Synergistic Effects of Sanglifehrin-Based Cyclophilin Inhibitor NV651 with Cisplatin in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC), commonly diagnosed at an advanced stage, is the most common primary liver cancer. Owing to a lack of effective HCC treatments and the commonly acquired chemoresistance, novel therapies need to be investigated. Cyclophilins-intracellular proteins with peptidyl-prolyl isomerase activity-have been shown to play a key role in therapy resistance and cell proliferation.

Novel Cyclophilin Inhibitor Decreases Cell Proliferation and Tumor Growth in Models of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC), the most common primary liver cancer, is usually diagnosed in its late state. Tyrosine kinase inhibitors such as sorafenib and regorafenib are one of the few treatment options approved for advanced HCC and only prolong the patient's life expectancy by a few months. Therefore, there is a need for novel effective treatments.

Mutant p53 controls tumor metabolism and metastasis by regulating PGC-1α.

Mutant forms of p53 protein often possess protumorigenic functions, conferring increased survival and migration to tumor cells via their "gain-of-function" activity. Whether and how a common polymorphism in at amino acid 72 (Pro72Arg; referred to here as P72 and R72) impacts this gain of function has not been determined. We show that mutant p53 enhances migration and metastasis of tumors through the ability to bind and regulate PGC-1α and that this regulation is markedly impacted by the codon 72 polymorphism.

Transgenic Expression of the Mitochondrial Chaperone TNFR-associated Protein 1 (TRAP1) Accelerates Prostate Cancer Development.

Protein homeostasis, or proteostasis, is required for mitochondrial function, but its role in cancer is controversial. Here we show that transgenic mice expressing the mitochondrial chaperone TNFR-associated protein 1 (TRAP1) in the prostate develop epithelial hyperplasia and cellular atypia. When examined on a Pten background, a common alteration in human prostate cancer, TRAP1 transgenic mice showed accelerated incidence of invasive prostatic adenocarcinoma, characterized by increased cell proliferation and reduced apoptosis, in situ Conversely, homozygous deletion of TRAP1 delays prostatic tumorigenesis in Pten mice without affecting hyperplasia or prostatic intraepithelial neoplasia.

Deletion of Cyclophilin D Impairs β-Oxidation and Promotes Glucose Metabolism.

Cyclophilin D (CypD) is a mitochondrial matrix protein implicated in cell death, but a potential role in bioenergetics is not understood. Here, we show that loss or depletion of CypD in cell lines and mice induces defects in mitochondrial bioenergetics due to impaired fatty acid β-oxidation. In turn, CypD loss triggers a global compensatory shift towards glycolysis, with transcriptional upregulation of effectors of glucose metabolism, increased glucose consumption and higher ATP production.

Adaptive mitochondrial reprogramming and resistance to PI3K therapy.

Background: Small molecule inhibitors of phosphatidylinositol-3 kinase (PI3K) have been developed as molecular therapy for cancer, but their efficacy in the clinic is modest, hampered by resistance mechanisms.

Methods: We studied the effect of PI3K therapy in patient-derived tumor organotypic cultures (from five patient samples), three glioblastoma (GBM) tumor cell lines, and an intracranial model of glioblastoma in immunocompromised mice (n = 4-5 mice per group). Mechanisms of therapy-induced tumor reprogramming were investigated in a global metabolomics screening, analysis of mitochondrial bioenergetics and cell death, and modulation of protein phosphorylation.

Deletion of the mitochondrial chaperone TRAP-1 uncovers global reprogramming of metabolic networks.

Reprogramming of metabolic pathways contributes to human disease, especially cancer, but the regulators of this process are unknown. Here, we have generated a mouse knockout for the mitochondrial chaperone TRAP-1, a regulator of bioenergetics in tumors. TRAP-1(-/-) mice are viable and showed reduced incidence of age-associated pathologies, including obesity, inflammatory tissue degeneration, dysplasia, and spontaneous tumor formation.

DNA-PK inhibition by NU7441 sensitizes breast cancer cells to ionizing radiation and doxorubicin.

DNA-dependent protein kinase (DNA-PK) plays a key role in the repair of DNA double-strand breaks (DSBs) that are probably the most deleterious form of DNA damage. Inhibition of DNA-PK has been considered as an attractive approach to decrease resistance to therapeutically induced DNA DSBs. Ionizing radiation (IR) and doxorubicin, which induce DSBs, are used in the treatment of breast cancer.

Cyclophilin D extramitochondrial signaling controls cell cycle progression and chemokine-directed cell motility.

