APOBEC3B (A3B)-catalyzed DNA cytosine deamination contributes to the overall mutational landscape in breast cancer. Molecular mechanisms responsible for
upregulation in cancer are poorly understood. ...Here we show that a single E2F cis-element mediates repression in normal cells and that expression is activated by its mutational disruption in a reporter construct or the endogenous
gene. The same E2F site is required for
induction by polyomavirus T antigen indicating a shared molecular mechanism. Proteomic and biochemical experiments demonstrate the binding of wildtype but not mutant E2F promoters by repressive PRC1.6/E2F6 and DREAM/E2F4 complexes. Knockdown and overexpression studies confirm the involvement of these repressive complexes in regulating A3B expression. Altogether, these studies demonstrate that
expression is suppressed in normal cells by repressive E2F complexes and that viral or mutational disruption of this regulatory network triggers overexpression in breast cancer and provides fuel for tumor evolution.
Highlights • Hypoxia is an inherent negative factor for outcome after chemo- and radio-therapy. • PI3-K/AKT signaling affects hypoxia via HIF-1-dependent & -independent mechanisms. • PI3-K/AKT ...blockers act synergistic with chemo- or radio-therapy in animal models. • Reduction of hypoxia by PI3-K/AKT blockers seems important for tumor sensitization. • Phase I trials show the feasibility of combining AKT inhibition with chemoradiation.
Three-dimensional spheroids of non-malignant MCF10A and malignant SKBR3 breast cells were used for subsequent 3D Cell-SELEX to generate aptamers for specific binding and treatment of breast cancer ...cells. Using 3D Cell-SELEX combined with Next-Generation Sequencing and bioinformatics, ten abundant aptamer families with specific structures were identified that selectively bind to SKBR3, and not to MCF10A cells. Multivalent aptamer polymers were synthesized by co-polymerization and analyzed for binding performance as well as therapeutic efficacy. Binding performance was determined by confocal fluorescence imaging and revealed specific binding and efficient internalization of aptamer polymers into SKBR3 spheroids. For therapeutic purposes, DNA sequences that intercalate the cytotoxic drug doxorubicin were co-polymerized into the aptamer polymers. Viability tests show that the drug-loaded polymers are specific and effective in killing SKBR3 breast cancer cells. Thus, the 3D-selected aptamers enhanced the specificity of doxorubicin against malignant over non-malignant breast cells. The innovative modular DNA aptamer platform based on 3D Cell SELEX and polymer multivalency holds great promise for diagnostics and treatment of breast cancer.
Objective
The plasminogen activator system (PAS) and vascular endothelial growth factor (VEGF) are important in the carcinogenesis and play a key role in cancer invasion and mediating metastasis of ...carcinomas. The aim of the study was to evaluate the correlation of serum levels of VEGF and components of the PAS with clinicopathological risk factors and outcome in patients with endometrial cancer (EC).
Methods
Preoperative blood was collected from 173 patients treated for EC between 1999 and 2009. Serum concentrations of VEGF, urokinase plasminogen activator (uPA) tissue plasminogen activator (tPA), plasminogen activator inhibitor type-1 (PAI-1) and -2 (PAI-2) were assessed by enzyme-linked immunosorbent assays (ELISA).
Results
Serum levels of VEGF and components of the PAS were significantly associated with stage of the disease, tumor histology, tumor grade, myometrial invasion (MI), presence of lymphovascular space invasion (LVSI) and lymph node metastases (LNM). Preoperative serum levels of PAI-1 and -2 and tPA were higher in patients who experienced a recurrence than in patients who remained disease free (
p
< 0.01). PAI-1 and -2 and tPA were significantly independent prognostic factors for DFS with a HR of 3.85 (95% CI 1.84–8.07), 3.90 (95% CI 1.75–8.66) and 2.53 (95% CI 1.16–5.55), respectively. PAI-1 and tPA turned out to be independent prognostic factors for OS, with a HR of 2.09 (95% CI 1.08–4.05) and 2.16 (95% CI 1.06–4.44), respectively.
