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1.
J Med Chem ; 65(6): 4783-4797, 2022 03 24.
Article in English | MEDLINE | ID: mdl-35290041

ABSTRACT

A set of meta-substituted 3-arylisoquinolinones have been identified that show substantial cytotoxicity in breast, liver, lung and colon cancer cell lines; these are up to 700-fold more active than the corresponding para analogues. These compounds were initially proposed as inhibitors of N-ribosyl dihydronicotinamide (NRH): quinone oxidoreductase 2 (NQO2) but were found to be inactive against the enzyme. Instead, COMPARE analysis suggested that 6-fluoro-3-(meta-fluorophenyl)isoquinolin-1(2H)-one (4) could mimic colchicine and interact with microtubules, a recognized target for cancer therapy. Subsequent docking, molecular dynamics simulations, and free energy analysis further suggested that compound 4 bound well into the colchicine-binding pocket of tubulin. Indeed, 4 suppressed tubulin polymerization, caused G2/M cell cycle arrest, and induced apoptosis. Also, 4 inhibited the formation of endothelial cell capillary-like tubes and further disrupted the structure of preestablished tubes; the effects were not observed with para analogue 5. In accordance with this, the computed free energy of binding of 5 to tubulin was lower in magnitude than that for 4 and appeared to arise in part from the inability of the para substituent to occupy a tubulin subpocket, which is possible in the meta orientation. In conclusion, the antiproliferative potential of the novel 3-arylisoquinolinones is markedly influenced by a subtle change in the structure (meta versus para). The meta-substituted isoquinolinone 4 is a microtubule-destabilizing agent with potential tumor-selectivity and antiangiogenic and vascular disrupting features.


Subject(s)
Antineoplastic Agents , Tubulin , Antineoplastic Agents/chemistry , Cell Line, Tumor , Cell Proliferation , Colchicine/metabolism , Drug Screening Assays, Antitumor , Microtubules , Molecular Structure , Structure-Activity Relationship , Tubulin/metabolism , Tubulin Modulators/chemistry
2.
Br J Cancer ; 122(1): 62-71, 2020 01.
Article in English | MEDLINE | ID: mdl-31819173

ABSTRACT

BACKGROUND: High expression of Ki67, a proliferation marker, is associated with reduced endometrial cancer-specific survival. Pre-surgical metformin reduces tumour Ki-67 expression in some women with endometrial cancer. Metformin's anti-cancer activity may relate to effects on cellular energy metabolism. Since tumour hypoxia and glucose availability are major cellular redox determinants, we evaluated their role in endometrial cancer response to metformin. METHODS: Endometrial cancer biopsies from women treated with pre-surgical metformin were tested for the hypoxia markers, HIF-1α and CA-9. Endometrial cancer cell lines were treated with metformin in variable glucose concentrations in normoxia or hypoxia and cell viability, mitochondrial biogenesis, function and energy metabolism were assessed. RESULTS: In women treated with metformin (n = 28), Ki-67 response was lower in hypoxic tumours. Metformin showed minimal cytostatic effects towards Ishikawa and HEC1A cells in conventional medium (25 mM glucose). In low glucose (5.5 mM), a dose-dependent cytostatic effect was observed in normoxia but attenuated in hypoxia. Tumours treated with metformin showed increased mitochondrial mass (n = 25), while in cultured cells metformin decreased mitochondrial function. Metformin targets mitochondrial respiration, however, in hypoxic, high glucose conditions, there was a switch to glycolytic metabolism and decreased metformin response. CONCLUSIONS: Understanding the metabolic adaptations of endometrial tumours may identify patients likely to derive clinical benefit from metformin.


Subject(s)
Cell Hypoxia/drug effects , Cytostatic Agents/therapeutic use , Endometrial Neoplasms/drug therapy , Endometrial Neoplasms/metabolism , Hyperglycemia/metabolism , Hypoglycemic Agents/therapeutic use , Metformin/therapeutic use , Antigens, Neoplasm/genetics , Antigens, Neoplasm/metabolism , Carbonic Anhydrase IX/genetics , Carbonic Anhydrase IX/metabolism , Cell Hypoxia/genetics , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Survival/drug effects , Cytostatic Agents/administration & dosage , Endometrial Neoplasms/pathology , Female , Gene Expression Regulation, Neoplastic , Glucose/metabolism , Humans , Hypoglycemic Agents/administration & dosage , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Ki-67 Antigen/metabolism , Metformin/administration & dosage , Metformin/adverse effects , Mitochondria/drug effects , Mitochondria/metabolism , Preoperative Care/methods , Signal Transduction/drug effects , Treatment Outcome
3.
Adv Healthc Mater ; 8(24): e1901182, 2019 12.
Article in English | MEDLINE | ID: mdl-31738017

