Nanoparticle Delivery of RIG-I Agonist Enables Effective and Safe Adjuvant Therapy in Pancreatic Cancer.

Local immunomodulation can be a promising strategy to augment the efficacy and decrease off-target toxicities associated with cancer treatment. Pancreatic cancer is resistant to immunotherapies due to the immunosuppressive tumor microenvironment. Herein, we investigated a therapeutic approach involving delivery of a short interfering double-stranded RNA (dsRNA), specific to Bcl2, with 5' triphosphate ends, by lipid calcium phosphate nanoparticles, in an orthotopic allograft KPC model of pancreatic cancer. Retinoic acid-inducible gene I (RIG-I)-like receptors can bind to 5' triphosphate dsRNA (ppp dsRNA), a pathogen-associated molecular pattern, producing type I interferon, while Bcl2 silencing can drive apoptosis of cancer cells. Our approach demonstrated a robust enrichment of tumor tissue with therapeutic nanoparticles and enabled a significant tumor growth inhibition, prolonging median overall survival. Nanoparticles encapsulating dual-therapeutic ppp dsRNA allowed strong induction in levels of pro-inflammatory Th1 cytokines, further increasing proportions of CD8+ T cells over regulatory T cells, M1 over M2 macrophages, and decreased levels of immunosuppressive B regulatory and plasma cells in the tumor microenvironment. Thus, these results provide a new immunotherapy approach for pancreatic cancer.

Protein corona-mediated targeting of nanocarriers to B cells allows redirection of allergic immune responses.

BACKGROUND: Nanoparticle (NP)-based vaccines are attractive immunotherapy tools because of their capability to codeliver antigen and adjuvant to antigen-presenting cells. Their cellular distribution and serum protein interaction ("protein corona") after systemic administration and their effect on the functional properties of NPs is poorly understood. OBJECTIVES: We analyzed the relevance of the protein corona on cell type-selective uptake of dextran-coated NPs and determined the outcome of vaccination with NPs that codeliver antigen and adjuvant in disease models of allergy. METHODS: The role of protein corona constituents for cellular binding/uptake of dextran-coated ferrous nanoparticles (DEX-NPs) was analyzed both in vitro and in vivo. DEX-NPs conjugated with the model antigen ovalbumin (OVA) and immunostimulatory CpG-rich oligodeoxynucleotides were administered to monitor the induction of cellular and humoral immune responses. Therapeutic effects of this DEX-NP vaccine in mouse models of OVA-induced anaphylaxis and allergic asthma were assessed. RESULTS: DEX-NPs triggered lectin-induced complement activation, yielding deposition of activated complement factor 3 on the DEX-NP surface. In the spleen DEX-NPs targeted predominantly B cells through complement receptors 1 and 2. The DEX-NP vaccine elicited much stronger OVA-specific IgG2a production than coadministered soluble OVA plus CpG oligodeoxynucleotides. B-cell binding of the DEX-NP vaccine was critical for IgG2a production. Treatment of OVA-sensitized mice with the DEX-NP vaccine prevented induction of anaphylactic shock and allergic asthma accompanied by IgE inhibition. CONCLUSIONS: Opsonization of lectin-coated NPs by activated complement components results in selective B-cell targeting. The intrinsic B-cell targeting property of lectin-coated NPs can be exploited for treatment of allergic immune responses.

Differences in risk behaviours and HIV/STI prevalence between low-fee and medium-fee female sex workers in three provinces in China.

