Carol Ward

Randomized Phase II Trial of the Efficacy and Safety of Trastuzumab Combined With Docetaxel in Patients With Human Epidermal Growth Factor Receptor 2–Positive Metastatic Breast Cancer Administered As First-Line Treatment: The M77001 Study Group

PURPOSE This randomized, multicenter trial compared first-line trastuzumab plus docetaxel versus docetaxel alone in patients with human epidermal growth factor receptor 2 (HER2) -positive metastatic breast cancer (MBC). PATIENTS AND METHODS Patients were randomly assigned to six cycles of docetaxel 100 mg/m2 every 3 weeks, with or without trastuzumab 4 mg/kg loading dose followed by 2 mg/kg weekly until disease progression. RESULTS A total of 186 patients received at least one dose of the study drug. Trastuzumab plus docetaxel was significantly superior to docetaxel alone in terms of overall response rate (61% v 34%; P = .0002), overall survival (median, 31.2 v 22.7 months; P = .0325), time to disease progression (median, 11.7 v 6.1 months; P = .0001), time to treatment failure (median, 9.8 v 5.3 months; P = .0001), and duration of response (median, 11.7 v 5.7 months; P = .009). There was little difference in the number and severity of adverse events between the arms. Grade 3 to 4 neutropenia was seen more commonly with the combination (32%) than with docetaxel alone (22%), and there was a slightly higher incidence of febrile neutropenia in the combination arm (23% v 17%). One patient in the combination arm experienced symptomatic heart failure (1%). Another patient experienced symptomatic heart failure 5 months after discontinuation of trastuzumab because of disease progression, while being treated with an investigational anthracycline for 4 months. CONCLUSION Trastuzumab combined with docetaxel is superior to docetaxel alone as first-line treatment of patients with HER2-positive MBC in terms of overall survival, response rate, response duration, time to progression, and time to treatment failure, with little additional toxicity.

Prospective Molecular Marker Analyses of EGFR and KRAS From a Randomized, Placebo-Controlled Study of Erlotinib Maintenance Therapy in Advanced Non–Small-Cell Lung Cancer

PURPOSE The phase III, randomized, placebo-controlled Sequential Tarceva in Unresectable NSCLC (SATURN; BO18192) study found that erlotinib maintenance therapy extended progression-free survival (PFS) and overall survival in patients with advanced non-small-cell lung cancer (NSCLC) who had nonprogressive disease following first-line platinum-doublet chemotherapy. This study included prospective analysis of the prognostic and predictive value of several biomarkers. PATIENTS AND METHODS Mandatory diagnostic tumor specimens were collected before initiating first-line chemotherapy and were tested for epidermal growth factor receptor (EGFR) protein expression by using immunohistochemistry (IHC), EGFR gene copy number by using fluorescent in situ hybridization (FISH), and EGFR and KRAS mutations by using DNA sequencing. An EGFR CA simple sequence repeat in intron 1 (CA-SSR1) polymorphism was evaluated in blood. RESULTS All 889 randomly assigned patients provided tumor samples. EGFR IHC, EGFR FISH, KRAS mutation, and EGFR CA-SSR1 repeat length status were not predictive for erlotinib efficacy. A profound predictive effect on PFS of erlotinib relative to placebo was observed in the EGFR mutation-positive subgroup (hazard ratio [HR], 0.10; P < .001). Significant PFS benefits were also observed with erlotinib in the wild-type EGFR subgroup (HR, 0.78; P = .0185). KRAS mutation status was a significant negative prognostic factor for PFS. CONCLUSION This large prospective biomarker study found that patients with activating EGFR mutations derive the greatest PFS benefit from erlotinib maintenance therapy. No other biomarkers were predictive for outcomes with erlotinib, although the study was not powered for clinical outcomes in biomarker subgroups other than EGFR IHC-positive [corrected]. KRAS mutations were prognostic for reduced PFS. The study demonstrated the feasibility of prospective tissue collection for biomarker analyses in NSCLC.

