Inorganic Nanoparticles Change Cancer-Cell-Derived Extracellular Vesicle Secretion Levels and Cargo Composition, Resulting in Secondary Biological Effects.

Over the past decades, the medical exploitation of nanotechnology has been largely increasing and finding its way into translational research and clinical applications. Despite their biomedical potential, uncertainties persist regarding the intricate role that nanomaterials may play on altering physiology in healthy and diseased tissues. Extracellular vesicles (EVs) are recognized as an important pathway for intercellular communication and known to be mediators of cellular stress. EVs are currently explored for targeted delivery of therapeutic agents, including nanoformulations, to treat and diagnose cancer or other diseases. Here, we aimed to investigate whether nanomaterials could have a possible impact on EV functionality, their safety, and whether EVs can play a role in nanomaterial toxicity profiles. To evaluate this, the impact of inorganic nanomaterial administration on EVs derived from murine melanoma and human breast cancer cells was tested. Cells were incubated with subtoxic concentrations of 4 different biomedically relevant inorganic nanoparticles (NPs): gold, silver, silicon dioxide, or iron oxide. The results displayed a clear NP and cell-type-dependent effect on increasing or decreasing EV secretion. Furthermore, the expression pattern of several EV-derived miRNAs was significantly changed upon NP exposure, compared to nontreated cells. Detailed pathway analysis and additional studies confirmed that EVs obtained from NP-exposed cells could influence immunological responses and cellular physiology. Together, these data reveal that NPs can have wide-ranging effects which can result in toxicity concerns or enhanced therapeutic potential as a secondary enhanced effect mediated and enhanced by EVs.

The Efficacy of Nanoparticle Delivery to Hypoxic Solid Tumors by ciRGD Co-Administration Depends on Neuropilin-1 and Neutrophil Levels.

The ability to improve nanoparticle delivery to solid tumors is an actively studied domain, where various mechanisms are looked into. In previous work, the authors have looked into nanoparticle size, tumor vessel normalization, and disintegration, and here it is aimed to continue this work by performing an in-depth mechanistic study on the use of ciRGD peptide co-administration. Using a multiparametric approach, it is observed that ciRGD can improve nanoparticle delivery to the tumor itself, but also to tumor cells specifically better than vessel normalization strategies. The effect depends on the level of tumor perfusion, hypoxia, neutrophil levels, and vessel permeability. This work shows that upon characterizing tumors for these parameters, conditions can be selected that can optimally benefit from ciRGD co-administration as a means to improve NP delivery to solid tumors.

Cu-doped TiO2 nanoparticles improve local antitumor immune activation and optimize dendritic cell vaccine strategies.

Nanoparticle-mediated cancer immunotherapy holds great promise, but more efforts are needed to obtain nanoformulations that result in a full scale activation of innate and adaptive immune components that specifically target the tumors. We generated a series of copper-doped TiO2 nanoparticles in order to tune the kinetics and full extent of Cu2+ ion release from the remnant TiO2 nanocrystals. Fine-tuning nanoparticle properties resulted in a formulation of 33% Cu-doped TiO2 which enabled short-lived hyperactivation of dendritic cells and hereby promoted immunotherapy. The nanoparticles result in highly efficient activation of dendritic cells ex vivo, which upon transplantation in tumor bearing mice, exceeded the therapeutic outcomes obtained with classically stimulated dendritic cells. Efficacious but simple nanomaterials that can promote dendritic cancer cell vaccination strategies open up new avenues for improved immunotherapy and human health.

Gold nanoparticle delivery to solid tumors: a multiparametric study on particle size and the tumor microenvironment.

Nanoparticle (NP) delivery to solid tumors remains an actively studied field, where several recent studies have shed new insights into the underlying mechanisms and the still overall poor efficacy. In the present study, Au NPs of different sizes were used as model systems to address this topic, where delivery of the systemically administered NPs to the tumor as a whole or to tumor cells specifically was examined in view of a broad range of tumor-associated parameters. Using non-invasive imaging combined with histology, immunohistochemistry, single-cell spatial RNA expression and image-based single cell cytometry revealed a size-dependent complex interaction of multiple parameters that promoted tumor and tumor-cell specific NP delivery. Interestingly, the data show that most NPs are sequestered by tumor-associated macrophages and cancer-associated fibroblasts, while only few NPs reach the actual tumor cells. While perfusion is important, leaky blood vessels were found not to promote NP delivery, but rather that delivery efficacy correlated with the maturity level of tumor-associated blood vessels. In line with recent studies, we found that the presence of specialized endothelial cells, expressing high levels of CD276 and Plvap promoted both tumor delivery and tumor cell-specific delivery of NPs. This study identifies several parameters that can be used to determine the suitability of NP delivery to the tumor region or to tumor cells specifically, and enables personalized approaches for maximal delivery of nanoformulations to the targeted tumor.