OBJECTIVES: To better understand risk behaviours and factors associated with low-fee female sex workers (FSW) and support HIV/sexually transmitted infections (STI) epidemic control among this key population in China. METHODS: A cross-sectional study using convenience sampling to recruit 1487 eligible low-fee and medium-fee FSW was conducted in 2012 in three provinces. The participants were interviewed using a structured questionnaire and tested for HIV-1, herpes simplex virus (HSV)-2 and syphilis antibody. Log-binomial modelling was used to estimate prevalence ratios (PR) and examine factors associated with low-fee sex work. RESULTS: Prevalence of HIV-1, syphilis and HSV-2 antibody positive were 0.5%, 4.8% and 27.8%, respectively. Low-fee FSW were more likely to have HSV-2 infection (adjusted prevalence ratio (APR)=1.3, 95% CI 1.1 to 1.7), but not more likely to have HIV-1 and syphilis infection compared with medium-fee FSW. Compared with medium-fee FSW, low-fee FSW were more likely to be ≥35 years of age (APR=2.1, 95% CI 1.3 to 3.6), engage in sex work ≥6 days/per week (APR=1.7, 95% CI 1.2 to 2.6), have ≥3 clients per day (APR=2.2, 95% CI 1.5 to 3.3), have clients decide condom use (APR=1.6, 95% CI 1.1 to 2.3), fail to persuade clients to use condoms (APR=1.6, 95% CI 1.1 to 2.6), express willingness to have unprotected sex in return for receipt of a higher fee (APR=1.8, 95% CI 1.2 to 2.8), have had genital symptoms in the past year (APR=1.4, 95% CI 1.1 to 1.8) and have migrated from another city. CONCLUSIONS: Low-fee FSW in China have unique risks for acquiring HIV/STI, in part due to greater economic pressures. Tailored interventions targeting low-fee FSW and incorporating their prevailing perception of HIV/STI risks and condom use negotiation challenges that they face are urgently needed.

Dendritic cell motility and T cell activation requires regulation of Rho-cofilin signaling by the Rho-GTPase activating protein myosin IXb.

Directed migration of stimulated dendritic cells (DCs) to secondary lymphoid organs and their interaction with Ag-specific T cells is a prerequisite for the induction of primary immune responses. In this article, we show that murine DCs that lack myosin IXB (Myo9b), a motorized negative regulator of RhoA signaling, exhibit increased Rho signaling activity and downstream acto-myosin contractility, and inactivation of the Rho target protein cofilin, an actin-depolymerizing factor. On a functional level, Myo9b(-/-) DCs showed impaired directed migratory activity both in vitro and in vivo. Moreover, despite unaltered Ag presentation and costimulatory capabilities, Myo9b(-/-) DCs were poor T cell stimulators in vitro in a three-dimensional collagen matrix and in vivo, associated with altered DC-T cell contact dynamics and T cell polarization. Accordingly, Myo9b(-/-) mice showed an attenuated ear-swelling response in a model of contact hypersensitivity. The impaired migratory and T cell stimulatory capacity of Myo9b(-/-) DCs was restored in large part by pharmacological activation of cofilin. Taken together, these results identify Myo9b as a negative key regulator of the Rho/RhoA effector Rho-kinase [Rho-associated coiled-coil-forming kinase (ROCK)]/LIM domain kinase signaling pathway in DCs, which controls cofilin inactivation and myosin II activation and, therefore may control, in part, the induction of adaptive immune responses.

[Syphilis incidence and its risk factors in a cohort of young men who have sex with men].

OBJECTIVES: To investigate the incidence of syphilis infection and to determine the risk factors related to syphilis infection among young men who had sex with men (YMSM), which were documented for developing effective intervention to prevent sexually transmitted diseases among YMSM. METHODS: A cohort study was conducted in 8 cities (Beijing, Shanghai, Kunming,Guiyang, Chongqing, Chengdu, Urumqi and Nanning) from May to December, 2009. A total of 1 037 syphilis-negative YMSM aged 18-25 were enrolled in the cohort and the two follow-up surveys were carried out every six months. The contents of study included sociodemographic characteristics, HIV-related knowledge, sexual behavior and condom use in the 6 months prior to survey. All participants were tested for syphilis with whole blood specimens. Chi-square test was used to compare demographic characteristics of participants in baseline with those of two follow-up, and Cox regression analysis was used to identify risk factors associated with syphilis infection. RESULTS: The rates of participants in 6, 12 months follow-up surveys was 79.85% (828/1 037) and 82.16% (852/1 037) respectively.39 syphilis seroconversions were found in the 12 months follow-up survey. Cumulative observed person-years during follow-up time was 1 106.67. The syphilis incidence rate was 3.5%. The Multivariate Cox regression analysis showed that the education of senior high school (senior high school vs some college or higher, RR = 2.19, 95% CI:1.21-3.98), bisexual orientation (bisexual orientation vs homosexual orientation, RR = 2.19, 95% CI:1.21-3.97), score of HIV/AIDS knowledge <8 (score of HIV/AIDS knowledge <8 vs knowledge = 8, RR = 2.39, 95%CI:1.35-4.21), had two and more sexual partners and inconsistent condom use in the past 6 months (inconsistent condom use vs consistent condom use, RR = 3.10, 95% CI:1.39-6.91) were significantly associated with syphilis seroconversion in the 12-month period. CONCLUSIONS: The syphilis incidence was high and risk behaviors were common among YMSM of China.