Pharmacological manipulation of granulocyte apoptosis: potential therapeutic targets

Resolution of inflammation involves the clearance of excess or effete inflammatory cells by a process of physiological programmed cell death (apoptosis) and the subsequent recognition and removal of apoptotic cells by phagocytes. The therapeutic induction of apoptosis for the resolution of chronic inflammation and the general pharmacology of apoptosis have become subjects of increasing interest. In this article, some of the unique and important differences in the control of apoptosis of various inflammatory cells (particularly neutrophil and eosinophil granulocytes) are highlighted. It is suggested that apoptosis can be specifically regulated pharmacologically and could be exploited to develop new drug therapies.

New strategies for targeting the hypoxic tumour microenvironment in breast cancer

Radiation and drug resistance remain major challenges and causes of mortality in the treatment of locally advanced, recurrent and metastatic breast cancer. Metabolic reprogramming is a recently recognised hallmark of cancer with the hypoxic acidic extracellular environment as a major driver of invasion and metastases. Nearly 40% of all breast cancers and 50% of locally advanced breast cancers are hypoxic and their altered metabolism is strongly linked to resistance to radiotherapy and systemic therapy. The dependence of metabolically adapted breast cancer cells on alterations in cell function presents a wide range of new therapeutic targets such as carbonic anhydrase IX (CAIX). This review highlights preclinical approaches to evaluating an array of targets against tumour metabolism in breast cancer and early phase clinical studies on efficacy.

Prostaglandin D2 and Its Metabolites Induce Caspase-Dependent Granulocyte Apoptosis That Is Mediated Via Inhibition of IκBα Degradation Using a Peroxisome Proliferator-Activated Receptor-γ-Independent Mechanism

Many inflammatory mediators retard granulocyte apoptosis. Most natural PGs studied herein (e.g., PGE2, PGA2, PGA1, PGF2α) either delayed apoptosis or had no effect, whereas PGD2 and its metabolite PGJ2 selectively induced eosinophil, but not neutrophil apoptosis. This novel proapoptotic effect does not appear to be mediated via classical PG receptor ligation or by elevation of intracellular cAMP or Ca2+. Intriguingly, the sequential metabolites Δ12PGJ2 and 15-deoxy-Δ12, Δ14-PGJ2 (15dPGJ2) induced caspase-dependent apoptosis in both granulocytes, an effect that did not involve de novo protein synthesis. Despite the fact that Δ12PGJ2 and 15dPGJ2 are peroxisome proliferator-activated receptor-γ (PPAR-γ) activators, apoptosis was not mimicked by synthetic PPAR-γ and PPAR-α ligands or blocked by an irreversible PPAR-γ antagonist. Furthermore, Δ12PGJ2 and 15dPGJ2 inhibited LPS-induced IκBα degradation and subsequent inhibition of neutrophil apoptosis, suggesting that apoptosis is mediated via PPAR-γ-independent inhibition of NF-κB activation. In addition, we show that TNF-α-mediated loss of cytoplasmic IκBα in eosinophils is inhibited by 15dPGJ2 in a concentration-dependent manner. The selective induction of eosinophil apoptosis by PGD2 and PGJ2 may help define novel therapeutic pathways in diseases in which it would be desirable to specifically remove eosinophils but retain neutrophils for antibacterial host defense. The powerful proapoptotic effects of Δ12PGJ2 and 15dPGJ2 in both granulocyte types suggest that these natural products control the longevity of key inflammatory cells and may be relevant to understanding the control and resolution of inflammation.

WWOX gene expression abolishes ovarian cancer tumorigenicity in vivo and decreases attachment to fibronectin via integrin alpha3.

