miRNAs) biogenesis in acute myeloid leukemia (AML) blasts. We showed that FLT3-ITD interferes with the canonical biogenesis of intron-hosted miRNAs such as miR-126, by phosphorylating SPRED1 protein and inhibiting the "gatekeeper" Exportin 5 (XPO5)/RAN-GTP complex that regulates the nucleus-to-cytoplasm transport of pre-miRNAs for completion of maturation into mature miRNAs. Of note, despite the blockage of "canonical" miRNA biogenesis, miR-155 remains upregulated in FLT3-ITD+ AML blasts, suggesting activation of alternative mechanisms of miRNA biogenesis that circumvent the XPO5/RAN-GTP blockage. MiR-155, a BIC-155 long noncoding (lnc) RNA-hosted oncogenic miRNA, has previously been implicated in FLT3-ITD+ AML blast hyperproliferation. We showed that FLT3-ITD upregulates miR-155 by inhibiting DDX3X, a protein implicated in the splicing of lncRNAs, via p-AKT. Inhibition of DDX3X increases unspliced BIC-155 that is then shuttled by NXF1 from the nucleus to the cytoplasm, where it is processed into mature miR-155 by cytoplasmic DROSHA, thereby bypassing the XPO5/RAN-GTP blockage via "non-canonical" mechanisms of miRNA biogenesis.

Treatment-induced arteriolar revascularization and miR-126 enhancement in bone marrow niche protect leukemic stem cells in AML.

BACKGROUND: During acute myeloid leukemia (AML) growth, the bone marrow (BM) niche acquires significant vascular changes that can be offset by therapeutic blast cytoreduction. The molecular mechanisms of this vascular plasticity remain to be fully elucidated. Herein, we report on the changes that occur in the vascular compartment of the FLT3-ITD+ AML BM niche pre and post treatment and their impact on leukemic stem cells (LSCs). METHODS: BM vasculature was evaluated in FLT3-ITD+ AML models (MllPTD/WT/Flt3ITD/ITD mouse and patient-derived xenograft) by 3D confocal imaging of long bones, calvarium vascular permeability assays, and flow cytometry analysis. Cytokine levels were measured by Luminex assay and miR-126 levels evaluated by Q-RT-PCR and miRNA staining. Wild-type (wt) and MllPTD/WT/Flt3ITD/ITD mice with endothelial cell (EC) miR-126 knockout or overexpression served as controls. The impact of treatment-induced BM vascular changes on LSC activity was evaluated by secondary transplantation of BM cells after administration of tyrosine kinase inhibitors (TKIs) to MllPTD/WT/Flt3ITD/ITD mice with/without either EC miR-126 KO or co-treatment with tumor necrosis factor alpha (TNFα) or anti-miR-126 miRisten. RESULTS: In the normal BM niche, CD31+Sca-1high ECs lining arterioles have miR-126 levels higher than CD31+Sca-1low ECs lining sinusoids. We noted that during FLT3-ITD+ AML growth, the BM niche lost arterioles and gained sinusoids. These changes were mediated by TNFα, a cytokine produced by AML blasts, which induced EC miR-126 downregulation and caused depletion of CD31+Sca-1high ECs and gain in CD31+Sca-1low ECs. Loss of miR-126high ECs led to a decreased EC miR-126 supply to LSCs, which then entered the cell cycle and promoted leukemia growth. Accordingly, antileukemic treatment with TKI decreased the BM blast-produced TNFα and increased miR-126high ECs and the EC miR-126 supply to LSCs. High miR-126 levels safeguarded LSCs, as shown by more severe disease in secondary transplanted mice. Conversely, EC miR-126 deprivation via genetic or pharmacological EC miR-126 knock-down prevented treatment-induced BM miR-126high EC expansion and in turn LSC protection. CONCLUSIONS: Treatment-induced CD31+Sca-1high EC re-vascularization of the leukemic BM niche may represent a LSC extrinsic mechanism of treatment resistance that can be overcome with therapeutic EC miR-126 deprivation.

Rabies virus glycoprotein-amplified hierarchical targeted hybrids capable of magneto-electric penetration delivery to orthotopic brain tumor.

Compact nanohybrids can potentially unite various therapeutic features and reduce side effects for precise cancer therapy. However, the poor accumulation and limited tumor penetration of drugs at the tumor impede the manifestation of nanomedicine. We developed a rabies virus glycoprotein (RVG)-amplified hierarchical targeted hybrid that acts as a stealthy and magnetolytic carrier that transports dual tumor-penetrating agents incorporating two drugs (boron-doped graphene quantum dots (B-GQDs)/doxorubicin and pH-responsive dendrimers (pH-Den)/palbociclib). The developed RVG-decorated hybrids (RVG-hybrids) enhance the accumulation of drugs at tumor by partially bypassing the BBB via spinal cord transportation and pH-induced aggregation of hierarchical targeting. The penetrated delivery of dual pH-Den and B-GQD drugs to deep tumors is actuated by magnetoelectric effect, which are able to generate electrons to achieve electrostatic repulsion and disassemble the hybrids into components of a few nanometers in size. The synergy of magnetoelectric drug penetration and chemotherapy was achieved by delivery of the B-GQDs and pH-Den to orthotopic tumors, which prolonged the host survival time. This RVG-amplified dual hierarchical delivery integrated with controlled and penetrated release from this hybrid improve the distribution of the therapeutic agents at the brain tumor for synergistic therapy, exhibiting potential for clinic use.