Free PEG Suppresses Anaphylaxis to PEGylated Nanomedicine in Swine.

Covalent conjugation of poly(ethylene glycol) (PEG) is frequently employed to enhance the pharmacokinetics and biodistribution of various protein and nanoparticle therapeutics. Unfortunately, some PEGylated drugs can induce elevated levels of antibodies that can bind PEG, i.e., anti-PEG antibodies (APA), in some patients. APA in turn can reduce the efficacy and increase the risks of allergic reactions, including anaphylaxis. There is currently no intervention available in the clinic that specifically mitigates allergic reactions to PEGylated drugs without the use of broad immunosuppression. We previously showed that infusion of high molecular weight free PEG could safely and effectively suppress the induction of APA in mice and restore prolonged circulation of various PEGylated therapeutics. Here, we explored the effectiveness of free PEG as a prophylaxis against anaphylaxis induced by PEG-specific allergic reactions in swine. Injection of PEG-liposomes (PL) resulted in anaphylactoid shock (pseudoanaphylaxis) within 1-3 min in both naïve and PL-sensitized swine. In contrast, repeated injection of free PEG alone did not result in allergic reactions, and injection of free PEG effectively suppressed allergic reactions to PL, including in previously PL-sensitized swine. These results strongly support the further investigation of free PEG for reducing APA and allergic responses to PEGylated therapeutics.

[Effects of Corbrin Shugan capsule on dimethylnitrosamine-induced hepatic fibrosis in rats].

OBJECTIVE: To investigate the effects of Corbrin Shugan capsule on dimethylnitrosamine (DMN)-induced hepatic fibrosis in rats. METHODS: Hepatic fibrosis was induced by DMN in AD rats. The serum concentrations of III pro-collagen (III PC),laminin (LN) and tissue inhibitor of metalloproteinase-1(TIMP-1) were determined with ELISA. The concentration of albumin (ALB) in sera and the content of hydroxyproline (Hyp) in liver tissues were determined with chemical colorimetric and HPLC, respectively. The fibrosis area was measured with Motic Med 6.0 digital medical image analysis system. RESULTS: Compared to model group the high-dose (450 mg kg(-1)),mid-dose (270 mg kg(-1)) and low-dose (90 mg kg(-1)) groups of Corbrin Shugan capsule had significantly lower serum content of III PC [34.46 ± 13.95),(36.15 ± 9.46), and (40.58 ± 7.72)ng ml(-1) compared with (49.38 ± 10.95)ng ml(-1),P<0.05 or P<0.01],TIMP-1 [(16.65 ± 4.24),(16.66 ± 4.34),and (18.99 ± 6.05)ng ml(-1) compared with (30.84 ± 14.48)ng ml(-1), P<0.05 or P<0.01], LN [(12.94 ± 4.29), (12.96 ± 3.21),and (15.32 ± 8.00)ng ml(-1) compared with (30.22 ± 17.00)ng ml(-1),P<0.05 or P<0.01] and smaller hepatic fibrosis area [(0.02240 ± 0.01337), (0.02176 ± 0.01460) and (0.02384 ± 0.01405)µm(2) compared with vs (0.03929 ± 0.01732)µm2, P<0.05 or P<0.01]; the high-dose and mid-dose groups of Corbrin Shugan capsule had significantly lower content of Hyp in liver tissues [(0.77 ± 0.09) and (0.81 ± 0.09)µg µmg(-1) compared with (1.06 ± 0.33)µg mg(-1),P<0.05 or P<0.01]; and the high-dose group of Corbrin Shugan capsule significantly increased the content of ALB in sera [(34.02 ± 4.17)g L(-1) compared with (30.25 ± 4.21)g L(-1),P<0.05]. CONCLUSION: Corbrin Shugan capsule is effective in treatment of DMN-induced hepatic fibrosis in rats.

