Pharmacokinetic characterization of amrubicin cardiac safety in an ex vivo human myocardial strip model. II. Amrubicin shows metabolic advantages over doxorubicin and epirubicin.

Anthracycline-related cardiotoxicity correlates with cardiac anthracycline accumulation and bioactivation to secondary alcohol metabolites or reactive oxygen species (ROS), such as superoxide anion (O₂·⁻) and hydrogen peroxide H₂O₂). We reported that in an ex vivo human myocardial strip model, 3 or 10 μM amrubicin [(7S,9S)-9-acetyl-9-amino-7-[(2-deoxy-β-D-erythro-pentopyranosyl)oxy]-7,8,9,10-tetrahydro-6,11-dihydroxy-5,12-napthacenedione hydrochloride] accumulated to a lower level compared with equimolar doxorubicin or epirubicin (J Pharmacol Exp Ther 341:464-473, 2012). We have characterized how amrubicin converted to ROS or secondary alcohol metabolite in comparison with doxorubicin (that formed both toxic species) or epirubicin (that lacked ROS formation and showed an impaired conversion to alcohol metabolite).

Pharmacokinetic characterization of amrubicin cardiac safety in an ex vivo human myocardial strip model. I. Amrubicin accumulates to a lower level than doxorubicin or epirubicin.

Antitumor anthracyclines such as doxorubicin and epirubicin are known to cause cardiotoxicity that correlates with anthracycline accumulation in the heart. The anthracycline amrubicin [(7S,9S)-9-acetyl-9-amino-7-[(2-deoxy-β-d-erythro-pentopyranosyl)oxy]-7,8,9,10-tetrahydro-6,11-dihydroxy-5,12-napthacenedione hydrochloride] has not shown cardiotoxicity in laboratory animals or patients in approved or investigational settings; therefore, we conducted preclinical work to characterize whether amrubicin attained lower levels than doxorubicin or epirubicin in the heart. Anthracyclines were evaluated in ex vivo human myocardial strips incubated in plasma to which anthracycline concentrations of 3 or 10 μM were added.

Increased cellular accumulation and distribution of amrubicin contribute to its activity in anthracycline-resistant cancer cells.

Purpose: Multi-drug resistance and cumulative cardiotoxicity are major limitations for the clinical use of anthracyclines. Here, we evaluated and compared the cross-resistance of amrubicin, a third-generation synthetic anthracycline and potent topoisomerase (topo)-II inhibitor with little or no observed cardiotoxicity to other anthracyclines and the topo-II inhibitor etoposide in drug-resistant tumor models in order to elucidate its potential mechanisms of action.

Methods: Amrubicin activity was assessed in multi-drug-resistant cell lines and human tumor explants using cytotoxicity assays, confocal microscopy, fluorescence time-lapse imaging, flow cytometry, immunoblotting, and gene expression profiling techniques.

A comparison of azacitidine and decitabine activities in acute myeloid leukemia cell lines.

Background: The cytidine nucleoside analogs azacitidine (AZA) and decitabine (DAC) are used for the treatment of patients with myelodysplastic syndromes and acute myeloid leukemia (AML). Few non-clinical studies have directly compared the mechanisms of action of these agents in a head-to-head fashion, and the agents are often viewed as mechanistically similar DNA hypomethylating agents. To better understand the similarities and differences in mechanisms of these drugs, we compared their in vitro effects on several end points in human AML cell lines.

The antileukemia activity of a human anti-CD40 antagonist antibody, HCD122, on human chronic lymphocytic leukemia cells.

B-cell chronic lymphocytic leukemia (B-CLL) is a lymphoproliferative disorder characterized by the surface expression of CD20, CD5 antigens, as well as the receptor CD40. Activation of CD40 by its ligand (CD40L) induces proliferation and rescues the cells from spontaneous and chemotherapy-induced apoptosis. CD40 activation also induces secretion of cytokines, such as IL-6, IL-10, TNF-alpha, IL-8, and GM-CSF, which are involved in tumor cell survival, migration, and interaction with cells in the tumor microenvironment.