ABSTRACT

CD44 is an endocytic hyaluronic acid (HA) receptor, and is overexpressed in many carcinomas. This has encouraged the use of HA to design CD44-targeting carriers. This paper is about dissecting the mechanistic role of CD44. Here, HA-decorated nanoparticles are used to deliver siRNA to both tumoral (AsPC-1, PANC-1, HT-29, HCT-116) and non-tumoral (fibroblasts, differently polarized THP-1 macrophages, HUVEC) human cell lines, evaluating the initial binding of the nanoparticles, their internalization rate, and the silencing efficiency (cyclophilin B (PPIB) gene). Tumoral cells internalize faster and experience higher silencing than non-tumoral cells. This is promising as it suggests that, in a tumor, HA nanocarriers may have limited off-target effects. More far-reaching is the inter-relation between the four parameters of the study: CD44 expression, HA binding on cell surfaces, internalization rate, and silencing efficiency. No correlation is found between binding (an early event) and any of the other parameters, whereas silencing correlates both with speed of the internalization process and CD44 expression. This study confirms on one hand that HA-based carriers can perform a targeted action, but on the other it suggests that this may not be due to a selective binding event, but rather to a later recognition leading to selective internalization.


Subject(s)
Hyaluronan Receptors/chemistry , Hyaluronic Acid/chemistry , Nanoparticles/chemistry , Cell Line , Cell Line, Tumor , Chitosan/chemistry , Drug Delivery Systems/methods , Dynamic Light Scattering , HCT116 Cells , HT29 Cells , Human Umbilical Vein Endothelial Cells , Humans , Kinetics , RNA, Small Interfering/chemistry , THP-1 Cells
4.
Eur J Med Chem ; 182: 111649, 2019 Nov 15.
Article in English | MEDLINE | ID: mdl-31514018

ABSTRACT

(NRH):quinone oxidoreductase 2 (NQO2) is associated with various processes involved in cancer initiation and progression probably via the production of ROS during quinone metabolism. Thus, there is a need to develop inhibitors of NQO2 that are active in vitro and in vivo. As part of a strategy to achieve this we have used the 4-aminoquinoline backbone as a starting point and synthesized 21 novel analogues. The syntheses utilised p-anisidine with Meldrum's acid and trimethyl orthoacetate or trimethyl orthobenzoate to give the 4-hydrazin-quinoline scaffold, which was derivatised with aldehydes or acid chlorides to give hydrazone or hydrazide analogues, respectively. The hydrazones were the most potent inhibitors of NQO2 in cell free systems, some with low nano-molar IC50 values. Structure-activity analysis highlighted the importance of a small substituent at the 2-position of the 4-aminoquinoline ring, to reduce steric hindrance and improve engagement of the scaffold within the NQO2 active site. Cytotoxicity and NQO2-inhibitory activity in vitro was evaluated using ovarian cancer SKOV-3 and TOV-112 cells (expressing high and low levels of NQO2, respectively). Generally, the hydrazones were more toxic than hydrazide analogues and further, toxicity is unrelated to cellular NQO2 activity. Pharmacological inhibition of NQO2 in cells was measured using the toxicity of CB1954 as a surrogate end-point. Both the hydrazone and hydrazide derivatives are functionally active as inhibitors of NQO2 in the cells, but at different inhibitory potency levels. In particular, 4-((2-(6-methoxy-2-methylquinolin-4-yl)hydrazono)methyl)phenol has the greatest potency of any compound yet evaluated (53 nM), which is 50-fold lower than its toxicity IC50. This compound and some of its analogues could serve as useful pharmacological probes to determine the functional role of NQO2 in cancer development and response to therapy.


Subject(s)
Aminoquinolines/pharmacology , Drug Discovery , Enzyme Inhibitors/pharmacology , Hydrazones/pharmacology , Quinone Reductases/antagonists & inhibitors , Aminoquinolines/chemical synthesis , Aminoquinolines/chemistry , Cell Survival/drug effects , Dose-Response Relationship, Drug , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Humans , Hydrazones/chemical synthesis , Hydrazones/chemistry , Models, Molecular , Molecular Structure , Quinone Reductases/metabolism , Structure-Activity Relationship , Tumor Cells, Cultured
5.
Chembiochem ; 20(22): 2841-2849, 2019 11 18.
Article in English | MEDLINE | ID: mdl-31165578

ABSTRACT

NAD(P)H quinone oxidoreductase-1 (NQO1) is a homodimeric protein that acts as a detoxifying enzyme or as a chaperone protein. Dicourmarol interacts with NQO1 at the NAD(P)H binding site and can both inhibit enzyme activity and modulate the interaction of NQO1 with other proteins. We show that the binding of dicoumarol and related compounds to NQO1 generates negative cooperativity between the monomers. This does not occur in the presence of the reducing cofactor, NAD(P)H, alone. Alteration of Gly150 (but not Gly149 or Gly174) abolished the dicoumarol-induced negative cooperativity. Analysis of the dynamics of NQO1 with the Gaussian network model indicates a high degree of collective motion by monomers and domains within NQO1. Ligand binding is predicted to alter NQO1 dynamics both proximal to the ligand binding site and remotely, close to the second binding site. Thus, drug-induced modulation of protein motion might contribute to the biological effects of putative inhibitors of NQO1.