Pegloticase co-administered with high MW polyethylene glycol effectively reduces PEG-immunogenicity and restores prolonged circulation in mouse.

The interactions between polymers and the immune system remains poorly controlled. In some instances, the immune system can produce antibodies specific to polymer constituents. Indeed, roughly half of pegloticase patients without immunomodulation develop high titers of anti-PEG antibodies (APA) to the PEG polymers on pegloticase, which then quickly clear the drug from circulation and render the gout treatment ineffective. Here, using pegloticase as a model drug, we show that addition of high molecular weight (MW) free (unconjugated) PEG to pegloticase allows us to control the immunogenicity and mitigates APA induction in mice. Compared to pegloticase mixed with saline, mice repeatedly dosed with pegloticase containing different MW or amount of free PEG possessed 4- to 12- fold lower anti-PEG IgG, and 6- to 10- fold lower anti-PEG IgM, after 3 rounds of pegloticase dosed every 2 weeks. The markedly reduced APA levels, together with competitive inhibition by free PEG, restored the prolonged circulation of pegloticase to levels observed in APA-naïve animals. In contrast, mice with pegloticase-induced APA eliminated nearly all pegloticase from the circulation within just four hours post-injection. These results support the growing literature demonstrating free PEG may effectively suppress drug-induced APA, which in turn may offer sustained therapeutic benefits without requiring broad immunomodulation. We also showed free PEG effectively blocked the PEGylated protein from binding with cells expressing PEG-specific B cell receptors. It provides a template of how we may be able to tune the interactions and immunogenicity of other polymer-modified therapeutics. STATEMENT OF SIGNIFICANCE: A major challenge with engineering materials for drug delivery is their interactions with the immune system. For instance, our body can produce high levels of anti-PEG antibodies (APA). Unfortunately, the field currently lack tools to limit immunostimulation or overcome pre-existing anti-PEG antibodies, without using broad immunosuppression. Here, we showed that simply introducing free PEG into a clinical formulation of PEG-uricase can effectively limit induction of anti-PEG antibodies, and restore their prolonged circulation upon repeated dosing. Our work offers a readily translatable method to safely and effectively restore the use PEG-drugs in patients with PEG-immunity, and provides a template to use unconjugated polymers with low immunogenicity to regulate interactions with the immune system for other polymer-modified therapeutics.

[Corbrin shugan capsule for treatment of alcoholic hepatic fibrosis in rats].

OBJECTIVE: To investigate the therapeutic effect of Corbrin shugan capsule for treatment of alcoholic hepatic fibrosis in rats. METHODS: The rat model of alcoholic hepatic fibrosis was induced by intragastric administration of alcohol repeatedly. The serum procollagen III (PC III), laminin (LN) and tissue inhibitors of metalloproteinase-1 (TIMP-1) levels were measured with ELISA, and the content of hydroxyproline (Hyp) in liver tissue were determined with colorimetric method. Collagen deposition in liver tissue was observed with Masson's staining, and the fibrosis area was measured with digital medical image analysis system (Motic Med 6.0). RESULTS: Compared with the model control group, the serum TIMP-1 and LN levels and hepatic fibrosis area in liver tissue significantly decreased in Corbrin shugan capsule groups with doses of 0.09,0.27 and 0.45 g*kg(-1), and the serum PC III and the Hyp contents in liver tissue also decreased of Corbrin shugan capsule groups with doses of 0.27 and 0.45g*kg(-1). CONCLUSION: Corbrin shugan capsule can decrease serum PC III, TIMP-1 and LN levels and Hyp levels in liver tissue and hepatic fibrosis area in rats, indicating it may have therapeutic effect on alcoholic hepatic fibrosis.