Recombinant interleukin-2 significantly augments activity of rituximab in human tumor xenograft models of B-cell non-Hodgkin lymphoma.

Recombinant interleukin-2 (rIL-2) is a pleiotropic cytokine that activates select immune effector cell responses associated with antitumor activity, including antibody-dependent cellular cytotoxicity (ADCC). Rituximab is an anti-CD20 monoclonal antibody that activates ADCC in non-Hodgkin lymphoma (NHL). The ability of rIL-2 to augment rituximab-dependent tumor responses was investigated.

Novel robust hepatitis C virus mouse efficacy model.

The lack of a robust small-animal model for hepatitis C virus (HCV) has hindered the discovery and development of novel drug treatments for HCV infections. We developed a reproducible and easily accessible xenograft mouse efficacy model in which HCV RNA replication is accurately monitored in vivo by real-time, noninvasive whole-body imaging of gamma-irradiated SCID mice implanted with a mouse-adapted luciferase replicon-containing Huh-7 cell line (T7-11). The model was validated by demonstrating that both a small-molecule NS3/4A protease inhibitor (BILN 2061) and human alpha interferon (IFN-alpha) decreased HCV RNA replication and that treatment withdrawal resulted in a rebound in replication, which paralleled clinical outcomes in humans.

CHIR-258: a potent inhibitor of FLT3 kinase in experimental tumor xenograft models of human acute myelogenous leukemia.

Purpose: Fms-like tyrosine kinase 3 (FLT3) encodes a receptor tyrosine kinase (RTK) for which activating mutations have been identified in a proportion of acute myelogenous leukemia (AML) patients and associated with poor clinical prognosis. Given the relevance of FLT3 mutations in AML, we investigated the activity of CHIR-258, an orally active, multitargeted small molecule, with potent activity against FLT3 kinase and class III, IV, and V RTKs involved in endothelial and tumor cell proliferation in AML models.

Experimental Design: CHIR-258 was tested on two human leukemic cell lines in vitro and in vivo with differing FLT3 mutational status [MV4;11 cells express FLT3 internal tandem duplications (ITD) versus RS4;11 cells with wild-type (WT) FLT3].

In vivo target modulation and biological activity of CHIR-258, a multitargeted growth factor receptor kinase inhibitor, in colon cancer models.

Purpose: To evaluate the therapeutic and biological effects of CHIR-258, an orally bioavailable, potent inhibitor of class III-V receptor tyrosine kinases, in colon cancer models.

Experimental Design: The pharmacologic activity of CHIR-258 was characterized by monitoring target modulation as well as by evaluating the antitumor and antiangiogenic effects in human colon xenograft models.

Results: CHIR-258 inhibits vascular endothelial growth factor receptor 1/2, fibroblast growth factor receptor 1/3, and platelet-derived growth factor receptor beta (PDGFRbeta) and shows both antitumor and antiangiogenic activities in vivo.

beta-Catenin regulates vascular endothelial growth factor expression in colon cancer.

To evaluate whether beta-catenin signaling has a role in the regulation of angiogenesis in colon cancer, a series of angiogenesis-related gene promoters was analyzed for beta-catenin/TCF binding sites. Strikingly, the gene promoter of human vascular endothelial growth factor (VEGF, or VEGF-A) contains seven consensus binding sites for beta-catenin/TCF. Analysis of laser capture microdissected human colon cancer tissue indicated a direct correlation between up-regulation of VEGF-A expression and adenomatous polyposis coli (APC) mutational status (activation of beta-catenin signaling) in primary tumors.

Matrix immobilization enhances the tissue repair activity of growth factor gene therapy vectors.

Although growth factor proteins display potent tissue repair activities, difficulty in sustaining localized therapeutic concentrations limits their therapeutic activity. We reasoned that enhanced histogenesis might be achieved by combining growth factor genes with biocompatible matrices capable of immobilizing vectors at delivery sites. When delivered to subcutaneously implanted sponges, a platelet-derived growth factor B-encoding adenovirus (AdPDGF-B) formulated in a collagen matrix enhanced granulation tissue deposition 3- to 4-fold (p < or = 0.

