Multifunctional LUV liposomes decorated for BBB and amyloid targeting - B. In vivo brain targeting potential in wild-type and APP/PS1 mice.

Multifunctional liposomes (mf-LIPs) having a curcumin-lipid ligand (to target amyloids) together with two ligands to target the transferrin, and the low-density apolipoprotein receptor of the blood-brain-barrier (BBB) on their surface, were previously studied (in vitro) as potential theranostic systems for Alzheimer's disease (AD) (Papadia et al., 2017, Eur. J. Pharm. Sciences; 101:140-148). Herein, the targeting potential of mf-LIPs was compared to that of BBB-LIPs (liposomes having only the two BBB-specific ligands) in FVB mice (normal), as well as in double transgenic mice (APP/PS1) and their corresponding littermates (WT), by live-animal (in vivo) and explanted organ (ex vivo) imaging. In FVB mice, the head-signals of mf-LIPs and BBB-LIPs are either similar, or signals from mf-LIP are higher, suggesting that the co-presence of the curcumin derivative on the liposome surface does not disturb the functionality of the BBB-specific ligands. Higher brain/liver+spleen ratios (ex vivo) were calculated post-injection of mf-LIP, compared to those found after BBB-LIP injection, due to the reduced distribution of mf-LIPs in the liver and spleen; showing that the curcumin ligand increases the stealth properties of liposomes by reducing their uptake by liver and spleen. The later effect is more pronounced when the density of the BBB-specific ligands on the mf-LIPs is 0.1mol%, compared to 0.2%, highlighting the importance of this parameter. When a high lipid dose (4mg/mouse) is injected in WT and APP/PS1 mice, the head-signals of mf-LIPs are significantly higher than those of BBB-LIPs, but no differences are observed between WT and APP/PS1 mice. However, after administration of a low liposome dose (0.05mg/mouse) of mf-LIPs, significant differences in the head-signals are found between WT and transgenic mice, highlighting the AD theranostic potential of the multifunctional liposomes, as well as the importance of the experimental parameters used in such in vivo screening studies

Multifunctional LUV liposomes decorated for BBB and amyloid targeting. A. In vitro proof-of-concept.

Multifunctional LUV liposomes (mf-LIPs) were developed, having a curcumin-lipid ligand (TREG) with affinity towards amyloid species, together with ligands to target the transferrin and the LDL receptors of the blood-brain-barrier (BBB), on their surface. mf-LIPs were evaluated for their brain targeting, on hCMEC/D3 monolayers, and for their ability to inhibit Aβ-peptide aggregation. The transport of mf-LIP across hCMEC/D3 monolayers was similar to that of BBB-LIPs, indicating that the presence of TREG on their surface does not reduce their brain targeting potential. Likewise, mf-LIP inhibitory effect on Aβ aggregation was similar to that of LIPs functionalized only with TREG, proving that the presence of brain targeting ligands does not reduce the functionality of the amyloid-specific ligand. Addition of the curcumin-lipid in some liposome types was found to enhance their integrity and reduce the effect of serum proteins on their interaction with brain endothelial cells. Finally, preliminary in vivo results confirm the in vitro findings. Concluding, the current results reveal the potential of the specific curcumin-lipid derivative as a component of multifunctional LIPs with efficient brain targeting capability, intended to act as a theragnostic system for AD

Geminin ablation in vivo enhances tumorigenesis through increased genomic instability

Geminin, a DNA replication licensing inhibitor, ensures faithful DNA replication in vertebrates. Several studies have shown that geminin depletion in vitro results in rereplication and DNA damage, whereas increased expression of geminin has been observed in human cancers. However, conditional inactivation of geminin during embryogenesis has not revealed any detectable DNA replication defects. In order to examine its role in vivo, we conditionally inactivated geminin in the murine colon and lung, and assessed chemically induced carcinogenesis. We show here that mice lacking geminin develop a significantly higher number of tumors and bear a larger tumor burden than sham-treated controls in urethane-induced lung and azoxymethane/dextran sodium sulfate-induced colon carcinogenesis. Survival is also significantly reduced in mice lacking geminin during lung carcinogenesis. A significant increase in the total number and grade of lesions (hyperplasias, adenomas, and carcinomas) was also confirmed by hematoxylin and eosin staining. Moreover, increased genomic aberrations, identified by increased ATR and γH2AX expression, was detected with immunohistochemistry analysis. In addition, we analyzed geminin expression in human colon cancer, and found increased expression, as well as a positive correlation with ATM/ATR levels and a non-monotonic association with γH2AX. Taken together, our data demonstrate that geminin acts as a tumor suppressor by safeguarding genome stability, whereas its overexpression is also associated with genomic instability. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd

Osteopontin drives KRAS-mutant lung adenocarcinoma.