Conclusion
Serum levels of VEGF and components of the PAS at primary diagnosis were associated with well-known clinicopathological risk factors such as; FIGO stage, tumor histology, tumor grade, MI, LVSI and LNM. High concentrations of PAI-1 and-2 and tPA are independent factors for poor prognosis in patients with endometrial cancer.
•The AKT pathway is activated by hypoxia in HPV-positive and HPV-negative head and neck squamous cell carcinoma (HNSCC) cells.•HPV-positive HNSCC cells are more sensitive to AKT inhibition under ...hypoxia.•AKT inhibition radiosensitises hypoxic HPV-positive HNSCC.
Hypoxia negatively affects treatment outcome in both Human papillomavirus (HPV)-positive and -negative head and neck squamous cell carcinomas (HNSCC). Despite HPV-positive patients having a relatively good prognosis, hypoxic HPV-positive tumours are associated with poor treatment outcome, and do not respond to hypoxia modification. Earlier, we showed that hypoxia induces the pro-survival AKT pathway. In this study, we aim to investigate whether AKT inhibition affects the response to radiotherapy under hypoxia, and determine whether this is a viable treatment strategy for HNSCC patients with hypoxic HPV-positive tumours.
Nine HPV-negative and 4 HPV-positive HNSCC cell lines were characterized. AKT activation was assessed by western blot. Survival in response to hypoxic incubation, AKT inhibition and/or irradiation was assessed using CCK8 assays and colony forming assays.
AKT was activated under hypoxia in both HPV-negative and -positive cell lines, which could be abrogated by the AKT inhibitor MK2206. HPV-positive cell lines were highly sensitive to MK2206 at normoxia. In all HNSCC cell lines, AKT inhibition was significantly more effective in inhibiting cell growth during hypoxic conditions than under normoxia. Hypoxia significantly reduced radiosensitivity irrespective of HPV-status, yet specifically in HPV-positive cells this could be efficiently reversed by AKT inhibition.
These data suggest that HNSCC tumours are dependent on AKT to survive hypoxia, and that AKT inhibition is specifically effective in radioresistant hypoxic HPV-positive cells. Targeting AKT may thus be a potential way to overcome hypoxia induced radioresistance, particularly in HPV-positive HNSCC tumours.
Carbonic anhydrase IX (CAIX) plays a pivotal role in pH homeostasis, which is essential for tumor cell survival. We examined the effect of the CAIX inhibitor 4-(3'(3",5"-dimethylphenyl)-ureido)phenyl ...sulfamate (S4) on the tumor microenvironment in a laryngeal tumor model by analyzing proliferation, apoptosis, necrosis, hypoxia, metabolism and CAIX ectodomain shedding.
SCCNij202 tumor bearing-mice were treated with S4 for 1, 3 or 5 days. CAIX ectodomain shedding was measured in the serum after therapy. Effects on tumor cell proliferation, apoptosis, necrosis, hypoxia (pimonidazole) and CAIX were investigated with quantitative immunohistochemistry. Metabolic transporters and enzymes were quantified with qPCR.
CAIX ectodomain shedding decreased after treatment with S4 (p<0.01). S4 therapy did neither influence tumor cell proliferation nor the amount of apoptosis and necrosis. Hypoxia (pimonidazole) and CAIX expression were also not affected by S4. CHOP and MMP9 mRNA as a reference of intracellular pH did not change upon treatment with S4. Compensatory mechanisms of pH homeostasis at the mRNA level were not observed.
As the clinical and biological meaning of the decrease in CAIX ectodomain shedding after S4 therapy is not clear, studies are required to elucidate whether the CAIX ectodomain has a paracrine or autocrine signaling function in cancer biology. S4 did not influence the amount of proliferation, apoptosis, necrosis and hypoxia. Therefore, it is unlikely that S4 can be used as single agent to influence tumor cell kill and proliferation, and to target primary tumor growth.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Abstract Autophagy is a process in which cells can generate energy and building materials, by degradation of redundant and/or damaged organelles and proteins. Especially during conditions of stress, ...autophagy helps to maintain homeostasis. In addition, autophagy has been shown to influence malignant transformation and cancer progression. The precise molecular events in autophagy are complex and the core autophagic machinery described to date consists of nearly thirty proteins. Apart from these factors that execute the process of autophagy, several signalling pathways are involved in converting internal and external stimuli into an autophagic response. In this review we provide an overview of the signalling pathways that influence autophagy, particularly in cancer cells. We will illustrate that interference with multiple of these signalling pathways can have significant effects on cancer cell survival.