Fibroblast growth factor 2-retargeted adenoviral vectors exhibit a modified biolocalization pattern and display reduced toxicity relative to native adenoviral vectors.

Targeted vectors provide a number of advantages for systemic and local gene delivery strategies. Several groups have investigated the utility of using various ligands to alter the tropism of adenovirus (Ad) vectors. We have previously demonstrated that fibroblast growth factor (FGF) ligands can specifically target DNA transfection and Ad transduction through high-affinity FGF receptors (FGFRs).

Fibroblast growth factor 2 retargeted adenovirus has redirected cellular tropism: evidence for reduced toxicity and enhanced antitumor activity in mice.

Adenovirus (Ad) have been used as vectors to deliver genes to a wide variety of tissues. Despite achieving high expression levels in vivo, Ad vectors display normal tissue toxicity, transient expression, and antivector immune responses that limit therapeutic potential. To circumvent these problems, several retargeting strategies to abrogate native tropism and redirect Ad uptake through defined receptors have been attempted.

Prevalent expression of fibroblast growth factor (FGF) receptors and FGF2 in human tumor cell lines.

Basic fibroblast growth factor (FGF2) has potent mitogenic and angiogenic activities that have been implicated in tumor development and malignant progression. The biological effects of FGF2 and other members of the FGF ligand family are mediated by 4 transmembrane tyrosine kinase receptors (FGFRs). To better understand the roles of FGFRs in cancer, the expression of FGF2 and each of the 4 FGFRs was assessed by RNase protection analysis of 60 human tumor cell lines, representing 9 tumor types.

Targeting tumor cells via EGF receptors: selective toxicity of an HBEGF-toxin fusion protein.

Over-expression of the epidermal growth factor receptor (EGFR) is a hallmark of numerous solid tumors, thus providing a means of selectively targeting therapeutic agents. Heparin-binding epidermal growth factor (HBEGF) binds to EGFRs with high affinity and to heparan sulfate proteoglycans, resulting in increased mitogenic potential compared to other EGF family members. We have investigated the feasibility of using HBEGF to selectively deliver a cytotoxic protein into EGFR-expressing tumor cells.

Keratinocyte growth factor protects mice from chemotherapy and radiation-induced gastrointestinal injury and mortality.

Keratinocyte growth factor (KGF) stimulates the proliferation and differentiation of epithelial cells including those of the gastrointestinal tract. Although chemotherapeutics and radiation exposure kill rapidly proliferating tumor cells, rapidly dividing normal cells of the host's gastrointestinal tract are also frequently damaged, leading to the clinical condition broadly termed "mucositis." In this report, recombinant human KGF used as a pretreatment in several mouse models of chemotherapy and/or radiation-induced gastrointestinal injury significantly improved mouse survival.

Keratinocyte growth factor ameliorates mucosal injury in an experimental model of colitis in rats.

Background & Aims: Keratinocyte growth factor (KGF) is known to enhance tissue repair in the skin; however, its role in the gastrointestinal tract is largely unknown. The aim of this study was to evaluate the effects of exogenous KGF in an experimental model of colitis in rats.

Methods: KGF was administered before or after induction of colitis with 2,4,6-trinitrobenzenesulfonic acid/ethanol.

Impaired tumor growth in colony-stimulating factor 1 (CSF-1)-deficient, macrophage-deficient op/op mouse: evidence for a role of CSF-1-dependent macrophages in formation of tumor stroma.

Macrophages have been suggested to play a major role in the immune response to cancer. They have also been suggested to stimulate the formation of tumor stroma and to promote tumor vascularization. The availability of the op/op mouse, which has no endogenous colony-stimulating factor 1 (CSF-1) and which possesses a profound macrophage deficiency, provides a new model to verify these notions.

Keratinocyte growth factor causes cystic dilation of the mammary glands of mice. Interactions of keratinocyte growth factor, estrogen, and progesterone in vivo.