Subject(s)
Allosteric Regulation/drug effects , Dicumarol/pharmacology , Enzyme Inhibitors/pharmacology , NAD(P)H Dehydrogenase (Quinone)/antagonists & inhibitors , Amino Acid Substitution , Catalytic Domain , Cell Line, Tumor , Dicumarol/metabolism , Enzyme Inhibitors/metabolism , Humans , Ligands , NAD(P)H Dehydrogenase (Quinone)/genetics , NAD(P)H Dehydrogenase (Quinone)/metabolism , Protein Binding , Tumor Suppressor Protein p53/metabolism
6.
Mol Pharm ; 16(6): 2481-2493, 2019 06 03.
Article in English | MEDLINE | ID: mdl-31013093

ABSTRACT

The development of delivery systems capable of tumor targeting represents a promising strategy to overcome issues related to nonspecific effects of conventional anticancer therapies. Currently, one of the most investigated agents for cancer targeting is hyaluronic acid (HA), since its receptor, CD44, is overexpressed in many cancers. However, most of the studies on CD44/HA interaction have been so far performed in cell-free or genetically modified systems, thus leaving some uncertainty regarding which cell-related factors influence HA binding and internalization (collectively called "uptake") into CD44-expressing cells. To address this, the expression of CD44 (both standard and variants, designated CD44s and CD44v, respectively) was evaluated in human dermal fibroblasts (HDFs) and a large panel of cancer cell lines, including breast, prostate, head and neck, pancreatic, ovarian, colorectal, thyroid, and endometrial cancers. Results showed that CD44 isoform profiles and expression levels vary across the cancer cell lines and HDF and are not consistent within the cell origin. Using composite information of CD44 expression, HA binding, and internalization, we found that the expression of CD44v can negatively influence the uptake of HA, and, instead, when cells primarily expressed CD44s, a positive correlation was observed between expression and uptake. In other words, CD44shigh cells bound and internalized more HA compared to CD44slow cells. Moreover, CD44shigh HDFs were less efficient in uptaking HA compared to CD44shigh cancer cells. The experiments described here are the first step toward understanding the interplay between CD44 expression, its functionality, and the underlying mechanism(s) for HA uptake. The results show that factors other than the amount of CD44 receptor can play a role in the interaction with HA, and this represents an important advance with respect to the design of HA-based carriers and the selection of tumors to treat according to their CD44 expression profile.


Subject(s)
Hyaluronan Receptors/metabolism , Hyaluronic Acid/chemistry , Hyaluronic Acid/therapeutic use , Cell Line, Tumor , Drug Delivery Systems/methods , Flow Cytometry , Humans , Immunohistochemistry
7.
Sci Rep ; 8(1): 16804, 2018 11 14.
Article in English | MEDLINE | ID: mdl-30429503

ABSTRACT

Prognosis of HPV negative head and neck squamous cell carcinoma (HNSCC) patients remains poor despite surgical and medical advances and inadequacy of predictive and prognostic biomarkers in this type of cancer highlights one of the challenges to successful therapy. Statins, widely used for the treatment of hyperlipidaemia, have been shown to possess anti-tumour effects which were partly attributed to their ability to interfere with metabolic pathways essential in the survival of cancer cells. Here, we have investigated the effect of statins on the metabolic modulation of HNSCC cancers with a vision to predict a personalised anticancer therapy. Although, treatment of tumour-bearing mice with simvastatin did not affect tumour growth, pre-treatment for 2 weeks prior to tumour injection, inhibited tumour growth resulting in strongly increased survival. This was associated with increased expression of the monocarboxylate transporter 1 (MCT1) and a significant reduction in tumour lactate content, suggesting a possible reliance of these tumours on oxidative phosphorylation for survival. Since MCT1 is responsible for the uptake of mitochondrial fuels into the cells, we reasoned that inhibiting it would be beneficial. Interestingly, combination of simvastatin with AZD3965 (MCT1 inhibitor) led to further tumour growth delay as compared to monotherapies, without signs of toxicity. In clinical biopsies, prediagnostic statin therapy was associated with a significantly higher MCT1 expression and was not of prognostic value following conventional chemo-radiotherapy. These findings provide a rationale to investigate the clinical effectiveness of MCT1 inhibition in patients with HNSCC who have been taking lipophilic statins prior to diagnosis.


