Lewis Lung Carcinoma Tumor-bearing Research Articles

Antibody-directed myostatin inhibition enhances muscle mass and function in tumor-bearing mice

Cancer cachexia describes the progressive skeletal muscle wasting and weakness in many cancer patients and accounts for >20% of cancer-related deaths. We tested the hypothesis that antibody-directed myostatin inhibition would attenuate the atrophy and loss of function in muscles of tumor-bearing mice. Twelve-week-old C57BL/6 mice received a subcutaneous injection of saline (control) or Lewis lung carcinoma (LLC) tumor cells. One week later, mice received either once weekly injections of saline (control, n = 12; LLC, n = 9) or a mouse chimera of anti-human myostatin antibody (PF-354, 10 mg·kg⁻¹·wk⁻¹, LLC+PF-354, n = 11) for 5 wk. Injection of LLC cells reduced muscle mass and maximum force of tibialis anterior (TA) muscles by 8-10% (P < 0.05), but the muscle atrophy and weakness were prevented with PF-354 treatment (P > 0.05). Maximum specific (normalized) force of diaphragm muscle strips was reduced with LLC injection (P < 0.05) but was not improved with PF-354 treatment (P > 0.05). PF-354 enhanced activity of oxidative enzymes in TA and diaphragm muscles of tumor-bearing mice by 118% and 89%, respectively (P < 0.05). Compared with controls, apoptosis that was not of myofibrillar or satellite cell origin was 140% higher in TA muscle cross sections from saline-treated LLC tumor-bearing mice (P < 0.05) but was not different in PF-354-treated tumor-bearing mice (P > 0.05). Antibody-directed myostatin inhibition attenuated the skeletal muscle atrophy and loss of muscle force-producing capacity in a murine model of cancer cachexia, in part by reducing apoptosis. The improvements in limb muscle mass and function highlight the therapeutic potential of antibody-directed myostatin inhibition for cancer cachexia.

Abstract C78: The mu opioid receptor regulates Lewis lung carcinoma tumor growth and metastasis

Abstract Introduction: Expression of the mu opioid receptor (MOR) is increased in patients with non-small cell lung cancer (NSCLC), a disease with poor prognosis and limited therapies. Previous work by our laboratory and others suggested a role of opioids in several aspects of tumor growth (Anticancer Res 2009;29:3195–3205, Mol Cancer Ther 2008;7:1669–1679, Cancer Res 2002;62:4491–4498). In this study, we investigated MOR as a potential therapeutic target for lung carcinoma. Methods: Lewis lung carcinoma (LLC) cells were treated with MOR siRNA or the peripheral MOR antagonist, methylnaltrexone (MNTX, 250 nM), prior to the addition of EGF (10 ng/ml), IGF (10 ng/ml), DAMGO (1 nM), morphine (1 nM) or serum (1, 5 or 10%). In vitro functional (cell proliferation and invasion) and biochemical studies (immunoblotting) were then conducted. For in vivo assays, C57BL/6J mice were injected intravenously with dual color (GFP-RFP labeled) LLC cells, with or without MOR siRNA treatment. In another experiment, MOR knockout as well as wildtype control mice were injected with LLC cells (subcutaneously into the flank) and followed for 12 weeks. In a parallel experiment, LLC tumor bearing C57BL/6J mice received continuous infusion of MNTX (10 mg/kg/day) for 2 weeks. Tumor growth and lung metastasis were evaluated using tumor volume measurements and/or in vivo fluorescent microscopy (Olympus OV-100) with or without intravenous injection of ProSense and/or MMP Sense probes. Results: Our data indicate that LLC cells have ∼5 fold increased expression of MOR (compared to primary lung epithelial or BEAS-2B cells). Inhibition of MOR with siRNA or MNTX treatment reduced in vitro LLC proliferation (90%) and invasion (50–75%). In vivo lung metastasis was reduced by ∼75% in wildtype C57BL/6J mice with LLC lacking MOR (siRNA) versus control siRNA-treated LLC (3 weeks post-i.v. injection, quantitated with OV-100). In addition, primary LLC tumor volume was substantially reduced (&amp;gt;90%) in MOR knockout mice compared to wildtype mice (flank injection). Finally, we observed that continuous infusion of MNTX (10 mg/kg/day) in mice with pre-existing LLC primary tumors attenuates further tumor growth. Conclusion: We have shown that MOR is a potential therapeutic target for inhibition of LLC proliferation, invasion and metastasis. In addition, MNTX can attenuate LLC primary tumor growth. Further study of MNTX as a potential therapeutic agent is warranted. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C78.

Use of a Novel Arg-Gly-Asp Radioligand,18F-AH111585, to Determine Changes in Tumor Vascularity After Antitumor Therapy

Despite the recent development of various radiolabeled Arg-Gly-Asp (RGD) peptides for imaging the alphavbeta3 integrin receptor, relatively little attention has been focused on the ability of these radiotracers to monitor changes in tumor vascularity after antitumor therapies. This study describes the favorable in vivo kinetics and tumor-targeting properties of 18F-AH111585, a novel 18F-RGD peptide, and its ability to monitor tumor vascularity noninvasively. Mice bearing Lewis lung carcinoma (LLC) tumors or Calu-6 non-small cell lung tumor xenografts were used for in vivo biodistribution and small-animal PET imaging studies. In addition, some animals were treated with either low-dose paclitaxel or the vascular endothelial growth factor receptor-2 tyrosine kinase inhibitor ZD4190. Tumor uptake of 18F-AH111585 and microvessel density were then assessed. Biodistribution of 18F-AH111585 demonstrated rapid clearance from the blood and key background organs and good tumor accumulation, with 1.5 percentage injected dose per gram (%ID/g) present at 2 h after injection in LLC tumors. Small-animal PET imaging of Calu-6 tumors allowed visualization of tumors above background tissue, with mean baseline uptake of 2.2 %ID/g. Paclitaxel therapy reduced the microvessel density in LLC tumor-bearing mice and resulted in significantly reduced 18F-AH111585 tumor uptake (P<0.05). ZD4190 therapy resulted in a significant (31.8%) decrease in 18F-AH111585 uptake in Calu-6 tumors, compared with the vehicle control-treated Calu-6 tumors, which had a 26.9% increase in 18F-AH111585 uptake over the same period (P<0.01). 18F-AH111585 is a promising 18F-labeled RGD tracer that offers a new approach to noninvasively image tumor vasculature. This tracer may reveal important information in the assessment of the impact of antitumor therapies, in particular those that predominantly target tumor blood vessels.