Mitochondria control bioenergetics and cell fate decisions, but how they influence nuclear gene expression is understood poorly. Here, we show that deletion or reduction in the levels of cyclophilin D (CypD, also called Ppif), a mitochondrial matrix peptidyl prolyl isomerase and apoptosis regulator, results in increased cell proliferation and enhanced cell migration and invasion. These responses are associated with extensive transcriptional changes, modulation of a chemokine/chemokine receptor gene signature, and activation of the pleiotropic inflammatory mediator, STAT3.

Chk2 phosphorylation of survivin-DeltaEx3 contributes to a DNA damage-sensing checkpoint in cancer.

Survivin is an oncogene that functions in cancer cell cytoprotection and mitosis. Here we report that differential expression in cancer cells of a C-terminal splice variant of survivin, termed survivin-ΔEx3, is tightly associated with aggressive disease and markers of unfavorable prognosis. In contrast to other survivin variants, survivin-ΔEx3 localized exclusively to nuclei in tumor cells and was phosphorylated at multiple residues by the checkpoint kinase Chk2 during DNA damage.

Chemosensitization of cancer cells by KU-0060648, a dual inhibitor of DNA-PK and PI-3K.

DNA double-strand breaks (DSB) are the most cytotoxic lesions induced by topoisomerase II poisons. Nonhomologous end joining (NHEJ) is a major pathway for DSB repair and requires DNA-dependent protein kinase (DNA-PK) activity. DNA-PK catalytic subunit (DNA-PKcs) is structurally similar to PI-3K, which promotes cell survival and proliferation and is upregulated in many cancers.

Further characterisation of the cellular activity of the DNA-PK inhibitor, NU7441, reveals potential cross-talk with homologous recombination.

Purpose: Inhibition of DNA repair is emerging as a new therapeutic strategy for cancer treatment. One promising target is DNA-PK, a pivotal kinase in double-strand break repair. The purpose of this study was to further characterise the activity of the DNA-PK inhibitor NU7441, giving some new insights into the biology of DNA-PK.

Targeted inhibition of mitochondrial Hsp90 suppresses localised and metastatic prostate cancer growth in a genetic mouse model of disease.

Background: The molecular chaperone heat shock protein-90 (Hsp90) is a promising cancer drug target, but current Hsp90-based therapy has so far shown limited activity in the clinic.

Methods: We tested the efficacy of a novel mitochondrial-targeted, small-molecule Hsp90 inhibitor, Gamitrinib (GA mitochondrial matrix inhibitor), in the Transgenic Adenocarcinoma of the Mouse Prostate (TRAMP) model. The TRAMP mice receiving 3-week or 5-week systemic treatment with Gamitrinib were evaluated for localised or metastatic prostate cancer, prostatic intraepithelial neoplasia (PIN) or localised inflammation using magnetic resonance imaging, histology and immunohistochemistry.

Antitumor and anti-inflammatory effects of trabectedin on human myxoid liposarcoma cells.

Inflammatory mediators present in the tumor milieu may promote cancer progression and are considered promising targets of novel biological therapies. We previously reported that the marine antitumor agent trabectedin, approved in Europe in 2007 for soft tissue sarcomas and in 2009 for ovarian cancer, was able to downmodulate the production of selected cytokines/chemokines in immune cells. Patients with myxoid liposarcoma (MLS), a subtype characterized by the expression of the oncogenic transcript FUS-CHOP, are highly responsive to trabectedin.

The isothiocyanate produced from glucomoringin inhibits NF-kB and reduces myeloma growth in nude mice in vivo.

Glucosinolates (GLs), natural compounds extracted from Brassicaceae and precursors of isothiocyanates (ITCs), have been studied in the last decades mostly due to their chemopreventive activity and, more recently, for their potential use as novel chemotherapeutics. The aim of the present study was to investigate the in vitro and in vivo activity of glucomoringin (GMG), an uncommon member of the GLs family, and to compare it with glucoraphanin (GRA), one of the most studied GL. We have evaluated the potency of both compounds in inducing cell death, cell cycle perturbations, apoptosis, NF-kB inhibition and GST-pi activity in human carcinoma cells with different GST-pi contents as well as in human multiple myeloma and leukaemia cell lines.

Trabectedin (ET-743) promotes differentiation in myxoid liposarcoma tumors.

Differentiation is a complex set of events that can be blocked by rearrangements of regulatory genes producing fusion proteins with altered properties. In the case of myxoid liposarcoma (MLS) tumors, the causative abnormality is a fusion between the CHOP transcription factor and the FUS or EWS genes. CHOP belongs to and is a negative regulator of the large CAAT/enhancer binding protein family whose alpha, beta, and delta members are master genes of adipogenesis.

The expression of the DeltaNp73beta isoform of p73 leads to tetraploidy.