Subject(s)
Head and Neck Neoplasms/diagnosis , Hydroxymethylglutaryl-CoA Reductase Inhibitors/pharmacology , Monocarboxylic Acid Transporters/antagonists & inhibitors , Animals , Biomarkers/metabolism , Head and Neck Neoplasms/drug therapy , Head and Neck Neoplasms/metabolism , Humans , Hydroxymethylglutaryl-CoA Reductase Inhibitors/therapeutic use , Lactic Acid/metabolism , Mice , Oxidative Phosphorylation , Precision Medicine , Prognosis , Pyrimidinones/pharmacology , Thiophenes/pharmacology
9.
Bioorg Med Chem Lett ; 28(8): 1292-1297, 2018 05 01.
Article in English | MEDLINE | ID: mdl-29567345

ABSTRACT

Inhibitors of the enzyme NQO2 (NRH: quinone oxidoreductase 2) are of potential use in cancer chemotherapy and malaria. We have previously reported that non-symmetrical furan amidines are potent inhibitors of NQO2 and here novel analogues are evaluated. The furan ring has been changed to other heterocycles (imidazole, N-methylimidazole, oxazole, thiophene) and the amidine group has been replaced with imidate, reversed amidine, N-arylamide and amidoxime to probe NQO2 activity, improve solubility and decrease basicity of the lead furan amidine. All compounds were fully characterised spectroscopically and the structure of the unexpected product N-hydroxy-4-(5-methyl-4-phenylfuran-2-yl)benzamidine was established by X-ray crystallography. The analogues were evaluated for inhibition of NQO2, which showed lower activity than the lead furan amidine. The observed structure-activity relationship for the furan-amidine series with NQO2 was rationalized by preliminary molecular docking and binding mode analysis. In addition, the oxazole-amidine analogue inhibited the growth of Plasmodium falciparum with an IC50 value of 0.3 µM.


Subject(s)
Amidines/pharmacology , Enzyme Inhibitors/pharmacology , Furans/pharmacology , Quinone Reductases/antagonists & inhibitors , Amidines/chemical synthesis , Amidines/chemistry , Antimalarials/chemical synthesis , Antimalarials/chemistry , Antimalarials/pharmacology , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Furans/chemical synthesis , Furans/chemistry , Imidazoles/chemical synthesis , Imidazoles/chemistry , Imidazoles/pharmacology , Molecular Structure , Oxazoles/chemical synthesis , Oxazoles/chemistry , Oxazoles/pharmacology , Oximes/chemical synthesis , Oximes/chemistry , Oximes/pharmacology , Plasmodium falciparum/drug effects , Structure-Activity Relationship , Thiophenes/chemical synthesis , Thiophenes/chemistry , Thiophenes/pharmacology
10.
EMBO Mol Med ; 9(12): 1646-1659, 2017 12.
Article in English | MEDLINE | ID: mdl-29084756

ABSTRACT

Radiotherapy is an important anti-cancer treatment, but tumour recurrence remains a significant clinical problem. In an effort to improve outcomes further, targeted anti-cancer drugs are being tested in combination with radiotherapy. Here, we have studied the effects of Akt inhibition with AZD5363. AZD5363 administered as an adjuvant after radiotherapy to FaDu and PE/CA PJ34 tumours leads to long-term tumour control, which appears to be secondary to effects on the irradiated tumour microenvironment. AZD5363 reduces the downstream effectors VEGF and HIF-1α, but has no effect on tumour vascularity or oxygenation, or on tumour control, when administered prior to radiotherapy. In contrast, AZD5363 given after radiotherapy is associated with marked reductions in tumour vascular density, a decrease in the influx of CD11b+ myeloid cells and a failure of tumour regrowth. In addition, AZD5363 is shown to inhibit the proportion of proliferating tumour vascular endothelial cells in vivo, which may contribute to improved tumour control with adjuvant treatment. These new insights provide promise to improve outcomes with the addition of AZD5363 as an adjuvant therapy following radiotherapy.


Subject(s)
Head and Neck Neoplasms/drug therapy , Protein Kinase Inhibitors/therapeutic use , Proto-Oncogene Proteins c-akt/antagonists & inhibitors , Pyrimidines/therapeutic use , Pyrroles/therapeutic use , Tumor Microenvironment , Animals , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Survival/drug effects , Combined Modality Therapy , Endothelial Cells/drug effects , Endothelial Cells/metabolism , Endothelial Cells/pathology , Female , Gamma Rays/therapeutic use , Head and Neck Neoplasms/pathology , Head and Neck Neoplasms/radiotherapy , Humans , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Mice , Mice, Nude , Positron Emission Tomography Computed Tomography , Protein Kinase Inhibitors/pharmacology , Proto-Oncogene Proteins c-akt/metabolism , Pyrimidines/pharmacology , Pyrroles/pharmacology , Transplantation, Heterologous , Tumor Microenvironment/drug effects , Vascular Endothelial Growth Factor A/metabolism
12.
Clin Endocrinol (Oxf) ; 87(5): 557-565, 2017 Nov.
Article in English | MEDLINE | ID: mdl-28748640