The WW domain-containing oxidoreductase (WWOX) gene is located at FRA16D, a common fragile site involved in human cancer. Targeted deletion of Wwox in mice causes increased spontaneous tumor incidence, confirming that WWOX is a bona fide tumor suppressor gene. We show that stable transfection of WWOX into human PEO1 ovarian cancer cells, containing homozygous WWOX deletion, abolishes in vivo tumorigenicity, but this does not correlate with alteration of in vitro growth. Rather, WWOX restoration in PEO1, or WWOX overexpression in SKOV3 ovarian cancer cells, results in reduced attachment and migration on fibronectin, an extracellular matrix component linked to peritoneal metastasis. Conversely, siRNA-mediated knockdown of endogenous WWOX in A2780 ovarian cancer cells increases adhesion to fibronectin. In addition, whereas there is no WWOX-dependent difference in cell death in adherent cells, WWOX-transfected cells in suspension culture display a proapoptotic phenotype. We further show that WWOX expression reduces membranous integrin alpha(3) protein but not integrin alpha(3) mRNA levels, and that adhesion of PEO1 cells to fibronectin is predominantly mediated through integrin alpha(3). We therefore propose that WWOX acts as an ovarian tumor suppressor by modulating the interaction between tumor cells and the extracellular matrix and by inducing apoptosis in detached cells. Consistent with this, the suppression of PEO1 tumorigenicity by WWOX can be partially overcome by implanting these tumor cells in Matrigel. These data suggest a possible role for the loss of WWOX in the peritoneal dissemination of human ovarian cancer cells.

Inhibition of nuclear factor-κB activation un-masks the ability of TNF-α to induce human eosinophil apoptosis

Apoptosis renders eosinophils functionally effete and marks them for ‘silent’ removal from inflamed sites by macrophages. We show, for the first time, that eosinophils exposed to TNF‐α rapidly lose their cytoplasmic levels of IκBα, the inhibitory subunit of NF‐κB. Consequently, TNF‐α triggers NF‐κB mobilization from the cytoplasm to the nucleus, as determined by tracking the NF‐κB subunit p65 by immunofluorescence and Western blot analysis. Inhibition of TNF‐α‐mediated IκBα degradation and NF‐κB activation by gliotoxin or the proteasome inhibitor MG‐132 un‐masks the caspase‐dependent pro‐apoptotic properties of TNF‐α. In addition, cycloheximide similarly renders TNF‐α pro‐apoptotic, suggesting that NF‐κB activation controls the production of a protein(s) that protects eosinophils from the cytotoxic effects of TNF‐α. Evidence is presented suggesting that TNF‐α triggered apoptosis is more susceptible to NF‐κB inhibition than constitutive apoptosis, leading to the possibility of the specific targeting of apoptosis in eosinophil sub‐populations. Prior to morphological signs of apoptosis, TNF‐α‐induced IL‐8 synthesis is abrogated by inhibition of NF‐κB. We propose that NF‐κB activation plays a critical role in controlling eosinophil responsiveness and apoptosis, and speculate that selective inhibitors of eosinophil NF‐κB activation may ultimately provide alternative therapeutic agents for the treatment of eosinophilic diseases, including asthma and allergic rhinitis.

Targeting tumour hypoxia to prevent cancer metastasis: from biology, biosensing and technology to drug development : the METOXIA consortium