CKT0353, a novel microtubule targeting agent, overcomes paclitaxel induced resistance in cancer cells.

Microtubule targeting agents (MTAs) are extensively used in cancer treatment and many have achieved substantial clinical success. In recent years, targeting microtubules to inhibit cell division has become a widespread pharmaceutical approach for treatment of various cancer types. Nevertheless, the development of multidrug resistance (MDR) in cancer remains a major obstacle for successful application of these agents. Herein, we provided the evidence that CKT0353, α-branched α,ß-unsaturated ketone, possesses the capacity to successfully evade the MDR phenotype as an MTA. CKT0353 induced G2/M phase arrest, delayed cell division via spindle assembly checkpoint activation, disrupted the mitotic spindle formation and depolymerized microtubules in human breast, cervix, and colorectal carcinoma cells. Molecular docking analysis revealed that CKT0353 binds at the nocodazole binding domain of ß-tubulin. Furthermore, CKT0353 triggered apoptosis via caspase-dependent mechanism. In addition, P-glycoprotein overexpressing colorectal carcinoma cells showed higher sensitivity to this agent when compared to their sensitive counterpart, demonstrating the ability of CKT0353 to overcome this classic MDR mechanism involved in resistance to various MTAs. Taken together, these findings suggest that CKT0353 is an excellent candidate for further optimization as a therapeutic agent against tumors with MDR phenotype.

Model-Based Nanoengineered Pharmacokinetics of Iron-Doped Copper Oxide for Nanomedical Applications.

The progress in nanomedicine (NM) using nanoparticles (NPs) is mainly based on drug carriers for the delivery of classical chemotherapeutics. As low NM delivery rates limit therapeutic efficacy, an entirely different approach was investigated. A homologous series of engineered CuO NPs was designed for dual purposes (carrier and drug) with a direct chemical composition-biological functionality relationship. Model-based dissolution kinetics of CuO NPs in the cellular interior at post-exposure conditions were controlled through Fe-doping for intra/extra cellular Cu2+ and biological outcome. Through controlled ion release and reactions taking place in the cellular interior, tumors could be treated selectively, in vitro and in vivo. Locally administered NPs enabled tumor cells apoptosis and stimulated systemic anti-cancer immune responses. We clearly show therapeutic effects without tumor cells relapse post-treatment with 6 % Fe-doped CuO NPs combined with myeloid-derived suppressor cell silencing.

Adaptive resistance to trastuzumab impairs response to neratinib and lapatinib through deregulation of cell death mechanisms.

Small molecule inhibitors (TKIs) of HER2 have demonstrated clinical benefit in HER2-positive breast tumors. One of them, lapatinib, is used once advanced tumors become refractory to the HER2 antibody trastuzumab. Another one, neratinib, has shown benefit in high-risk early-stage breast cancer after trastuzumab-based therapies. A common characteristic is that patients are formerly treated with trastuzumab. We have explored whether trastuzumab previous therapy affects its antitumoral action. Long time exposure of the HER2+ cell line BT474 to trastuzumab resulted in trastuzumab-insensitive cells (BTRH cells). While treatment of wild type BT474 cells with lapatinib or neratinib resulted in decreased viability, BTRH cells were resistant to the action of these TKIs. Analogous results were obtained using trastuzumab-resistant cells derived from a PDX. Functional transcriptomic analyses and biochemical studies demonstrated that the TKIs caused DNA damage and apoptosis in wild type cells, but not in BTRH. Moreover, previous treatment with trastuzumab impairs response to small TKIs, by eliminating their proapoptotic action. Moreover, actioning on the apoptotic machinery using a chemical library of proapoptotic compounds led to the identification of clinical-stage drugs that may be used to fight trastuzumab-TKI resistance.