Morphogenesis is a tightly‐regulated developmental process by which tissues acquire the morphology that is critical to their function. For example, epithelial cells exhibit different 2D and 3D ...morphologies, induced by distinct biochemical and biophysical cues from their environment. In this work, novel hybrid matrices composed of a Matrigel and synthetic oligo(ethylene glycol)‐grafted polyisocyanides (PICs) hydrogels are used to form a highly tailorable environment. Through precise control of the stiffness and cell‐matrix interactions, while keeping other properties constant, a broad range of morphologies induced in Madin‐Darby Canine Kidney (MDCK) cells is observed. At relatively low matrix stiffness, a large morphological shift from round hollow cysts to 2D monolayers is observed, without concomitant translocation of the mechanotransduction protein Yes‐associated protein (YAP). At higher stiffness levels and enhanced cell‐matrix interactions, tuned by controlling the adhesive peptide density on PIC, the hybrid hydrogels induce a flattened cell morphology with simultaneous YAP translocation, suggesting activation. In 3D cultures, the latter matrices lead to the formation of tubular structures. Thus, mixed synthetic and natural gels, such as the hybrids presented here, are ideal platforms to dissect how external physical factors can be used to regulate morphogenesis in MDCK model system, and in the future, in more complex environments.
The cellular environment plays a key role in determining tissue morphogenesis. This work uses hybrid matrices, composed of synthetic polyisocyanides (PICs) and Matrigel, that allows for precise control of matrix stiffness and cell‐matrix interactions. Variations in hybrid matrices induce a full spectrum of morphologies in Madin‐Darby canine kidney (MDCK) cells, and includes both YAP‐dependent and ‐independent processes.
Limited diffusion of oxygen in combination with increased oxygen consumption leads to chronic hypoxia in most solid malignancies. This scarcity of oxygen is known to induce radioresistance and leads ...to an immunosuppressive microenvironment. Carbonic anhydrase IX (CAIX) is an enzyme functioning as a catalyzer for acid export in hypoxic cells and is an endogenous biomarker for chronic hypoxia. The aim of this study is to develop a radiolabeled antibody that recognizes murine CAIX to visualize chronic hypoxia in syngeneic tumor models and to study the immune cell population in these hypoxic areas. An anti-mCAIX antibody (MSC3) was conjugated to diethylenetriaminepentaacetic acid (DTPA) and radiolabeled with indium-111 (111In). CAIX expression on murine tumor cells was determined using flow cytometry, and in vitro affinity of 111InIn-MSC3 was analyzed in a competitive binding assay. Ex vivo biodistribution studies were performed to determine in vivo radiotracer distribution. CAIX+ tumor fractions were determined by mCAIX microSPECT/CT, and the tumor microenvironment was analyzed using immunohistochemistry and autoradiography. We showed that 111InIn-MSC3 binds to CAIX-expressing (CAIX+) murine cells in vitro and accumulates in CAIX+ areas in vivo. We optimized the use of 111InIn-MSC3 for preclinical imaging such that it can be applied in syngeneic mouse models and showed that we can quantitatively distinguish between tumor models with varying CAIX+ fractions by ex vivo analyses and in vivo mCAIX microSPECT/CT. Analysis of the tumor microenvironment identified these CAIX+ areas as less infiltrated by immune cells. Together these data demonstrate that mCAIX microSPECT/CT is a sensitive technique to visualize hypoxic CAIX+ tumor areas that exhibit reduced infiltration of immune cells in syngeneic mouse models. In the future, this technique may enable visualization of CAIX expression before or during hypoxia-targeted or hypoxia-reducing treatments. Thereby, it will help optimize immuno- and radiotherapy efficacy in translationally relevant syngeneic mouse tumor models.
PDF