Optimization of Interface Materials between Bi2Te3-Based Films and Cu Electrodes Enables a High Performance Thin-Film Thermoelectric Cooler.

Thermoelectric interface materials (TEiMs) are key to optimizing the electrical contact and stability of the interface between thermoelectric material and metal electrode in high-performance thin-film thermoelectric coolers (TECs). Herein, we explored TEiMs applicable to representative Bi-Te films and found that Cr and Ag are effective TEiMs for p-type Bi0.5Sb1.5Te3 and n-type Bi2Te3, respectively. By introducing 200 nm Cr and 200 nm Ag as TEiMs for p-type Bi0.5Sb1.5Te3/Cu and n-type Bi2Te3/Cu interfaces, Cu diffusion is suppressed, and excellent electrical contact is achieved (1.81 × 10-12 Ω m2 for p-type and 3.32 × 10-12 Ω m2 for n-type) and remains stable after heat treatment (2.37 × 10-12 Ω m2 for p-type and 1.63 × 10-12 Ω m2 for n-type). Furthermore, the cooling flux of TECs with optimized TEiMs increases from 122.74 to 296.56 W/cm2, while the performance degradation caused by contact resistance decreases from 50.81 to 4.15%. In addition, our results show that diffusion occurs between not only Cu but also Ag and the thermoelectric material, as TEiMs diffuse slightly. The diffusion of Cu and Ag at the interface can optimize the electrical contact of Bi2Te3/Cu but strongly degrade the electrical contacts of Bi0.5Sb1.5Te3/Cu. Our work provides an optimal selection of TEiMs for high-performance Bi-Te thin film coolers and provides guidance for further miniaturization of devices.

Nano-trapping CXCL13 reduces regulatory B cells in tumor microenvironment and inhibits tumor growth.

Interactions between different cell types in the tumor microenvironment (TME) affect tumor growth. Tumor-associated fibroblasts produce C-X-C motif chemokine ligand 13 (CXCL13) which recruits B cells to the TME. B-cells in the TME differentiate into regulatory B cells (Bregs) (IL-10+CD1d+CD5+CD138+CD19+). We highlight these Breg cells as a new important factor in the modulation of the immunosuppressive TME in different desmoplastic murine tumor models. In addition, CXCL13 also stimulates epithelial-mesenchymal transition (EMT) of the tumor cells. The tumorigenic roles of CXCL13 led us to explore an innovative anti-cancer strategy based on delivering plasmid DNA encoding a CXCL13 trap to reduce Bregs differentiation and normalize EMT, thereby suppressing tumor growth. CXCL13 trap suppressed tumor growth in pancreatic cancer, BRAF-mutant melanoma, and triple-negative breast cancer. In this study, following treatment, the affected tumor remained dormant resulting in prolonged progression-free survival of the host.

Celastrol nanoemulsion induces immunogenicity and downregulates PD-L1 to boost abscopal effect in melanoma therapy.

Programmed cell death-ligand 1 (PD-L1)-based immune checkpoint blockade therapy using the anti-PD-L1 antibody is effective for a subset of patients with advanced metastatic melanoma but about half of the patients do not respond to the therapy because of the tumor immunosuppressive microenvironment. Immunogenic cell death (ICD) induced by cytotoxins such as doxorubicin (DOX) allows damaged dying tumor cells to release immunostimulatory danger signals to activate dendritic cells (DCs) and T-cells; however, DOX also makes tumor cells upregulate PD-L1 expression and thus deactivate T-cells via the PD-1/PD-L1 pathway. Herein, we show that celastrol (CEL) induced not only strong ICD but also downregulation of PD-L1 expression of tumor cells. Thus, CEL was able to simultaneously activate DCs and T-cells and interrupt the PD-1/PD-L1 pathway between T-cells and tumor cells. In a bilateral tumor model, intratumorally (i.t.) injected celastrol nanoemulsion retaining a high tumor CEL concentration activated the immune system efficiently, which inhibited both the treated tumor and the distant untreated tumor in the mice (i.e., abscopal effect). Thus, this work demonstrates a new and much cost-effective immunotherapy strategy - chemotherapy-induced immunotherapy against melanoma without the need for expensive immune-checkpoint inhibitors.