Keratinocyte growth factor (KGF) is a paracrine mediator of epithelial cell proliferation that has been reported to induce marked proliferation of mammary epithelium in rats. In this study, systemic administration of KGF into naive and oophorectomized mice causes mammary gland proliferation, as evidenced histologically by the appearance of cysts lined by a single layer of epithelium and by hyperplastic epithelium. Whole mount preparations of the mammary glands reveal that the histologically noted cysts are actually ducts that are dilated along much of their length.

Keratinocyte growth factor induces proliferation of hepatocytes and epithelial cells throughout the rat gastrointestinal tract.

Keratinocyte growth factor (KGF), a member of the fibroblast growth factor (FGF) family, was identified as a specific keratinocyte mitogen after isolation from a lung fibroblast line. Recently, recombinant (r)KGF was found to influence proliferation and differentiation patterns of multiple epithelial cell lineages within skin, lung, and the reproductive tract. In the present study, we designed experiments to identify additional target tissues, and focused on the rat gastrointestinal (GI) system, since a putative receptor, K-sam, was originally identified in a gastric carcinoma.

Keratinocyte growth factor induces pancreatic ductal epithelial proliferation.

Keratinocyte growth factor (KGF) causes a proliferation of pancreatic ductal epithelial cells in adult rats after daily systemic administration for 1 to 2 weeks. Even before the proliferation of intralobular ducts is histologically evident, KGF also induces proliferating cell nuclear antigen expression within the ductal epithelium of intercalated, intralobular, and interlobular ducts. KGF also causes incorporation of 5-bromodeoxyuridine in ductal epithelial cells.

Antifungal activity of recombinant human macrophage colony-stimulating factor in models of acute and chronic candidiasis in the rat.

Models of acute and chronic candidiasis were developed in Fischer 344 rats to evaluate the therapeutic efficacy of recombinant human macrophage colony-stimulating factor (rhM-CSF) alone and in combination with the antifungal agent fluconazole. In the acute model, rats were challenged by intravenous injection with 2 x 10(6) Candida albicans, approximately 4 times the LD50. Daily subcutaneous (sc) bolus injections of rhM-CSF for 10 days plus a single sc bolus dose of 0.

Effect of macrophage-modulating agents on in vivo growth of transplantable Lewis lung cancer in mice.

C57Bl/6 mice, bearing transplantable Lewis lung cancer (non-metastatic subline) implanted either subcutaneously or intraperitoneally were treated with macrophage colony stimulating factor (M-CSF, 10(6) units per mouse, per day for 19 days), Escherichia coli lipopolysaccharide or both. Lipopolysaccharide (5 micrograms per mouse) administered daily once a day for up to 30 days impaired both subcutaneous and intraperitoneal tumor growth and prolonged survival of tumor bearing mice. Macrophage colony stimulating factor, administered daily, inhibited only subcutaneous tumor growth, both when administered alone and in combination with with lipopolysaccharide, and had no effect on intraperitoneal tumor.

Antitumor protection from the murine T-cell leukemia/lymphoma EL4 by the continuous subcutaneous coadministration of recombinant macrophage-colony stimulating factor and interleukin-2.

Combined continuous s.c. coadministration of macrophage-colony stimulating factor (M-CSF) plus interleukin-2 (IL-2) by osmotic pump protected mice given i.

The skeletal effects of colony-stimulating factor-1 in toothless (osteopetrotic) rats: persistent metaphyseal sclerosis and the failure to restore subepiphyseal osteoclasts.

Toothless (tl), one of four osteopetrotic mutations in the rat, is characterized by few osteoclasts, undetectable bone resorption, and failure of correction by bone marrow transplantation. We recently reported that CSF-1 treatment improves these skeletal problems but that metaphyseal sclerosis persists. In the present study we demonstrate that optimal reduction of the skeletal sclerosis in tl rats following CSF-1 treatment has lower and upper dosage thresholds and that skeletal sclerosis returns after CSF-1 withdrawal.