Abstract The hypoxic areas of solid cancers represent a negative prognostic factor irrespective of which treatment modality is chosen for the patient. Still, after almost 80 years of focus on the problems created by hypoxia in solid tumours, we still largely lack methods to deal efficiently with these treatment-resistant cells. The consequences of this lack may be serious for many patients: Not only is there a negative correlation between the hypoxic fraction in tumours and the outcome of radiotherapy as well as many types of chemotherapy, a correlation has been shown between the hypoxic fraction in tumours and cancer metastasis. Thus, on a fundamental basis the great variety of problems related to hypoxia in cancer treatment has to do with the broad range of functions oxygen (and lack of oxygen) have in cells and tissues. Therefore, activation–deactivation of oxygen-regulated cascades related to metabolism or external signalling are important areas for the identification of mechanisms as potential targets for hypoxia-specific treatment. Also the chemistry related to reactive oxygen radicals (ROS) and the biological handling of ROS are part of the problem complex. The problem is further complicated by the great variety in oxygen concentrations found in tissues. For tumour hypoxia to be used as a marker for individualisation of treatment there is a need for non-invasive methods to measure oxygen routinely in patient tumours. A large-scale collaborative EU-financed project 2009–2014 denoted METOXIA has studied all the mentioned aspects of hypoxia with the aim of selecting potential targets for new hypoxia-specific therapy and develop the first stage of tests for this therapy. A new non-invasive PET-imaging method based on the 2-nitroimidazole [18F]-HX4 was found to be promising in a clinical trial on NSCLC patients. New preclinical models for testing of the metastatic potential of cells were developed, both in vitro (2D as well as 3D models) and in mice (orthotopic grafting). Low density quantitative real-time polymerase chain reaction (qPCR)-based assays were developed measuring multiple hypoxia-responsive markers in parallel to identify tumour hypoxia-related patterns of gene expression. As possible targets for new therapy two main regulatory cascades were prioritised: The hypoxia-inducible-factor (HIF)-regulated cascades operating at moderate to weak hypoxia (<1% O2), and the unfolded protein response (UPR) activated by endoplasmatic reticulum (ER) stress and operating at more severe hypoxia (<0.2%). The prioritised targets were the HIF-regulated proteins carbonic anhydrase IX (CAIX), the lactate transporter MCT4 and the PERK/eIF2α/ATF4-arm of the UPR. The METOXIA project has developed patented compounds targeting CAIX with a preclinical documented effect. Since hypoxia-specific treatments alone are not curative they will have to be combined with traditional anti-cancer therapy to eradicate the aerobic cancer cell population as well.

Interleukin-10 inhibits lipopolysaccharide-induced survival and extracellular signal-regulated kinase activation in human neutrophils

Lipopolysaccharide (LPS) induces a marked delay in human neutrophil apoptosis that is reversed by the anti‐inflammatory cytokine IL‐10. The effect of IL‐10 is specific since other agents that delay neutrophil apoptosis are not affected. To investigate mechanisms underlying the actions of IL‐10, we examined signaling pathways activated by LPS per se and in response to IL‐10. The MAPK kinase (MEK) 1 inhibitor PD098059, the protein kinase C (PKC) inhibitor Ro31,8220, and the phosphatidylinositol‐3 kinase (PI3‐K) inhibitor LY294002 all partially reversed LPS‐mediated retardation of neutrophil apoptosis, but the p38 MAPK inhibitor SB203850 did not. LPS activates the transcription factor NF‐κB, however, IL‐10 did not affect the ability of LPS to activate NF‐κB as assessed by IκB‐α proteolysis. Although IL‐10 did not alter activation of ERK by GM‐CSF or TNF‐α, it did inhibit activation induced by LPS. Thus our data illustrate that LPS‐induced neutrophil survival is regulated by the MAPK, PKC and PI3‐K pathways as well as NF‐κB, and can be reversed by IL‐10, through a mechanism involving inhibition of ERK activation. Because of the specific nature of this inhibition, we conclude that IL‐10 interferes with an ERK activation pathway, which is not involved in GM‐CSF or TNF‐α signaling.

Regulation of neutrophil apoptosis and removal of apoptotic cells.

The accumulation of neutrophils during inflammation is essential for the destruction and removal of invading microorganisms. However, for resolution of inflammation to occur, neutrophils must also be removed from the inflammatory site since these cells are capable of releasing tissue toxic molecules. Neutrophil removal has been shown to occur via apoptosis and phagocyte clearance of apoptotic cells. Therefore, manipulation of these processes is likely to be a key therapeutic strategy in the management of inflammatory disease. In this review, we examine mediators of neutrophil survival and apoptosis and the signalling pathways that regulate the balance between life and death in these cells.