ABSTRACT

OBJECTIVE: Women with a prior history of polycystic ovary syndrome (PCOS) have an increased risk of endometrial cancer (EC). AIM: To investigate whether the endometrium of women with PCOS possesses gene expression changes similar to those found in EC. DESIGN AND METHODS: Patients with EC, PCOS and control women unaffected by either PCOS or EC were recruited into a cross-sectional study at the Nottingham University Hospital, UK. For RNA sequencing, representative individual endometrial biopsies were obtained from women with EC, PCOS and a woman unaffected by PCOS or EC. Expression of a subset of differentially expressed genes identified by RNA sequencing, including NAD(P)H quinone dehydrogenase 1 (NQO1), was validated by quantitative reverse transcriptase PCR validation (n = 76) and in the cancer genome atlas UCEC (uterine corpus endometrioid carcinoma) RNA sequencing data set (n = 381). The expression of NQO1 was validated by immunohistochemistry in EC samples from a separate cohort (n = 91) comprised of consecutive patients who underwent hysterectomy at St Mary's Hospital, Manchester, between 2011 and 2013. A further 6 postmenopausal women with histologically normal endometrium who underwent hysterectomy for genital prolapse were also included. Informed consent and local ethics approval were obtained for the study. RESULTS: We show for the first that NQO1 expression is significantly increased in the endometrium of women with PCOS and EC. Immunohistochemistry confirms significantly increased NQO1 protein expression in EC relative to nonmalignant endometrial tissue (P < .0001). CONCLUSIONS: The results obtained here support a previously unrecognized molecular link between PCOS and EC involving NQO1.


Subject(s)
Endometrial Neoplasms/metabolism , Endometrium/metabolism , NAD(P)H Dehydrogenase (Quinone)/metabolism , Polycystic Ovary Syndrome/metabolism , Adult , Aged , Aged, 80 and over , Case-Control Studies , Cross-Sectional Studies , Endometrial Neoplasms/enzymology , Endometrium/enzymology , Female , Gene Expression , Humans , Immunohistochemistry , Middle Aged , Polycystic Ovary Syndrome/enzymology , Young Adult
13.
Cancer Lett ; 403: 98-107, 2017 09 10.
Article in English | MEDLINE | ID: mdl-28624622

ABSTRACT

Organometallic complexes offer the prospect of targeting multiple pathways that are important in cancer biology. Here, the preclinical activity and mechanism(s) of action of a silver-bis(N-heterocyclic carbine) complex (Ag8) were evaluated. Ag8 induced DNA damage via several mechanisms including topoisomerase I/II and thioredoxin reductase inhibition and induction of reactive oxygen species. DNA damage induction was consistent with cytotoxicity observed against proliferating cells and Ag8 induced cell death by apoptosis. Ag8 also inhibited DNA repair enzyme PARP1, showed preferential activity against cisplatin resistant A2780 cells and potentiated the activity of temozolomide. Ag8 was substantially less active against non-proliferating non-cancer cells and selectively inhibited glycolysis in cancer cells. Ag8 also induced significant anti-tumour effects against cells implanted intraperitoneally in hollow fibres but lacked activity against hollow fibres implanted subcutaneously. Thus, Ag8 targets multiple pathways of importance in cancer biology, is less active against non-cancer cells and shows activity in vivo in a loco-regional setting.


Subject(s)
Antineoplastic Agents/pharmacology , Imidazoles/pharmacology , Neoplasms/drug therapy , Organometallic Compounds/pharmacology , Antigens, Neoplasm/metabolism , Antineoplastic Agents/chemistry , Antineoplastic Agents/toxicity , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Apoptosis/drug effects , Cell Line, Tumor , Cell Survival/drug effects , Cisplatin/pharmacology , DNA Damage , DNA Topoisomerases, Type I/metabolism , DNA Topoisomerases, Type II/metabolism , DNA-Binding Proteins/metabolism , Dacarbazine/analogs & derivatives , Dacarbazine/pharmacology , Dose-Response Relationship, Drug , Drug Resistance, Neoplasm , Drug Synergism , Glycolysis/drug effects , Humans , Imidazoles/chemistry , Imidazoles/toxicity , Inhibitory Concentration 50 , Neoplasms/metabolism , Neoplasms/pathology , Organometallic Compounds/chemistry , Organometallic Compounds/toxicity , Oxidative Stress/drug effects , Poly (ADP-Ribose) Polymerase-1/antagonists & inhibitors , Poly (ADP-Ribose) Polymerase-1/metabolism , Poly(ADP-ribose) Polymerase Inhibitors/pharmacology , Reactive Oxygen Species/metabolism , Signal Transduction/drug effects , Temozolomide , Thioredoxin Reductase 1/antagonists & inhibitors , Thioredoxin Reductase 1/metabolism , Topoisomerase I Inhibitors/pharmacology , Topoisomerase II Inhibitors/pharmacology
14.
Mol Pharm ; 14(7): 2422-2436, 2017 07 03.
Article in English | MEDLINE | ID: mdl-28597662