Pre-treatment with high molecular weight free PEG effectively suppresses anti-PEG antibody induction by PEG-liposomes in mice.

Immune responses against polyethylene glycol (PEG) can lead to the rapid clearance of PEGylated drugs and are associated with increased risk of serious adverse events such as infusion reactions and anaphylaxis. Although select PEGylated therapeutics can induce anti-PEG antibodies (APA), there is currently no readily deployable strategy to mitigate their negative effects. Given the large number of PEGylated therapeutics that are either FDA-approved or in clinical development, methods that suppress APA induction to ensure the safety and efficacy of PEGylated drugs in patients would be a valuable clinical tool. We previously showed that infusion of high molecular weight (MW) free PEG can safely and effectively restore the circulation of PEG liposomes in animals with high pre-existing titers of APA, without stimulating additional APA production. Here, we explored the effectiveness of prophylaxis with free PEG or tolerogenic PEGylated liposomes as a strategy to reduce the amount of APA induced by subsequently administered PEGylated liposomes. Surprisingly, we found that a single administration of free PEG alone was capable of markedly reducing the APA response to PEG-liposomes for ~2 months; the effectiveness was comparable to, and frequently exceeded, interventions with different tolerogenic PEG-liposomes. These results support further investigations of free PEG prophylaxis as a potential strategy to ameliorate the APA response to sensitizing PEGylated therapeutics.

[Impact of methadone maintenance treatment on HIV risk behaviors of heroin drug users].

OBJECTIVE: To evaluate HIV risk behaviors among heroin drug users who were treated in methadone maintenance treatment (MMT) clinics. METHODS: A prospective cohort study recruited and followed up clients of eight MMT clinics treated no more than two and half months in Guizhou province, China. Through face-to-face interviews, the baseline and following up informations were collected. And the baseline information included both demographic information and HIV risk behaviors, the following up informations included only HIV risk behaviors. The baseline investigation started in June, 2006 and the following up investigation finished in June, 2007. A total of 1003 heroin drug users were recruited at baseline, among them 666 (66.4%) were still at treatment by the end of follow up and 469 (70.4%) clients participated in the follow up interview. Wilcoxon two sample test and McNemar test were used to test for changes in HIV risk behaviors between baseline and following up investigation. RESULTS: Among the 469 MMT clients who were followed up, the average days of reported heroin use decreased from 26.4 to 0.9 in the past 30 days (Z = 27.21, P < 0.05). Average days of alcohol use at baseline were 3.3 but 3.7 at follow up (Z = 0.45, P = 0.96). Needle-sharing behavior reported in the past 30 days decreased from 1.3% at baseline to 0.2% at follow up (χ(2) = 5.00, P = 0.025). At baseline, 5.5% (26/469) subjects reported having multiple sex partners in the past 30 days compared to 3.4% (16/469) at following up (χ(2) = 3.18, P = 0.08).6.4% (30/469) subjects reported casual sex with non-regular sex partners in the past 30 days at baseline compared to 5.1% (24/469) at following up (χ(2) = 0.95, P = 0.33). Of those who reported having casual sex relationship in the past 30 days 56.7% (17/30) reported using condoms at baseline but 58.3% (14/24) reported using condoms at follow up (χ(2) = 1.96, P = 0.16). CONCLUSION: MMT was observed to decrease needle-sharing HIV risk behavior. However, decreased HIV sexual risk behaviors were not observed at statistical significant level.

Nanocarrier-mediated immunogenic chemotherapy for triple negative breast cancer.