Beclin-1 as a neutrophil-specific immune checkpoint.

Neutrophils are early wound healing and inflammation regulators that, due to functional plasticity, can adopt either pro- or antitumor functions. Until recently, beclin-1 was a protein known mainly for its role as a critical regulator of autophagy. In this issue of the JCI, Tan et al. describe the effects of the beclin-1 conditional myeloid cell-specific deletion in mice, in which immunostimulation resulted in hypersensitive neutrophils. The chronic proinflammatory effect of these neutrophils triggered spontaneous B cell malignancies to develop. Such tumorigenic effects were mediated primarily by IL-21 and CD40 signaling, leading to the upregulation of tolerogenic molecules, such as IL-10 and PD-L1. The authors went on to examine samples derived from patient lymphoid malignancies and showed that beclin-1 expression in neutrophils positively correlated with pre-B cell leukemia/lymphoma. Overall, the study provides an elegant model for neutrophil-driven carcinogenesis and identifies potential targets for immunotherapy of B cell malignancies.

Targeted Delivery of miRNA Antagonists to Myeloid Cells In Vitro and In Vivo.

Elevated levels of microRNAs in cancer cells are often associated with oncogenic effects and thus provide potential therapeutic targets. However, the lack of efficient delivery methods for synthetic miRNA inhibitors, antagomiR, or anti-miR oligonucleotides hindered clinical translation of such strategies. We recently developed an approach for targeted delivery of synthetic, 2'-O-methyl-modified antagomiR molecules to normal and malignant myeloid cells and B cells by tethering to the single-stranded, phosphorothioate oligodeoxynucleotides (PSO). The PSO-antagomiR are rapidly internalized through scavenger receptor-mediated endocytosis by human monocytes, dendritic cells, B cells, as well as myeloid leukemia and B-cell lymphoma cells, but not by T cells. Following internalization, the unformulated PSO-antagomiR potently reduces levels of target miRNA and modulates expression of downstream protein targets, both in vitro and in vivo. The simple design of PSO-antagomiR conjugates enable adaptation of this strategy for targeting oncogenic miRNAs in nonmalignant and malignant myeloid cells and B cells.

Functional Nanoparticles for Tumor Penetration of Therapeutics.

Theranostic nanoparticles recently received great interest for uniting unique functions to amplify therapeutic efficacy and reduce side effects. Despite the enhanced permeability and retention (EPR) effect, which amplifies the accumulation of nanoparticles at the site of a tumor, tumor heterogeneity caused by the dense extracellular matrix of growing cancer cells and the interstitial fluid pressure from abnormal angiogenesis in the tumor inhibit drug/particle penetration, leading to inhomogeneous and limited treatments. Therefore, nanoparticles for penetrated delivery should be designed with different strategies to enhance efficacy. Many strategies were developed to overcome the obstacles in cancer therapy, and they can be divided into three main parts: size changeability, ligand functionalization, and modulation of the tumor microenvironment. This review summarizes the results of ameliorated tumor penetration approaches and amplified therapeutic efficacy in nanomedicines. As the references reveal, further study needs to be conducted with comprehensive strategies with broad applicability and potential translational development.

STAT3 Inhibition Combined with CpG Immunostimulation Activates Antitumor Immunity to Eradicate Genetically Distinct Castration-Resistant Prostate Cancers.

PURPOSE: Prostate cancers show remarkable resistance to emerging immunotherapies, partly due to tolerogenic STAT3 signaling in tumor-associated myeloid cells. Here, we describe a novel strategy combining STAT3 inhibition with Toll-like Receptor 9 (TLR9) stimulation to unleash immune response against prostate cancers regardless of the genetic background. EXPERIMENTAL DESIGN: We developed and validated a conjugate of the STAT3 antisense oligonucleotide (ASO) tethered to immunostimulatory TLR9 agonist (CpG oligonucleotide) to improve targeting of human and mouse prostate cancer and myeloid immune cells, such as myeloid-derived suppressor cells (MDSC). RESULTS: CpG-STAT3ASO conjugates showed improved biodistribution and potency of STAT3 knockdown in target cells in vitro and in vivo. Systemic administration of CpG-STAT3ASO (5 mg/kg) eradicated bone-localized, Ras/Myc-driven, and Ptenpc -/- Smad4pc -/- Trp53c -/- prostate tumors in the majority of treated mice. These antitumor effects were primarily immune-mediated and correlated with an increased ratio of CD8+ to regulatory T cells and reduced pSTAT3+/PD-L1+ MDSCs. Both innate and adaptive immunity contributed to systemic antitumor responses as verified by the depletion of Gr1+ myeloid cells and CD8+ and CD4+ T cells, respectively. Importantly, only the bifunctional CpG-STAT3ASO, but not control CpG oligonucleotides, STAT3ASO alone, or the coinjection of both oligonucleotides, succeeded in recruiting neutrophils and CD8+ T cells into tumors. Thus, the concurrence of TLR9 activation with STAT3 inhibition in the same cellular compartment is indispensable for overcoming tumor immune tolerance and effective antitumor immunity against prostate cancer. CONCLUSIONS: The bifunctional, immunostimulatory, and tolerance-breaking design of CpG-STAT3ASO offers a blueprint for the development of effective and safer oligonucleotide strategies for treatment of immunologically "cold" human cancers.