ABSTRACT

Chitosan/hyaluronic acid (HA) nanoparticles can be used to deliver an RNA/DNA cargo to cells overexpressing HA receptors such as CD44. For these systems, unequivocal links have not been established yet between chitosan macromolecular (molecular weight; degree of deacetylation, i.e., charge density) and nanoparticle variables (complexation strength, i.e., stability; nucleic acid protection; internalization rate) on one hand, and transfection efficiency on the other hand. Here, we have focused on the role of avidity on transfection efficiency in the CD44-expressing HCT-116 as a cellular model; we have employed two differently sized payloads (a large luciferase-encoding mRNA and a much smaller anti-Luc siRNA), and a small library of chitosans (variable molecular weight and degree of deactylation). The RNA avidity for chitosan showed-as expected-an inverse relationship: higher avidity-higher polyplex stability-lower transfection efficiency. The avidity of chitosan for RNA appears to lead to opposite effects: higher avidity-higher polyplex stability but also higher transfection efficiency. Surprisingly, the best transfecting particles were those with the lowest propensity for RNA release, although this might be a misleading relationship: for example, the same macromolecular parameters that increase avidity can also boost chitosan's endosomolytic activity, with a strong enhancement in transfection. The performance of these nonviral vectors appears therefore difficult to predict simply on the basis of carrier- or payload-related variables, and a more holistic consideration of the journey of the nanoparticle, from cell uptake to cytosolic bioavailability of payload, is needed. It is also noteworthy that the nanoparticles used in this study showed optimal performance under slightly acidic conditions (pH 6.4), which is promising for applications in a tumoral extracellular environment. It is also worth pointing out that under these conditions we have for the first time successfully delivered mRNA with chitosan/HA nanoparticles.


Subject(s)
Chitosan/chemistry , Hyaluronic Acid/chemistry , Nanoparticles/chemistry , Dynamic Light Scattering , HCT116 Cells , Humans , Hyaluronan Receptors/genetics , Hyaluronan Receptors/metabolism , Molecular Weight , Peptides, Cyclic , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism
15.
BMC Clin Pathol ; 17: 27, 2017.
Article in English | MEDLINE | ID: mdl-29299023

ABSTRACT

BACKGROUND: Endometrial cancer (EC) is a major health concern due to its rising incidence. Whilst early stage disease is generally cured by surgery, advanced EC has a poor prognosis with limited treatment options. Altered energy metabolism is a hallmark of malignancy. Cancer cells drive tumour growth through aerobic glycolysis and must export lactate to maintain intracellular pH. The aim of this study was to evaluate the expression of the lactate/proton monocarboxylate transporters MCT1 and MCT4 and their chaperone CD147 in EC, with the ultimate aim of directing future drug development. METHODS: MCT1, MCT4 and CD147 expression was examined using immunohistochemical analysis in 90 endometrial tumours and correlated with clinico-pathological characteristics and survival outcomes. RESULTS: MCT1 and MCT4 expression was observed in the cytoplasm, the plasma membrane or both locations. CD147 was detected in the plasma membrane and associated with MCT1 (p = 0.003) but not with MCT4 (p = 0.207) expression. High MCT1 expression was associated with reduced overall survival (p = 0.029) and remained statistically significant after adjustment for survival covariates (p = 0.017). CONCLUSION: Our data suggest that MCT1 expression is an important marker of poor prognosis in EC. MCT1 inhibition may have potential as a treatment for advanced or recurrent EC.

16.
Adv Healthc Mater ; 6(4)2017 Feb.
Article in English | MEDLINE | ID: mdl-27990775

ABSTRACT

CD44 is a potentially rewarding target in cancer therapy, although its mechanisms of ligand binding and internalization are still poorly understood. In this study, we have established quantitative relationships between CD44 expression in differently polarized macrophages (M0, M1, and M2-polarized THP-1 human macrophages) and the uptake of hyaluronic acid (HA)-based materials, which are potentially usable for CD44 targeting. We have validated a robust method for macrophage polarization, which sequentially uses differentiating and polarizing factors, and allows to show that CD44 expression depends on polarization (M1 > M0 ≥ M2). It is noteworthy that THP-1 M2 expressed CD44v6, suggesting their suitability as a model of tumor-associated macrophages. In the uptake of HA, both as a soluble polymer and in the form of (siRNA-loaded) nanoparticles, CD44 expression correlated positively with binding, but negatively with internalization. Counterintuitively, it appears that a higher presence of CD44 (in M1) allows a more efficient capture of HA materials, but a lower expression (in M2) is conducive to better internalization. Although possibly cell-specific, this unexpected relationship indicates that the common paradigm "higher CD44 expression = better targetability" is too simplistic; mechanistic details of both receptor presentation and association still need to be elucidated for a predictable targeting behavior.