Triple negative breast cancer (TNBC) does not respond to checkpoint blockade immunotherapy as a result of immunosuppressive tumor microenvironment. To remodel the tumor microenvironment, we developed a liposome formulation to deliver a potential immunogenic cell death (ICD) inducing agent, 17-(allylamino)-17-demethoxygeldanamycin (17-AAG, or tanespimycin), in a tumor targeted manner to reverse the immunosuppressive microenvironment and facilitate the checkpoint blockade immunotherapy. The 17-AAG liposomes was prepared by thin film dispersion methods. The orthotopic 4T1 murine triple negative breast cancer model was studied. 17-AAG delivered by liposome remodeled the immunosuppressive microenvironment, significantly increased tumor infiltrating T cells, lowered the hypoxia level, decreased the suppressive lymphocytes such as tumor associated macrophages and myeloid derived suppressor cells in the tumor microenvironment. In addition, real-time PCR analysis revealed that chemokines and cytokines with immunosuppressive properties were notably reduced, which further facilitated the T cell mediated immunotherapy. Despite the fact that low dose 17-AAG liposomes demonstrated a limited therapeutic effect alone on 4T1 tumor, promising efficacy was observed when 17-AAG liposomes combined with checkpoint blockade immunotherapy. Taken together, 17-AAG liposomes could remodel the immunosuppressive microenvironment of triple negative breast cancer and facilitate the checkpoint blockade immunotherapy.

Genotyping of Mycobacterium leprae for understanding the distribution and transmission of leprosy in endemic provinces of China.

OBJECTIVES: Understanding the nature of Mycobacterium leprae transmission is vital to implement better control strategies for leprosy elimination. The present study expands the knowledge of county-level strain diversity, distribution, and transmission patterns of leprosy in endemic provinces of China. METHODS: We genetically characterized 290 clinical isolates of M. leprae from four endemic provinces using variable number tandem repeats (VNTR) and single nucleotide polymorphisms (SNPs). Attained genetic profiles and cluster consequences were contrasted with geographical and migration features of leprosy at county levels. RESULTS: Considering the allelic variability of 17 VNTR loci by the discriminatory index, (GTA)9, (AT)17, (AT)15, (TA)18, (TTC)21, and (TA)10 are reported to be more highly polymorphic than other loci. The VNTR profile generated the low-density clustering pattern in the counties of Sichuan and Yunnan, whereas clusters have been observed from the isolates from Huayuan (N = 6), Yongding (N = 3), Zixing (N = 3), Chenxi (N = 2) and Zhongfang (N = 2) counties of Hunan, and Zhijin (N = 3), Anlong (N = 2), Zhenning (N = 2), and Xixiu (N = 2) counties of Guizhou. In some clusters, people's social relations have been observed between villages. From the 290 clinical isolates, the most predominantly reported SNP was 3K (278, 95.8%), followed by SNP 1D (10, 3.4%), which are typically observed to be predominant in China. We also detected the novel SNP 3J (2, 0.8%), which has not yet been reported in China. CONCLUSION: The clustering pattern of M. leprae indicates the transmission of leprosy still persists at county levels, suggesting that there is a need to implement better approaches for tracing the close contacts of leprosy patients.

Inhibiting PI3 kinase-γ in both myeloid and plasma cells remodels the suppressive tumor microenvironment in desmoplastic tumors.

Phosphoinositide-3-kinases (PI3Ks) are part of signal transducing enzymes that mediate key cellular functions in cancer and immunity. PI3K-γ is crucial for cellular activation and migration in response to certain chemokines. PI3K-γ is highly expressed in myeloid cells and promotes their migration and the production of inflammatory mediators. We found that PI3K-γ was also highly expressed in tumor-associated B cells. IPI-549, the only PI3K-γ inhibitor in clinical development, offers a unique approach to enhance the anti-tumor immune response. We encapsulated IPI-549 in targeted polymeric nanoparticles (NP) and tested its activity in both murine pancreatic cancer and melanoma models. IPI-549 NP significantly decreased tumor growth and prolonged host survival in both models. Importantly, IPI-549 NP treatment reduced the suppressive tumor microenvironment by decreasing both suppressive myeloid and plasma cells in the tumor. We concluded that IPI-549 NP delivery could be a promising method for treating pancreatic cancer and other immune-suppressive tumors.

Nanoformulated Codelivery of Quercetin and Alantolactone Promotes an Antitumor Response through Synergistic Immunogenic Cell Death for Microsatellite-Stable Colorectal Cancer.