TRAIL receptor activation overcomes resistance to trastuzumab in HER2 positive breast cancer cells.

The appearance of resistance to the anti-HER2 targeted drug trastuzumab constitutes, nowadays, an important challenge in the oncology clinic. To fight such resistance, we searched for potential vulnerabilities in cells resistant to that drug. To that end, we used cell lines primary resistant to trastuzumab, as well as cells made secondarily resistant to the drug upon continuous exposure. Using genomic and proteomic approaches, a deregulation in cell death pathways was identified in trastuzumab-resistant cells. More precisely, an increased response to the death factor TRAIL, caused by an increase in the cellular receptors for this factor, was observed. In parallel, a decrease in inhibitory components of the pathway was detected. This combination produces a more efficient assembly of the functional complex in the trastuzumab-resistant cells that translates in the observed increased response to TRAIL. Analysis of HER2 positive patient samples confirmed deregulation of this pathway in trastuzumab-resistant patients. Taken together our data identify a vulnerability of trastuzumab-resistant cells that could be used to design new targeted therapies in that context.

Resistance to the Antibody-Drug Conjugate T-DM1 Is Based in a Reduction in Lysosomal Proteolytic Activity.

Trastuzumab-emtansine (T-DM1) is an antibody-drug conjugate (ADC) that was approved recently to treat HER2+ breast cancers. Despite its impressive clinical efficacy in many patients, intrinsic and acquired resistance to T-DM1 has emerged as a challenge. To identify mechanisms of T-DM1 resistance, we isolated several resistant HER2+ clones exhibiting stable drug refractoriness in vitro and in vivo Genomic comparisons showed substantial differences among three of the isolated clones, indicating several potential mechanisms of resistance to T-DM1. However, we observed no differences in HER2 levels and signaling among the resistant models and parental HER2+ cells. Bioinformatics studies suggested that intracellular trafficking of T-DM1 could underlie resistance to T-DM1, and systematic analysis of the path followed by T-DM1 showed that the early steps in the internalization of the drug were unaltered. However, in some of the resistant clones, T-DM1 accumulated in lysosomes. In these clones, lysosomal pH was increased and the proteolytic activity of these organelles was deranged. These results were confirmed in T-DM1-resistant cells from patient-derived HER2+ samples. We postulate that resistance to T-DM1 occurs through multiple mechanisms, one of which is impaired lysosomal proteolytic activity. Because other ADC may use the same internalization-degradation pathway to deliver active payloads, strategies aimed at restoring lysosomal functionality might overcome resistance to ADC-based therapies and improve their effectiveness. Cancer Res; 77(17); 4639-51. ©2017 AACR.

DTA0100, dual topoisomerase II and microtubule inhibitor, evades paclitaxel resistance in P-glycoprotein overexpressing cancer cells.

The efficacy of microtubule targeting agents in cancer treatment has been compromised by the development of drug resistance that may involve both, P-glycoprotein overexpression and the changes in ß-tubulin isoforms' expression. The anti-Topoisomerase II activity of methyl 4-((E)-2-(methoxycarbonyl)vinyloxy)oct-2-ynoate (DTA0100) was recently reported. Herein, we further evaluated this propargylic enol ether derivative and found that it exerts inhibitory effect on tubulin polymerization by binding to colchicine binding site. DTA0100 mitotic arrest properties were investigated in two multi-drug resistant cancer cell lines with P-glycoprotein overexpression (colorectal carcinoma and glioblastoma). The sensitivity of multi-drug resistant cancer cell lines to DTA0100 was not significantly changed in contrast to microtubule targeting agents such as paclitaxel, vinblastine and colchicine. DTA0100 clearly induced microtubule depolymerization, leading to disturbance of cell cycle kinetics and subsequent apoptosis. The fine-tuning in ß-tubulin isoforms expression observed in multi-drug resistant cancer cells may influence the efficacy of DTA0100. Importantly, DTA0100 blocked the P-glycoprotein function in both multi-drug resistant cancer cell lines without inducing the increase in P-glycoprotein expression. Therefore, DTA0100 acting as dual inhibitor of Topoisomerase II and microtubule formation could be considered as multi-potent anticancer agent. Besides, it is able to overcome the problem of drug resistance that emerges in the therapeutic approaches with either Topoisomerase II or microtubule targeting agents.