Increased expression of osteopontin (secreted phosphoprotein 1, SPP1) is associated with aggressive human lung adenocarcinoma (LADC), but its function remains unknown. Our aim was to determine the role of SPP1 in smoking-induced LADC. We combined mouse models of tobacco carcinogen-induced LADC, of deficiency of endogenous Spp1 alleles, and of adoptive pulmonary macrophage reconstitution to map the expression of SPP1 and its receptors and determine its impact during carcinogenesis. Co-expression of Spp1 and mutant Kras(G12C) in benign cells was employed to investigate SPP1/KRAS interactions in oncogenesis. Finally, intratracheal adenovirus encoding Cre recombinase was delivered to LSL.KRAS(G12D) mice lacking endogenous or overexpressing transgenic Spp1 alleles. SPP1 was overexpressed in experimental and human LADC and portended poor survival. In response to two different smoke carcinogens, Spp1-deficient mice developed fewer and smaller LADC with decreased cellular survival and angiogenesis. Both lung epithelial- and macrophage-secreted SPP1 drove tumor-associated inflammation, while epithelial SPP1 promoted early tumorigenesis by fostering the survival of KRAS-mutated cells. Finally, loss and overexpression of Spp1 was, respectively, protective and deleterious for mice harboring KRAS(G12D)-driven LADC. Our data support that SPP1 is functionally involved in early stages of airway epithelial carcinogenesis driven by smoking and mutant KRAS and may present an important therapeutic target

Osteopontin drives KRAS-mutant lung adenocarcinoma

Increased expression of osteopontin (secreted phosphoprotein 1, SPP1) is associated with aggressive human lung adenocarcinoma (LADC), but its function remains unknown. Our aim was to determine the role of SPP1 in smoking-induced LADC. We combined mouse models of tobacco carcinogen-induced LADC, of deficiency of endogenous Spp1 alleles, and of adoptive pulmonary macrophage reconstitution to map the expression of SPP1 and its receptors and determine its impact during carcinogenesis. Co-expression of Spp1 and mutant KrasG12C in benign cells was employed to investigate SPP1/KRAS interactions in oncogenesis. Finally, intratracheal adenovirus encoding Cre recombinase was delivered to LSL.KRASG12D mice lacking endogenous or overexpressing transgenic Spp1 alleles. SPP1 was overexpressed in experimental and human LADC and portended poor survival. In response to two different smoke carcinogens, Spp1-deficient mice developed fewer and smaller LADC with decreased cellular survival and angiogenesis. Both lung epithelial- A nd macrophage-secreted SPP1 drove tumor-associated inflammation, while epithelial SPP1 promoted early tumorigenesis by fostering the survival of KRAS-mutated cells. Finally, loss and overexpression of Spp1 was, respectively, protective and deleterious for mice harboring KRASG12D-driven LADC. Our data support that SPP1 is functionally involved in early stages of airway epithelial carcinogenesis driven by smoking and mutant KRAS and may present an important therapeutic target. © 2019 The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please email: [email protected]

Club cells form lung adenocarcinomas and maintain the alveoli of adult mice

Lung cancer and chronic lung diseases impose major disease burdens worldwide and are caused by inhaled noxious agents including tobacco smoke. The cellular origins of environmental-induced lung tumors and of the dysfunctional airway and alveolar epithelial turnover observed with chronic lung diseases are unknown. To address this, we combined mouse models of genetic labeling and ablation of airway (club) and alveolar cells with exposure to environmental noxious and carcinogenic agents. Club cells are shown to survive KRAS mutations and to form lung tumors after tobacco carcinogen exposure. Increasing numbers of club cells are found in the alveoli with aging and after lung injury, but go undetected since they express alveolar proteins. Ablation of club cells prevents chemical lung tumors and causes alveolar destruction in adult mice. Hence club cells are important in alveolar maintenance and carcinogenesis and may be a therapeutic target against premalignancy and chronic lung disease. © eLife Sciences Publications Ltd. All rights reserved

Mutant <em>KRAS </em>promotes malignant pleural effusion formation.

Malignant pleural effusion (MPE) is the lethal consequence of various human cancers metastatic to the pleural cavity. However, the mechanisms responsible for the development of MPE are still obscure. Here we show that mutant KRAS is important for MPE induction in mice. Pleural disseminated, mutant KRAS bearing tumour cells upregulate and systemically release chemokine ligand 2 (CCL2) into the bloodstream to mobilize myeloid cells from the host bone marrow to the pleural space via the spleen. These cells promote MPE formation, as indicated by splenectomy and splenocyte restoration experiments. In addition, KRAS mutations are frequently detected in human MPE and cell lines isolated thereof, but are often lost during automated analyses, as indicated by manual versus automated examination of Sanger sequencing traces. Finally, the novel KRAS inhibitor deltarasin and a monoclonal antibody directed against CCL2 are equally effective against an experimental mouse model of MPE, a result that holds promise for future efficient therapies against the human condition