Subject(s)
Drug Delivery Systems/methods , Hyaluronan Receptors/metabolism , Hyaluronic Acid , Macrophages/metabolism , Nanoparticles/chemistry , Cell Line, Tumor , Humans , Hyaluronic Acid/chemistry , Hyaluronic Acid/pharmacokinetics , Hyaluronic Acid/pharmacology , Macrophages/pathology , Neoplasms/drug therapy , Neoplasms/metabolism
17.
Nat Rev Clin Oncol ; 13(10): 627-42, 2016 10.
Article in English | MEDLINE | ID: mdl-27245279

ABSTRACT

In countries with the best cancer outcomes, approximately 60% of patients receive radiotherapy as part of their treatment, which is one of the most cost-effective cancer treatments. Notably, around 40% of cancer cures include the use of radiotherapy, either as a single modality or combined with other treatments. Radiotherapy can provide enormous benefit to patients with cancer. In the past decade, significant technical advances, such as image-guided radiotherapy, intensity-modulated radiotherapy, stereotactic radiotherapy, and proton therapy enable higher doses of radiotherapy to be delivered to the tumour with significantly lower doses to normal surrounding tissues. However, apart from the combination of traditional cytotoxic chemotherapy with radiotherapy, little progress has been made in identifying and defining optimal targeted therapy and radiotherapy combinations to improve the efficacy of cancer treatment. The National Cancer Research Institute Clinical and Translational Radiotherapy Research Working Group (CTRad) formed a Joint Working Group with representatives from academia, industry, patient groups and regulatory bodies to address this lack of progress and to publish recommendations for future clinical research. Herein, we highlight the Working Group's consensus recommendations to increase the number of novel drugs being successfully registered in combination with radiotherapy to improve clinical outcomes for patients with cancer.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Neoplasms/radiotherapy , Cell Hypoxia/radiation effects , Clinical Trials as Topic/methods , Combined Modality Therapy , Drug Approval , Humans , Neoplasms/drug therapy , Patient Education as Topic , Patient Participation , Quality Assurance, Health Care , Quality of Health Care , Radiation Dosage , Radiation Tolerance/radiation effects , Treatment Outcome
18.
Oncotarget ; 7(13): 17035-46, 2016 Mar 29.
Article in English | MEDLINE | ID: mdl-26959743

ABSTRACT

Strategies to augment anti-cancer immune responses have recently demonstrated therapeutic utility. To date clinical success has been achieved through targeting co-inhibitory checkpoints such as CTLA-4, PD-1, and PD-L1. However, approaches that target co-activatory pathways are also being actively being developed. Here we report that the novel TLR7-selective agonist DSR-29133 is well tolerated in mice and leads to acute immune activation. Administration of DSR-29133 leads to the induction of IFNα/γ, IP-10, TNFα, IL-1Ra and IL-12p70, and to a reduction in tumor burden in syngeneic models of renal cancer (Renca), metastatic osteosarcoma (LM8) and colorectal cancer (CT26). Moreover, we show that the efficacy of DSR-29133 was significantly improved when administered in combination with low-dose fractionated radiotherapy (RT). Effective combination therapy required weekly administration of DSR-29133 commencing on day 1 of a fractionated RT treatment cycle, whereas no enhancement of radiation response was observed when DSR-29133 was administered at the end of the fractionated RT cycle. Combined therapy resulted in curative responses in a high proportion of mice bearing established CT26 tumors which was dependent on the activity of CD8+ T-cells but independent of CD4+ T-cells and NK/NKT cells. Moreover, long-term surviving mice originally treated with DSR-29133 and RT were protected by a tumor-specific memory immune response which could prevent tumor growth upon rechallenge. These results demonstrate that DSR-29133 is a potent selective TLR7 agonist that when administered intravenously can induce anti-tumor immune responses that can be further enhanced through combination with low-dose fractionated RT.