Antiproliferative and quinone reductase-inducing activities of withanolides derivatives.

Two new and five known withanolides (jaborosalactones 2, 3, 4, 5, and 24) were isolated from the leaves of Jaborosa runcinata Lam. We also obtained some derivatives from jaborosalactone 5, which resulted to be the major isolated metabolite. The natural compounds as well as derivatives were evaluated for their antiproliferative activity and the induction of quinone reductase 1 (QR1; NQ01) activity. Structure-activity relationships revealed valuable information on the pharmacophore of withanolide-type compounds. Three compounds of this series showed significantly higher antiproliferative activity than jaborosalactone 5. The effect of these compounds on the cell cycle was determined. Furthermore, the ability of major compounds to induce QR1 was evaluated. It was found that all the active test compounds are monofunctional inducers that interact with Keap1. The most promising derivatives prepared from jaborosalactone 5 include (23R)-4ß,12ß,21-trihydroxy-1,22-dioxo-12,23-cycloergostan-2,5,17,24-tetraen-26,23-olide (18) and (23R)-21-acetoxy-12ß-hydroxy-1,22-dioxo-12,23-cycloergostan-2,5,17,24-tetraen-26,23-lactame (20).

QSAR on antiproliferative naphthoquinones based on a conformation-independent approach.

The antiproliferative activities of a series of 36 naphthoquinone derivatives were subjected to a Quantitative Structure-Activity Relationships (QSAR) study. For this purpose a panel of four human cancer cell lines was used, namely HBL-100 (breast), HeLa (cervix), SW-1573 (non-small cell lung) and WiDr (colon). A conformation-independent representation of the chemical structure was established in order to avoid leading with the scarce experimental information on X-ray crystal structure of the drug interaction. The 1179 theoretical descriptors derived with E-Dragon and Recon software were simultaneously analyzed through linear regression models based on the Replacement Method variable subset selection technique. The established models were validated and tested through the use of external test sets of compounds, the Leave-One-Out Cross Validation method, Y-Randomization and Applicability Domain analysis.

Molecular docking studies of the interaction between propargylic enol ethers and human DNA topoisomerase IIα.

Having identified a novel human DNA topoisomerase IIα (TOP2) catalytic inhibitor from a small and structure-focused library of propargylic enol ethers, we decided to analyze if the chirality of these compounds plays a determinant role in their antiproliferative activity. In this study, we describe for the first time the synthesis of the corresponding enantiomers and the biological evaluation against a panel of representative human solid tumor cell lines. Experimental results show that chirality does not influence the reported antiproliferative activity of these compounds. Docking studies of corresponding enantiomers against TOP2 reinforce the finding that the biological effect is not chiral-dependent and that these family of compounds seem to act as TOP2 catalytic inhibitors.

Novel clioquinol and its analogous platinum complexes: importance, role of the halogen substitution and the hydroxyl group of the ligand.

In this communication, we present the synthesis of new platinum complexes based on hydroxyquinoline ligands. We demonstrate the importance and the role of the halogen substitution as well as the chelation, which are essential structural characteristics for finding good cytotoxicities.

Expanding the synthesis of new trans-sulfonamide platinum complexes: cytotoxicity, SAR, fluorescent cell assays and stability studies.

In this manuscript, we describe the synthesis of new trans-N-sulfonamide platinum complexes and their antiproliferative activity (GI50, µM) in human solid tumors cells. The structure activity relationships (SAR), with different new synthesized complexes by variation in ligand, halogen and also in the stereochemistry of the ligand, has been studied. Solubility and stability studies have also been carried out as well as fluorescent cell assays in order to clarify the final target in the tumor cells.

Cytotoxic bioactivity of some phenylpropanoic acid derivatives.

In this study, we synthesized a series of phenylpropanoic acid derivatives based on modifications at four selected points of the molecular scaffold. The in vitro antiproliferative activities of the compounds were examined in representative human solid tumor cell lines. A SAR was established pointing out the relevance of the substituents. The best activity profiles were obtained for the derivatives bearing more lipophilic esters (GI50 3.1-21 microM).

Anti-inflammatory activity of a novel family of aryl ureas compounds in an endotoxin-induced airway epithelial cell injury model.

