No abstract

Mammary glands develop postnatally in response to the hypothalamic-pituitary-gonadal axis. Obesityinduced changes in the local environment, however, retard mammary gland development during late pregnancy and lactation. To clarify the effects of obesity on fundamental duct development, we compared the mammary glands of nulliparous nonpregnant obese mice fed a high-fat diet with those of lean mice fed a normal diet. Obese mice had enlarged mammary glands, reflecting fat pad size, whereas the ducts in obese mice showed a less dense distribution with less frequent branching. Additionally, the ducts were surrounded by thick collagen layers, and were incompletely lined with myoepithelium. Because leptin receptors were localized in the epithelium region and leptin that was highly expressed in the obese glands suppressed mammary epithelial cell proliferation in vitro, the present results suggest that obesity disrupts mammary ductal development, possibly by remodeling the mammary microenvironment and promoting the expression of such paracrine factors as leptin.

Two major apoptosis pathways, the mitochondrial and death receptor pathways, are well recognized. Here we established cell lines from the fetal thymus of Apaf-1-, Caspase-9-, or Bax/Bak-deficient mice. These cell lines were resistant to apoptosis induced by DNA-damaging agents, RNA or protein synthesis inhibitors, or stress in the endoplasmic reticulum. However, they underwent efficient apoptosis when treated with kinase inhibitors such as staurosporine and H-89, indicating that these inhibitors induce a caspase-dependent apoptosis that is different from the mitochondrial pathway. CrmA, a Caspase-8 inhibitor, did not prevent staurosporine-induced apoptosis of fetal thymic cell lines, suggesting that the death receptor pathway was also not involved in this process. The staurosporine-induced cell death was inhibited by okadaic acid, a serine/threonine phosphatase inhibitor, suggesting that dephosphorylation of a proapoptotic molecule triggered the death process, or that phosphorylation of an antiapoptotic molecule could block the process. Cells of various types (fetal thymocytes, bone marrows, thymocytes, and splenocytes), but not embryonic fibroblasts, were sensitive to the noncanonical staurosporine-induced apoptosis, suggesting that the noncanonical apoptosis pathway is tissue specific.

Both adiponectin and secreted protein, acidic and rich in cysteine (SPARC) inhibit platelet-derived growth factor-BB (PDGF-BB)-induced and basic fibroblast growth factor (FGF2)-induced angiogenic activities through direct and indirect interactions. Although SPARC enhances nerve growth factor (NGF)-dependent neurogenesis, the physical interaction of NGFβ with adiponectin and SPARC remains obscure. Therefore, we first examined their intermolecular interaction by surface plasmon resonance method. NGFβ bound to immobilized SPARC with the binding constant of 59.4 nM, comparable with that of PDGF-BB (24.5 nM) but far less than that of FGF2 (14.4 µM). NGFβ bound to immobilized full length adiponectin with the binding constant of 103 nM, slightly higher than those of PDGF-BB (24.3 nM) and FGF2 (80.2 nM), respectively. Treatment of PC12 cells with SPARC did not cause mitogen-activated protein kinase (MAPK) activation and neurite outgrowth. However, simultaneous addition of SPARC with NGFβ enhanced NGFβ-induced MAPK phosphorylation and neurite outgrowth. Treatment of the cells with adiponectin increased AMP-activated protein kinase (AMPK) phosphorylation but failed to induce neurite outgrowth. Simultaneous treatment with NGFβ and adiponectin significantly reduced cell size and the number of cells with neurite, even after silencing the adiponectin receptors by their siRNA. These results indicate that NGFβ directly interacts with adiponectin and SPARC, whereas these interactions oppositely regulate NGFβ functions.

Background: Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have excellent effects on non-small cell lung cancer (NSCLC) harboring EGFR mutations. However, EGFR mutant NSCLC inevitably acquires resistance to EGFR-TKIs such as gefitinib, erlotinib, or afatinib. Previously, we found that bevacizumab, an anti-vascular endothelial growth factor-A (VEGF-A) antibody, enhanced the antitumor effects of gefitinib and afatinib in preclinical lung cancer models (Ichihara et al., Caner Res 2009; Ninomiya et al., Mol Cancer Ther 2013). In a phase II trial of OLCSG1001, we demonstrated relatively longer progression-free survival by adding bevacizumab to gefitinib in the treatment of patients harboring an EGFR exon 19 deletion. Seto et al. (Lancet Oncol 2015) also reported that adding bevacizumab to erlotinib in patients harboring EGFR mutations significantly prolonged progression-free survival. A new anti-angiogenic agent, the anti-human VEGF receptor-2 (VEGFR-2) antibody ramucirumab, has also been used clinically (Garon et al., Lancet 2014). We investigated the effect of an anti-VEGFR-2 antibody combined with erlotinib on the growth of lung cancers harboring EGFR mutations. Materials and Methods: NSCLC cell lines PC-9, which harbors EGFR exon 19 deletion and H3255, which harbors EGFR L858R mutation, were used in this study. Ramucirumab and the anti-murine VEGFR-2 antibody DC101 were kindly provided by Eli Lilly (Indianapolis, IN, USA). BALB/c nu/nu or transgenic mice expressing the delE748-752 mutant version of mouse Egfr driven by the SP-C promoter, which is equivalent to the delE746-A750 mutation of humans (C57BL/6/Egfr15DEL) (Ohashi et al. Cancer Sci 2009), were used for in vivo experiments. For xenograft models, PC-9 or H3255 cells were injected subcutaneously into BALB/c mice. The mice were divided into four treatment groups: vehicle, erlotinib, DC101, or erlotinib combined with DC101. Results: DC101 alone moderately inhibited the growth of PC-9 or H3255 tumor in the xenograft mouse models or suppressed lung cancers in C57BL/6/Egfr15DEL mice. Combined erlotinib and DC101 therapy inhibited the growth of PC-9 tumor in the xenograft mouse models more markedly than did erlotinib or DC101 alone. There were no differences in toxicity between monotherapy and combination therapy. Conclusion: Combination therapy with erlotinib and an anti-VEGFR-2 antibody had a stronger antitumor effect than those of the respective monotherapies in lung cancers harboring EGFR mutations in vivo. Citation Format: Hiroe Kayatani, Kadoaki Ohashi, Takeshi Imao, Kenichiro Kudo, Yuka Kato, Takashi Ninomiya, Toshio Kubo, Akiko Sato, Ktsuyuki Hotta, Mitsune Tanimoto, Katsuyuki Kiura. Combination effect of anti-VEGFR-2 antibody with erlotinib on EGFR mutant non-small cell lung cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5198.