Subject(s)
Adenine/analogs & derivatives , Antineoplastic Agents/pharmacology , Chemoradiotherapy/methods , Neoplasms, Experimental/drug therapy , Toll-Like Receptor 7/agonists , Adenine/pharmacology , Administration, Intravenous , Animals , CD8-Positive T-Lymphocytes/drug effects , CD8-Positive T-Lymphocytes/immunology , Dose Fractionation, Radiation , Humans , Lymphocyte Activation/drug effects , Mice , Neoplasms, Experimental/immunology , Neoplasms, Experimental/radiotherapy
19.
Eur J Med Chem ; 111: 33-45, 2016 Mar 23.
Article in English | MEDLINE | ID: mdl-26854376

ABSTRACT

NRH:quinone oxidoreductase 2 enzyme (NQO2) is a potential therapeutic target in cancer and neurodegenerative diseases, with roles in either chemoprevention or chemotherapy. Here we report the design, synthesis and evaluation of non-symmetrical furan-amidines and their analogues as novel selective NQO2 inhibitors with reduced adverse off-target effects, such as binding to DNA. A pathway for the synthesis of the non-symmetrical furan-amidines was established from the corresponding 1,4-diketones. The synthesized non-symmetrical furan-amidines and their analogues showed potent NQO2 inhibition activity with nano-molar IC50 values. The most active compounds were non-symmetrical furan-amidines with meta- and para-nitro substitution on the aromatic ring, with IC50 values of 15 nM. In contrast to the symmetric furan-amidines, which showed potent intercalation in the minor grooves of DNA, the synthesized non-symmetrical furan-amidines showed no affinity towards DNA, as demonstrated by DNA melting temperature experiments. In addition, Plasmodium parasites, which possess their own quinone oxidoreductase PfNDH2, were inhibited by the non-symmetrical furan-amidines, the most active possessing a para-fluoro substituent (IC50 9.6 nM). The high NQO2 inhibition activity and nanomolar antimalarial effect of some of these analogues suggest the lead compounds are worthy of further development and optimization as potential drugs for novel anti-cancer and antimalarial strategies.


Subject(s)
Amidines/pharmacology , Antimalarials/pharmacology , Antineoplastic Agents/pharmacology , Furans/pharmacology , Malaria/drug therapy , Plasmodium/drug effects , Amidines/chemical synthesis , Amidines/chemistry , Antimalarials/chemical synthesis , Antimalarials/chemistry , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Cell Proliferation/drug effects , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Furans/chemical synthesis , Furans/chemistry , Humans , Models, Molecular , Molecular Structure , Parasitic Sensitivity Tests , Structure-Activity Relationship
20.
Oncol Rep ; 35(4): 1925-32, 2016 Apr.
Article in English | MEDLINE | ID: mdl-26782976

ABSTRACT

Banoxantrone (AQ4N) is a prototype hypoxia selective cytotoxin that is activated by haem containing reductases such as inducible nitric oxide synthase (iNOS). In the present study, we evaluate whether elevated levels of iNOS in human tumour cells will improve their sensitivity to AQ4N. Further, we examine the potential of radiation to increase cellular toxicity of AQ4N under normoxic (aerobic) and hypoxic conditions. We employed an expression vector containing the cDNA for human iNOS to transfect human fibrosarcoma HT1080 tumour cells. Alternatively, parental cells were exposed to a cytokine cocktail to induce iNOS gene expression and enzymatic activity. The cells were then treated with AQ4N alone and in combination with radiation in the presence or absence of the iNOS inhibitor N-methyl-L­arginine. In parental cells, AQ4N showed little difference in toxicity under hypoxic verses normoxic conditions. Notably, cells with upregulated iNOS activity showed a significant increase in sensitivity to AQ4N, but only under conditions of reduced oxygenation. When these cells were exposed to the combination of AQ4N and radiation, there was much greater cell killing than that observed with either modality alone. In the clinical development of hypoxia selective cytotoxins it is likely they will be used in combination with radiotherapy. In the present study, we demonstrated that AQ4N can selectively kill hypoxic cells via an iNOS-dependent mechanism. This hypoxia-selective effect can be augmented by combining AQ4N with radiation without increasing cytotoxicity to well­oxygenated tissues. Collectively, these results suggest that targeting hypoxic tumours with high levels of iNOS with a combination of AQ4N and radiotherapy could be a useful clinical therapeutic strategy.


Subject(s)
Anthraquinones/pharmacology , Antineoplastic Agents/pharmacology , Fibrosarcoma/genetics , Nitric Oxide Synthase Type II/genetics , Cell Hypoxia/drug effects , Cell Hypoxia/radiation effects , Cell Line, Tumor , Combined Modality Therapy , Fibrosarcoma/drug therapy , Fibrosarcoma/radiotherapy , Gene Expression Regulation, Neoplastic/drug effects , Gene Expression Regulation, Neoplastic/radiation effects , Humans , Nitric Oxide Synthase Type II/metabolism , Up-Regulation/drug effects , Up-Regulation/radiation effects
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