New HER2-negative breast cancer subtype responsive to anti-HER2 therapy identified.

PURPOSE: HER2 signaling functional activity may be important to measure in addition to HER2 protein quantification when identifying patients eligible for HER2 therapies. A HER2 Signaling Function (CELx HSF) Test for HER2-negative patients uses patient's live tumor cells on a biosensor to identify patients with abnormally high HER2-related signaling (HSFs+) likely to respond to anti-HER2 therapies. METHODS: The CELx HSF test was employed to: (1) characterize the sensitivity and specificity of the test to detect abnormal levels of HER2 signaling; (2) evaluate the inhibitory effectiveness of five different anti-HER2 therapies; (3) assess the correlation between CELx HSF test detection of abnormal HER2 signaling and response to HER2 therapy using xenograft models; and (4) confirm the prevalence of abnormal HER2 signaling amongst HER2-negative breast cancer patients (HER2-/HSFs+). RESULTS: HER2-/HSFs+ breast cancer patient samples were identified and showed sensitivity to five approved anti-HER2 therapies. Xenograft studies using both HER2+ and HER2- cell lines confirmed that CELx HER2 signaling status better predicts HER2 inhibitor efficacy than HER2 receptor status. In a study of 114 HER2-negative breast tumor patient samples, 27 (23.7%; 95% CI = 17-32%) had abnormal HER2 signaling (HSFs+). A ROC curve constructed with this dataset projects the CELx HSF Test would have greater than 90% sensitivity and specificity to detect the HER2-/HSFs+ patient population. CONCLUSIONS: The CELx HSF test is a well-characterized functional biomarker assay capable of identifying dynamic HER2-driven signaling dysfunction in tumor cells from HER2-negative breast cancer patients. This test has demonstrated efficacy of various HER2 targeted therapies in live tumor cells from the HSFs+ population and correlated the test result to HER2 drug response in mouse xenograft studies. The proportion of HER2-negative breast cancer patients found to have abnormal HER2 signaling in a 114 patient sample study, 20-25%, is significant. A clinical trial to evaluate the efficacy of anti-HER2 therapies in this patient population is warranted.

Bioavailability, intracellular mobilization of nickel, and HIF-1α activation in human lung epithelial cells exposed to metallic nickel and nickel oxide nanoparticles.

Micron-sized particles of poorly soluble nickel compounds, but not metallic nickel, are established human and rodent carcinogens. In contrast, little is known about the toxic effects of a growing number of Ni-containing materials in the nano-sized range. Here, we performed physicochemical characterization of NiO and metallic Ni nanoparticles and examined their metal ion bioavailability and toxicological properties in human lung epithelial cells. Cellular uptake of metallic Ni and NiO nanoparticles, but not metallic Ni microparticles, was associated with the release of Ni(II) ions after 24-48 h as determined by Newport Green fluorescence. Similar to soluble NiCl2, NiO nanoparticles induced stabilization and nuclear translocation of hypoxia-inducible factor 1α (HIF-1α) transcription factor followed by upregulation of its target NRDG1 (Cap43). In contrast to no response to metallic Ni microparticles, nickel nanoparticles caused a rapid and prolonged activation of the HIF-1α pathway that was stronger than that induced by soluble Ni(II). Soluble NiCl2 and NiO nanoparticles were equally toxic to H460 human lung epithelial cells and primary human bronchial epithelial cells; metallic Ni nanoparticles showed lower toxicity and Ni microparticles were nontoxic. Cytotoxicity induced by all forms of Ni occurred concomitant with activation of an apoptotic response, as determined by dose- and time-dependent cleavage of caspases and poly (ADP-ribose) polymerase. Our results show that metallic Ni nanoparticles, in contrast to micron-sized Ni particles, activate a toxicity pathway characteristic of carcinogenic Ni compounds. Moderate cytotoxicity and sustained activation of the HIF-1α pathway by metallic Ni nanoparticles could promote cell transformation and tumor progression.

XRCC1 deficiency sensitizes human lung epithelial cells to genotoxicity by crocidolite asbestos and Libby amphibole.

