Summary Lung cancer is the leading cause of cancer death worldwide1. Recent data suggest that tumor-associated inflammatory cells may modify lung tumor growth and invasiveness2-3. To determine the role of neutrophil elastase (NE or Elane) on tumor progression, we utilized the LSL-K-ras model of murine lung adenocarcinoma4 to generate LSL-K-ras/Elane−/− mice. Tumor burden was markedly reduced in LSL-K-ras/Elane−/− mice at all time points following induction of mutant K-ras expression. Kaplan-Meier life survival analysis demonstrated that while 100% of LSL-K-ras/Elane+/+ mice died, none of the mice lacking NE died. NE directly induced tumor cell proliferation in both human and mouse lung adenocarcinomas by gaining access to an endosomal compartment within tumor cells where it degraded insulin receptor substrate-1 (IRS1). Co-immunoprecipitation studies showed that as NE degraded IRS1, there was increased interaction between PI3K and the potent mitogen platelet derived growth factor receptor (PDGFR) thereby skewing the PI3K axis toward tumor cell proliferation. The inverse relationship identified between NE and IRS1 in LSL-K-ras mice was also identified in human lung adenocarcinomas, thus translating these findings to human disease. This study identifies IRS1 as a key regulator of PI3K within malignant cells. Additionally, this is the first description of a secreted proteinase gaining access to a cell beyond its plasma membrane and altering intracellular signaling.

Studies have begun to emerge showing critical roles for neutrophils in tumorigenesis. Neutrophils can have a significant impact on the tumor microenvironment via their production of cytokines and chemokines, which influence inflammatory cell recruitment and activation. Additionally, products secreted from neutrophils, such as reactive oxygen species and proteinases, have defined and specific roles in regulating tumor cell proliferation, angiogenesis, and metastasis. Although evidence suggests that neutrophils act in a decidedly protumor capacity in vivo, recent studies indicate that neutrophils may be manipulated to exhibit cytotoxicity against tumors. Herein, we explore the idea of targeting tumor-associated neutrophils as a means of antitumor therapy and the important ramifications such manipulation could pose to host tissues. Cancer Res; 71(7); 2411-6. Ó2011 AACR.

Neutrophils are released from the bone marrow in a regulated fashion to maintain homeostatic levels in the blood and to respond to physiological stresses, including infection. We show that under basal conditions granulocyte colony-stimulating factor (G-CSF) is an essential regulator of neutrophil release from the bone marrow. Nonredundant signals generated by the membrane-proximal 87 amino acids of the G-CSF receptor (G-CSFR) are sufficient to mediate this response. Surprisingly, G-CSFR expression on neutrophils is neither necessary nor sufficient for their mobilization from the bone marrow, suggesting that G-CSF induces neutrophil mobilization indirectly through the generation of trans-acting signals. Evidence is provided suggesting that downregulation of stromal cell-derived factor 1 expression in the bone marrow may represent such a signal.

IPF is a progressive lung disorder characterized by fibroblast proliferation and myofibroblast differentiation. Although neutrophil accumulation within IPF lungs has been negatively correlated with outcomes, the role played by neutrophils in lung fibrosis remains poorly understood. We have demonstrated previously that NE promotes lung cancer cell proliferation and hypothesized that it may have a similar effect on fibroblasts. In the current study, we show that NE(-/-) mice are protected from asbestos-induced lung fibrosis. NE(-/-) mice displayed reduced fibroblast and myofibroblast content when compared with controls. NE directly both lung fibroblast proliferation and myofibroblast differentiation in vitro, as evidenced by proliferation assays, collagen gel contractility assays, and αSMA induction. Furthermore, αSMA induction occurs in a TGF-β-independent fashion. Treatment of asbestos-recipient mice with ONO-5046, a synthetic NE antagonist, reduced hydroxyproline content. Thus, the current study points to a key role for neutrophils and NE in the progression of lung fibrosis. Lastly, the study lends rationale to use of NE-inhibitory approaches as a novel therapeutic strategy for patients with lung fibrosis.

There is accumulating evidence that delivery of bone marrow cells to sites of ischemia by direct local injection or mobilization into the blood can stimulate angiogenesis. This has stimulated tremendous interest in the translational potential of angiogenic cell population(s) in the bone marrow to mediate therapeutic angiogenesis. However, the mechanisms by which these cells stimulate angiogenesis are unclear. Herein, we show that the inflammatory subset of monocytes is selectively mobilized into blood after surgical induction of hindlimb ischemia in mice and is selectively recruited to ischemic muscle. Adoptive-transfer studies show that delivery of a small number of inflammatory monocytes early (within 48 h) of induction of ischemia results in a marked increase in the local production of MCP-1, which in turn, is associated with a secondary, more robust wave of monocyte recruitment. Studies of mice genetically deficient in MCP-1 or CCR2 indicate that although not required for the early recruitment of monocytes, the secondary wave of monocyte recruitment and subsequent stimulation of angiogenesis are dependent on CCR2 signaling. Collectively, these data suggest a novel role for MCP-1 in the inflammatory, angiogenic response to ischemia.