BACKGROUND: Despite our increased understanding of the mechanisms involved in acute lung injury (ALI) and the acute respiratory distress syndrome (ARDS), there is no specific pharmacological treatment of proven benefit. We used a novel screening methodology to examine potential anti-inflammatory effects of a small structure-focused library of synthetic carbamate and urea derivatives in a well established cell model of lipopolysaccharide (LPS)-induced ALI/ARDS. METHODOLOGY/PRINCIPAL FINDINGS: After a pilot study to develop an in vitro LPS-induced airway epithelial cell injury model, a library of synthetic carbamate and urea derivates was screened against representative panels of human solid tumor cell lines and bacterial and fungal strains. Molecules that were non-cytotoxic and were inactive in terms of antiproliferative and antimicrobial activities were selected to study the effects on LPS-induced inflammatory response in an in vitro cell culture model using A549 human alveolar and BEAS-2B human bronchial cells. These cells were exposed for 18 h to LPS obtained from Escherichia coli, either alone or in combination with the test compounds. The LPS antagonists rhein and emodin were used as reference compounds. The most active compound (CKT0103) was selected as the lead compound and the impact of CKT0103 on pro-inflammatory IL-6 and IL-8 cytokine levels, expression of toll-like receptor-4 (TLR4) and nuclear factor kappa B inhibitor alpha (IκBα) was measured. CKT0103 significantly inhibited the synthesis and release of IL-6 and IL-8 induced by LPS. This suppression was associated with inhibition of TLR4 up-regulation and IκBα down-regulation. Immunocytochemical staining for TLR4 and IκBα supported these findings. CONCLUSIONS/SIGNIFICANCE: Using a novel screening methodology, we identified a compound - CKT0103 - with potent anti-inflammatory effects. These findings suggest that CKT0103 is a potential target for the treatment of the acute phase of sepsis and sepsis-induced ALI/ARDS.

ß-Lapachone analogs with enhanced antiproliferative activity.

In this study, we describe the synthesis of a series of α- and ß-lapachone containing hydroxyl or methoxyl groups on the benzene ring, by means of the selective acid promoted cyclization of the appropriate lapachol analog. The evaluation of the antiproliferative activity in human solid tumor cell lines provided 7-hydroxy-ß-lapachone as lead with enhanced activity over the parent drug ß-lapachone. Cell cycle studies, protein expression experiments, and reactive oxygen species analysis revealed that, similarly to ß-lapachone, ROS formation and DNA damage are critical factors in the cellular toxicity of 7-hydroxy-ß-lapachone.

Antiproliferative activity of withanolide derivatives from Jaborosa cabrerae and Jaborosa reflexa. Chemotaxonomic considerations.

Three withanolides were isolated from the aerial parts of Jaborosa reflexa Phil. Jaborosa cabrerae Barboza yielded five sativolide withanolides (including jaborosalactones R, S, 38, and 39) and two trechonolide withanolides epimeric at C-23 (trechonolide A and jaborosalactone 32). In addition, five derivatives were obtained by chemical derivatization of jaborosalactone 38, and all compounds were fully characterized by 1D and 2D NMR spectroscopic studies. The in vitro antiproliferative activities of the major natural withanolides and the semisynthetic derivatives were examined against HBL-100, HeLa, SW1573, T-47D, and WiDr human solid tumor cancer cell lines. Some chemotaxonomic considerations are discussed.

In vitro synergistic interaction between DTA0100 and radiation in human cancer cell lines.

DTA0100 is a new catalytic inhibitor of the human DNA topoisomerase IIα that induces G2/M phase cell cycle arrest in human solid tumor cells lines from various malignancies. In our study, we investigated the effectiveness of the combined treatment of ionizing radiation with DTA0100 on the survival of three representative human solid tumor cell lines: HeLa (cervix), WiDr (colon) and SW1573 (non-small cell lung cancer). The concomitant treatment of DTA0100 and irradiation showed a synergistic and antagonistic effect in the three cell lines tested. A synergistic cytotoxic effect of the combination of DTA0100 and radiation was confirmed by the median drug effect analysis method. It was found that in those protocols where the drug was administered after radiation the most synergistic effect was achieved. Our study constitutes the first in vitro evidence for synergistic effects between DTA0100 and radiation. This combination therapy might thus be expected to be more effective than either treatment alone in patients with cervical, colon and non-small cell lung cancer cells.