Purpose: Cancer tissue-specific and nuclei-targeted drug delivery is ideal for the delivery of chemotherapy. However, it has only been achieved in in vitro studies mainly due to low efficiency in vivo. In this study, we aimed to establish an efficient dual-targeted system that targets liver cancer tissue as well as the nuclei of cancer cells in vivo. Methods: We first synthesized TAT peptide (TATp)-mesoporous silica nanoparticle (MSN) complex (TATp-MSN) and generated liposomes that carried liver cancer-specific aptamer TLS11a (TLS11a-LB). We then generated the drug TLS11a-LB@TATp-MSN/doxorubicin (DOX) by mixing TLS11a-LB and DOX-loaded TATp-MSN. After physical and chemical characterization of the nanoparticles, DOX release from these formulations was evaluated at pH 5.0 and 7.4. Furthermore, we also evaluated nuclear localization and cytotoxicity of the drug in H22 cells in vitro and investigated the liver cancer targeting and antitumor activities of the nano-drug in vivo using a H22 tumor-bearing mice model. Results: TLS11a-LB@TATp-MSN/DOX and its controls were confirmed as nano-drugs (< 100 nm) using transmission electron microscopy (TEM). The DOX release rate of TLS11a-LB@TATp-MSN/DOX was significantly faster at pH 5.0 than at pH 7.4. TLS11a-LB@TATp-MSN/DOX effectively targeted the nuclei of H22 cells and released DOX with a higher efficiency than that of the control groups. In addition, TLS11a-LB@TATp-MSN/DOX exhibited slight cytotoxicity, but not significantly more than controls. In vivo studies showed that TLS11a-LB@TATp-MSN accumulated in subcutaneous H22 tumors in the right axilla of BALB/c mice, reaching peak levels at 48 h after intravenous injection, respectively, and demonstrated that TLS11a-LB@TATp-MSN/DOX group enhanced tumor treatment efficacy while reducing systemic side effects. Conclusion: TLS11a-LB@TATp-MSN/DOX can efficiently deliver DOX to the nuclei of liver cancer cells by dual targeting liver cancer tissue and the nuclei of the cancer cells in mice. Thus, it is a promising nano-drug for the treatment of liver cancer.

The liposome-based adjuvant AS01 incorporates two immune stimulants, 3-O-desacyl-4′-monophosphoryl lipid A and the saponin QS-21. AS01 is under investigation for use in several vaccines in clinical development. i.m. injection of AS01 enhances immune cell activation and dendritic cell (DC) Ag presentation in the local muscle-draining lymph node. However, cellular and Ag trafficking in the lymphatic vessels that connect an i.m. injection site with the local lymph node has not been investigated. The objectives of this study were: 1) to quantify the in vivo cellular immune response induced by AS01 in an outbred ovine model, 2) to develop a lymphatic cannulation model that directly collects lymphatic fluid draining the muscle, and 3) to investigate the function of immune cells entering and exiting the lymphatic compartments after s.c. or i.m. vaccination with AS01 administered with hepatitis B surface Ag (HBsAg). We show that HBsAg-AS01 induces a distinct immunogenic cellular signature within the blood and draining lymphatics following both immunization routes. We reveal that MHCII^high migratory DCs, neutrophils, and monocytes can acquire Ag within muscle and s.c. afferent lymph, and that HBsAg-AS01 uniquely induces the selective migration of Ag-positive neutrophils, monocytes, and an MHCII^high DC-like cell type out of the lymph node via the efferent lymphatics that may enhance Ag-specific immunity. We report the characterization of the immune response in the lymphatic network after i.m. and s.c. injection of a clinically relevant vaccine, all in real time using a dose and volume comparable with that administered in humans.