Enhanced Tumor Visualization by γ-Scintigraphy with111In-Labeled Polychelating-Polymer-Containing Immunoliposomes

Here, we have prepared long-circulating PEGylated liposomes heavily loaded with 111In via the liposome-incorporated polylysine-based (PLL-based) polychelating amphiphilic polymer (PAP) and additionally modified with the monoclonal antibody 2C5 (mAb 2C5) possessing the nucleosome-restricted (NS-restricted) specificity and capable of specific recognition of a broad variety of live cancer cells via the cancer cell surface bound NSs. These liposomes have been tested as a tumor-specific contrast agent for the gamma-scintigraphic visualization of model tumors in mice. The tumor accumulation of mAb 2C5 modified liposomes prepared in this study was significantly (3-to-5-fold) higher than in the neighboring muscle tissue at all times after administration (6, 24, and 48 h) in mice bearing murine Lewis lung carcinoma (LLC) and human HT-29 tumors. The whole body direct gamma-imaging of LLC tumor bearing mice at different times has demonstrated the superior in vivo tumor accumulation of the targeted mAb 2C5 modified PAP-containing PEGylated liposomes compared to nontargeted liposomal control formulations, which resulted in better and faster tumor imaging as shown with LLC-bearing mice.

Decreased T-Cell Proliferation and Skewed Immune Responses in LLC-Bearing Mice

Deficits in immune cell responses have been reported in cancer patients. We used the murine Lewis lung carcinoma (LLC) model to better understand these deficits. The goal of this study was to determine if the immune responses of LLC tumor-bearing (TB) mice differ from control mice and whether the difference could be attributed to either antigen-presenting cells (FPC) or to T cells. Tumors were first allowed to grow in vivo for approximately 2 weeks. Splenocytes were then isolated for in vitro proliferation and cytokine release studies. The results showed a decrease in mitogen-stimulated proliferation by unfractionated splenocyte cultures from TB mice when compared to control mice in response to concanavalin A (Con A), a T-cell mitogen. Decreased responses were also observed when the APC spleen cell fraction from TB mice was cultured with normal T cells, although proliferation was more prominently reduced in cultures of TB T cells plus normal APC. Also, splenocytes from TB mice secreted significantly increased levels of IFN-gamma, IL-4, and IL-10. Admixing APC from control mice with TB T cells significantly decreased levels of IL-4 and IL-10 secretion as compared to the levels secreted by cocultures of TB T cells and TB APC. The decreased cytokine profile in the presence of normal APC despite the presence of TB T cells suggests that APC contributes to the immune dysfunction, including Th skewing of tumor bearers, possibly through their influence on T-cell expansion and cytokine production. Finally, our assessment of the APC population contributing to the observed immune dysfunction--i.e., dendritic cells or macrophages--showed that the proliferation of TB T cells was decreased regardless of the APC population with which they were cocultured. However, normal T-cell proliferation was only reduced by the addition of TB macrophages and not by the addition of TB dendritic cells. In conclusion, our results demonstrate that LLC TB mice have a skewed immune response characterized by a decreased proliferative response with both T cells and APC affected by the presence of tumor.

Anti-Tumor Efficacy of Human Angiostatin Using Liver-Mediated Adeno-Associated Virus Gene Therapy

Angiostatin is a potent endogenous inhibitor of angiogenesis and tumor growth in vivo. The therapeutic potential of adeno-associated viral (AAV) gene delivery of angiostatin in modulating tumor growth in vivo was evaluated. Sustained levels of angiostatin were detected in the sera of mice for up to 6 months after they received a single injection of AAV-angiostatin. AAV-mediated stable expression of angiostatin inhibited tumor burden in the highly aggressive B16F10 melanoma and Lewis lung carcinoma (LLC) models of experimental metastasis. Moreover, AAV-angiostatin prolonged survival in B16F10 and LLC tumor-bearing mice compared to control groups. Anti-tumor efficacy was consistently observed when angiostatin serum levels of 15–50 ng/ml were detected following gene transfer, but the effect was minimal when the levels were lower or higher than this range. The combination of AAV-angiostatin gene therapy with chemotherapy was also shown to extend marginally the survival of mice bearing preestablished human tumors; however, the effect was evident only within a narrow dose of circulating angiostatin. These studies demonstrate the feasibility of using AAV antiangiogenic gene therapy as a cancer treatment modality and suggest that the optimal anti-tumor efficacy of angiostatin following gene transfer may be limited to a narrow dose range.

Alcohol use, conception time, and birth weight.

Predictors of birth weight and birth length were studied using sociodemographic data collected from 2259 women who resided in Funen County, Denmark, and delivered a healthy child during the period 1978-9 at Odense University Hospital. Low birth weight was significantly related to tobacco use in the year of delivery (p less than 0.01), alcohol use during the same period (p less than 0.05), and a delay in conception of over six months (p less than 0.01). Smoking history and a delay in conception were also significantly associated with short birth length (p less than 0.01 and p less than 0.01, respectively).