Microsatellite-stable colorectal cancer (CRC) is known to be resistant to immunotherapy. The combination of quercetin (Q) and alantolactone (A) was found to induce synergistic immunogenic cell death (ICD) at a molar ratio of 1:4 (Q:A). To achieve ratiometric loading and delivery, the micellar delivery of Q and A (QA-M) was developed with high entrapment efficiency and drug loading at an optimal ratio. QA-M achieved prolonged blood circulation and increased tumor accumulation for both drugs. More importantly, QA-M retained the desired drug ratio (molar ratio of Q to A = 1:4) in tumors at 2 and 4 h after intravenous injection for synergistic immunotherapy. Tumor growth was significantly inhibited in murine orthotopic CRC by the treatment of QA-M compared to PBS and the combination of free drugs (p < 0.005). The combination of nanotherapy stimulated the host immune response to induce long-term tumor destruction and induced memory tumor surveillance with a 1.3-fold increase in survival median time compared to PBS (p < 0.0001) and a combination of free drugs (p < 0.0005). The synergistic therapeutic effect induced by codelivery of Q and A is capable of reactivating antitumor immunity by inducing ICD, causing cell toxicity and modulating the immune-suppressive tumor microenvironment. Such a combination of Q and A with synergistic effects entrapped in a simple and safe nanodelivery system may provide the potential for scale-up manufacturing and clinical applications as immunotherapeutic agents for CRC.

Vasodilator Hydralazine Promotes Nanoparticle Penetration in Advanced Desmoplastic Tumors.

Desmoplastic tumors are normally resistant to nanoparticle-based chemotherapy due to dense stroma and limited particle permeability inside the tumor. Herein, we reported that hydralazine (HDZ)-an antihypertension vasodilator-would dramatically promote nanoparticle penetration in advanced desmoplastic tumors. First, a HDZ-liposome system was developed for tumor-selective delivery of HDZ. After three injections of HDZ-liposomes at a dose of 15 mg/kg, the tumor stroma was remarkably reduced, along with ameliorated tumor hypoxia in murine models of desmoplastic melanoma (BPD6). Furthermore, HDZ-liposome treatment altered the immunosuppressive tumor microenvironment, which provided opportunities for applying this therapeutic system to aid immunotherapy in desmoplastic tumors. Using DiD-loaded liposome as a model nanoparticle, we showed that HDZ-liposome treatment significantly increased nanoparticle accumulation and penetration inside desmoplastic tumors. As a result, one single injection of doxorubicin-liposomes at a dose of 5 mg/kg resulted in strong tumor inhibition effect after HDZ-liposome pretreatment in the advanced desmoplastic melanoma with sizes over 400 mm3. Because HDZ is a widely used antihypertension drug, the findings here should be readily translatable for clinical benefits.

Molecular surveillance of antimicrobial resistance and transmission pattern of Mycobacterium leprae in Chinese leprosy patients.

Reports on antimicrobial resistance (AMR) of Mycobacterium leprae, relationship with bacteriological index (BI), and transmission in China are limited. We investigated the emergence of AMR mutations, the relationship between BI and AMR in complete, moderate and lack of BI decline cases, and molecular epidemiological features of AMR cases by enrolling 290 leprosy cases from four endemic provinces. Seven (2.41%), one (0.34%), five (1.72%), one (0.34%), and one (0.34%) strains had single mutations in folP1, rpoC, gyrA, gyrB, and 23S rRNA, respectively. Double mutations in folP1 and gyrA, rpoB and gyrA, and gyrA and 23S rRNA were observed in one (0.34%) strain each. Mutated strains occurred in three out of 81 (95% CI-0.005-0.079, p = 0.083) cases with complete BI decline, in seven out of 103 (95% CI 0.018-0.117, p = 0.008) cases with moderate BI decline, and in four out of 34 (95% CI 0.003-0.231, p = 0.044) cases with lack of BI decline. Most of these mutated strains were geographically separated and diverged genotypically. AMR mutations may not be the main cause of the lack of BI decline. The low transmission of AMR strains at the county level indicates an ongoing transmission at close contact levels.