Determining the mechanisms involved in the immune-pathogenesis of the tropical liver fluke, Fasciola gigantica, is crucial to the development of any effective therapeutic intervention. Here, we examined the differential gene expression of cytokines and transcription factors in the liver of F. gigantica-infected buffaloes, over the course of infection. Water buffaloes (swamp type) were infected orally with 500 F. gigantica encysted metacercariae. Liver tissue samples were collected 3, 10, 28, 42, 70 and 98 days post-infection (dpi). Levels of gene expression of nine cytokines (IFN-γ, TGF-β, IL-1β, IL-4, IL-6, IL-10, IL-12B, IL-13 and IL-17A) and four transcription factors (T-bet, GATA-3, Foxp3 and ROR-γτ) were determined using quantitative real-time PCR (qRT-PCR). We evaluated any correlation between gene expression of these immune-regulatory factors and the severity of liver pathology. Histopathological examination revealed that cellular infiltration, hemorrhage and fibrosis without calcification in the liver parenchyma of infected buffaloes, increased over the course of infection. This progressive pathology was attributed to dysregulated and excessive inflammatory responses induced by infection. The early infection phase (3–10 dpi) was marked by a generalized immunosuppression and elevated TGF-β expression in order to facilitate parasite colonization. A mixed Th1/Th2 immune response was dominant from 28 to 70 dpi, to promote parasite survival while minimizing host tissue damage. During late infection (98 dpi), the response was biased towards Th1/Treg in order to inhibit the host’s Th2 protective response and promote chronic infection. Both IL-10 and IL-17A and the Th17/Treg balance, played key roles in mediating the inflammatory and immunoregulatory mechanisms in the liver during chronic fasciolosis. Our data showed distinct CD4+ T helper (Th) polarization and cytokine dysregulation in response to F. gigantica infection in water buffaloes over the course of infection. Characterizing the temporal expression profiles for host immune genes during infection should provide important information for defining how F. gigantica adapts and survives in the liver of buffaloes and how host immune responses influence F. gigantica pathogenicity.

Epimedii Folium (EF, Epimedium brevicornu Maxim.), a traditional botanical drug, is famous for treating bone fractures, joint diseases, and several chronic illnesses. However, some studies indicated that EF could induce idiosyncratic drug-induced liver injury (IDILI) in the clinic. The NLRP3 inflammasome plays a crucial role in the pathogenesis of various human diseases, including IDILI. In the present study, we showed that epimedin B could specifically facilitate nigericin- or ATP-induced NLRP3 inflammasome activation under synergistic induction of mitochondrial reactive oxygen species. Moreover, epimedin B resulted in activation of Caspase-1 and IL-1β secretion in a lipopolysaccharide (LPS)-mediated susceptibility mouse model. MCC950 pretreatment completely abrogated activation of the NLRP3 inflammasome and prevented liver injury. Importantly, several studies have confirmed that some active constituents of EF could enhance activation of the NLRP3 inflammasome and may be involved in the pathogenesis of EF-IDILI. No reports are available on whether the structure-activity relationship associated with the immunostimulatory activity in EF contributes to the pathogenesis of EF-IDILI. These findings have changed our conventional understanding about the more glycogen, the more immunostimulatory activity.

Background: The broader application of adoptive cell therapy (ACT) in cancer immunotherapies (particularly for solid tumors) has always been limited by the immunosuppressive tumor microenvironment (TME) and the insufficient targetability of effector T cells, resulting in unsatisfied therapeutic outcome. Here, we designed a new strategy by using aptamer-based immunoliposomes to modify PD-1-silencing T cells, which were activated by dendritic cell (DC)/tumor fusion cells (FCs) to improve the antitumor potency of cytotoxic T lymphocytes (CTLs/CD8⁺ T cells). Methods: PD-1 gene was knocked out from CD8⁺ T cells using CRISPR/Cas9 system to liberate T cell activity from immunosuppression. The PD-1⁻ T cells were stimulated with DC/tumor FCs, followed by further functional modification of tumor-specific nanoliposomes (hEnd-Apt/CD3-Lipo) to generate FC/PD-1⁻ CTLs. The activation and proliferation and specificity of the modified FC/PD-1⁻ CTLs were measured. The antitumor activity of these CTLs against HepG2-tumors was evaluated in xenograft NOD/SCID mice, and the antitumor mechanism was investigated based on tissue immunohistochemistry and serum ELISA. Results: Our results indicated that the modification of hEnd-Apt/CD3-Lipo nanocomposites on the FC/PD-1⁻ CTLs had a more substantial synergetic effect in inhibiting tumor growth and prolonging animal survival, rather than other control liposomes. Furthermore, the hEnd-Apt/CD3-Lipo-modified FC/PD-1⁻ CTLs showed a stronger antitumor outcome in the tumor-bearing mouse model, through the mechanisms of suppressing tumor cell proliferation, promoting tumor apoptosis, reducing angiogenesis but increasing the infiltration of the FC/PD-1⁻ CTLs in the tumor tissue, as well as upregulating the systemic levels of IFN-γ, IL-2, TNF-α and IL-6 cytokines, by comparison of the control settings. Conclusion: In sum, our investigation suggests an enhancement of antitumor effect by the surface modification of endoglin-targeting nanoliposomes upon DC/tumor FC-activated PD-1⁻ CTLs, therefore, provides a new tumoral endoglin-targeted approach as a promising strategy to reduce immunosuppression of tumor microenvironment and improve the immunotherapeutic outcome of anticancer ACT.