The p73 locus gene has a complex structure encoding a plethora of isoforms. The different DeltaN truncated isoforms of p73 may exert different activities depending on the cellular context. The beta isoform of DeltaNp73 seems to have a particular pattern of action even if its role in cell cycle and mitosis is still under investigation.

Atypical retinoids ST1926 and CD437 are S-phase-specific agents causing DNA double-strand breaks: significance for the cytotoxic and antiproliferative activity.

Retinoid-related molecules (RRM) are novel agents with tumor-selective cytotoxic/antiproliferative activity, a different mechanism of action from classic retinoids and no cross-resistance with other chemotherapeutics. ST1926 and CD437 are prototypic RRMs, with the former currently undergoing phase I clinical trials. We show here that ST1926, CD437, and active congeners cause DNA damage.

Checkpoint kinase 1 down-regulation by an inducible small interfering RNA expression system sensitized in vivo tumors to treatment with 5-fluorouracil.

Purpose: After DNA damage, checkpoints pathways are activated in the cells to halt the cell cycle, thus ensuring repair or inducing cell death. To better investigate the role of checkpoint kinase 1 (Chk1) in cellular response to different anticancer agents, Chk1 was knocked down in HCT-116 cell line and in its p53-deficient subline by using small interfering RNAs (siRNA).

Experimental Design: Chk1 was abrogated by transient transfection of specific siRNA against it, and stable tetracycline-inducible Chk1 siRNA clones were obtained transfecting cells with a plasmid expressing two siRNA against Chk1.

Role of homologous recombination in trabectedin-induced DNA damage.

Trabectedin (ET-743, Yondelis) is a natural marine compound with antitumour activity currently undergoing phase II/III clinical trials. The mechanism of the drug's action is still to be defined, even though it has been clearly demonstrated the key role of Nucleotide Excision Repair (NER). To get further insights into the drug's mode of action, we studied the involvement of the DNA-double strand break repair (DNA-DSB) pathways: homologous and non-homologous recombination, both in budding yeasts and in mammalian cells and the possible cross-talk between NER and these repair pathways.

Multi-parametric flow cytometric cell cycle analysis using TO-PRO-3 iodide (TP3): detailed protocols.

TO-PRO-3 iodide (TP3), a monomeric cyanine nucleic acid stain with a peak absorbance at 642 nm and emission at 661 nm, is best excited by a helium-neon (HeNe) laser (633nm). It was tested in monocytes and different cell lines under conditions of different fixatives, dye concentrations, labeling kinetics and RNAse concentrations for mono-, bi- and tri-parametric flow cytometric cell cycle analysis to establish the best protocol for DNA analysis in terms of G1 peak CV, G2/G1 ratio and minimal amount of debris. A linear increase in G1 peak position was found from 0.

Dynamics of cell cycle phase perturbations by trabectedin (ET-743) in nucleotide excision repair (NER)-deficient and NER-proficient cells, unravelled by a novel mathematical simulation approach.

Objectives: Trabectedin (ET-743, Yondelis) is a natural marine product, with antitumour activity, currently in phase II/III clinical trials. Previous studies have shown that cells hypersensitive to ultraviolet (UV)-rays because of nucleotide excision repair (NER) deficiency, were resistant to trabectedin. The purpose of this study was to investigate whether this resistance was associated with different drug-induced cell cycle perturbations.

Modulation of gene transcription by natural products--a viable anticancer strategy.

Drug design based on the structure of specific enzymes playing a role in carcinogenesis, e.g. tyrosine kinases, has been successful at identifying novel effective anticancer drugs.

Antitumor activity of the retinoid-related molecules (E)-3-(4'-hydroxy-3'-adamantylbiphenyl-4-yl)acrylic acid (ST1926) and 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid (CD437) in F9 teratocarcinoma: Role of retinoic acid receptor gamma and retinoid-independent pathways.

The retinoid-related molecules (RRMs) ST1926 [(E)-3-(4'-hydroxy-3'-adamantylbiphenyl-4-yl)acrylic acid] and CD437 (6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid) are promising anticancer agents. We compared the retinoic acid receptor (RAR) trans-activating properties of the two RRMs and all-trans-retinoic acid (ATRA). ST1926 and CD437 are better RARgamma agonists than ATRA.

New drugs from the sea.

This paper illustrates some biochemical and pharmacological properties of two natural marine products such as trabectedin (ET-743, Yondelis) and aplidine. Trabectedin has shown clinical antitumor activity in refractory soft tissue sarcoma and ovarian cancer. The lack of cross resistance of trabectedin with other chemotherapeutic drugs is presumably related to its peculiar mode of action, that seems to be related to a promoter-dependent transcription modulation.