BACKGROUND: Asbestos induces DNA and chromosomal damage, but the DNA repair pathways protecting human cells against its genotoxicity are largely unknown. Polymorphisms in XRCC1 have been associated with altered susceptibility to asbestos-related diseases. However, it is unclear whether oxidative DNA damage repaired by XRCC1 contributes to asbestos-induced chromosomal damage. OBJECTIVES: We sought to examine the importance of XRCC1 in protection against genotoxic effects of crocidolite and Libby amphibole asbestos. METHODS: We developed a genetic model of XRCC1 deficiency in human lung epithelial H460 cells and evaluated genotoxic responses to carcinogenic fibers (crocidolite asbestos, Libby amphibole) and nongenotoxic materials (wollastonite, titanium dioxide). RESULTS: XRCC1 knockdown sensitized cells to the clastogenic and cytotoxic effects of oxidants [hydrogen peroxide (H2O2), bleomycin] but not to the nonoxidant paclitaxel. XRCC1 knockdown strongly enhanced genotoxicity of amphibole fibers as evidenced by elevated formation of clastogenic micronuclei. Crocidolite induced primarily clastogenic micronuclei, whereas Libby amphibole induced both clastogenic and aneugenic micronuclei. Crocidolite and bleomycin were potent inducers of nuclear buds, which were enhanced by XRCC1 deficiency. Libby amphibole and H2O2 did not induce nuclear buds, irrespective of XRCC1 status. Crocidolite and Libby amphibole similarly activated the p53 pathway. CONCLUSIONS: Oxidative DNA damage repaired by XRCC1 (oxidized bases, single-strand breaks) is a major cause of chromosomal breaks induced by crocidolite and Libby amphibole. Nuclear buds are a novel biomarker of genetic damage induced by exposure to crocidolite asbestos, which we suggest are associated with clustered DNA damage. These results provide mechanistic evidence for the epidemiological association between XRCC1 polymorphisms and susceptibility to asbestos-related disease.

Biopersistence and potential adverse health impacts of fibrous nanomaterials: what have we learned from asbestos?

Human diseases associated with exposure to asbestos fibers include pleural fibrosis and plaques, pulmonary fibrosis (asbestosis), lung cancer, and diffuse malignant mesothelioma. The critical determinants of fiber bioactivity and toxicity include not only fiber dimensions, but also shape, surface reactivity, crystallinity, chemical composition, and presence of transition metals. Depending on their size and dimensions, inhaled fibers can penetrate the respiratory tract to the distal airways and into the alveolar spaces. Fibers can be cleared by several mechanisms, including the mucociliary escalator, engulfment, and removal by macrophages, or through splitting and chemical modification. Biopersistence of long asbestos fibers can lead to inflammation, granuloma formation, fibrosis, and cancer. Exposure to synthetic carbon nanomaterials, including carbon nanofibers and carbon nanotubes (CNTs), is considered a potential health hazard because of their physical similarities with asbestos fibers. Respiratory exposure to CNTs can produce an inflammatory response, diffuse interstitial fibrosis, and formation of fibrotic granulomas similar to that observed in asbestos-exposed animals and humans. Given the known cytotoxic and carcinogenic properties of asbestos fibers, toxicity of fibrous nanomaterials is a topic of intense study. The mechanisms of nanomaterial toxicity remain to be fully elucidated, but recent evidence suggests points of similarity with asbestos fibers, including a role for generation of reactive oxygen species, oxidative stress, and genotoxicity. Considering the rapid increase in production and use of fibrous nanomaterials, it is imperative to gain a thorough understanding of their biologic activity to avoid the human health catastrophe that has resulted from widespread use of asbestos fibers.

SV40 oncoproteins enhance asbestos-induced DNA double-strand breaks and abrogate senescence in murine mesothelial cells.

SV40 virus has emerged as a potential cofactor with asbestos in the development of diffuse malignant mesothelioma, but its precise role in the pathogenesis of this tumor is unclear. SV40 large T antigen is known to inactivate cellular proteins involved in DNA damage and senescence, including p53 and pRb. We hypothesize that SV40 oncoproteins will sensitize mesothelial cells to DNA damage induced by asbestos or chemotherapeutic agents. SV40 oncoprotein expression in murine mesothelial cell lines enhanced spontaneous and asbestos-induced double-strand breaks, indicated by gamma-H2AX foci, and potentiated micronucleus formation. Mesothelial cells exposed to asbestos or bleomycin for 96 h acquired senescent-like morphology and displayed elevated senescence-associated beta-galactosidase activity, reduced bromodeoxyuridine (BrdUrd) incorporation, and reduced colony formation. SV40 oncoprotein expression abrogated the senescent phenotype, and transfected cell lines showed an increase in both BrdUrd incorporation and colony formation after prolonged DNA damage. Murine mesothelial cell lines lacking wild-type p53 due to a point mutation or gene rearrangement also failed to senesce in response to asbestos or chemotherapeutic agents. In addition, stress-induced senescence in human mesothelial cell lines was impaired by SV40 oncoprotein expression (MeT-5A), p53 small interfering RNA, or spontaneous p53 mutation (REN). These studies suggest that exposure to DNA-damaging agents can induce senescence in both murine and human mesothelioma cell lines and suggest a major, although not exclusive, role for p53 in this response. SV40 virus may contribute to mesothelioma progression by impairing stress-induced senescence, in part through p53 inactivation, thereby favoring survival and proliferation of mesothelial cells that have sustained DNA damage.