Markers Of Tumor Invasion Research Articles

Expression of Notch 1 receptor associated with tumor aggressiveness in papillary thyroid carcinoma.

AimThe aim of this study was to assess if the expression of Notch 1 receptor is associated with tumor aggressiveness in papillary thyroid carcinomas (PTCs).Patients and methodsBy searching the electronic medical record system of Xin Hua Hospital, all cases of PTC patients who underwent thyroidectomy in the hospital between 2013 and 2014 were retrieved. Then, the cases of patients who had a history of any other malignancy or whose thyroid tumor specimen was not available for assay were rejected. Finally, 68 cases of PTC patients were obtained. Formalin-fixed paraffin-embedded tissue blocks of these patients were studied by immunohistochemistry to learn the expression of Notch 1 receptor. Meanwhile, the clinical data of these patients including sex, age, size of the tumor, presence of node metastasis or distant metastasis, and presence of capsule invasion and tumor multicentricity were collected. Pearson’s chi-square test or Fisher’s exact test was used for measuring statistical differences in categorical variables. All the statistical tests were two-sided. A P-value <0.05 was considered to be statistically significant.ResultsA total of 19 male and 49 female PTC patients with a mean age of 44.8±13.6 years (range 18–78 years) were studied. Notch 1 receptor expression was found in 15/68 (22%) samples of PTC. The expression of Notch 1 receptor was significantly associated with tumor size (P=0.021), distant metastasis (P=0.008), capsule invasion (P=0.001), tumor multicentricity (P=0.018), and age (P=0.033). However, the expression of Notch 1 receptor was not significantly correlated with node metastasis (P=0.096) and sex (P=0.901).ConclusionThe expression of Notch 1 receptor is associated with tumor aggressiveness in PTC, and may be used as a molecular marker of tumor invasiveness in PTC. PTC patients who show positive expression of Notch 1 receptor may benefit from radioiodine remnant ablation.

Chemopreventive and hepatoprotective roles of adiponectin (SULF2 inhibitor) in hepatocelluar carcinoma.

Sulfatase 2 (SULF2) is an extracellular enzyme that catalyzes the removal of 6-O-sulfate groups from the heparan sulfate (HS). As elevated SULF2 activity has been correlated with hepatocellular carcinoma (HCC), this study was conducted to evaluate the chemoprotective and the hepatoprotective roles of adiponectin, as a SULF2 inhibitor, against hepatocellular carcinoma both in vivo and in vitro. HCC was induced in rats using thioacetamide (200 mg/kg). Treated rats received adiponectin (5 μg/kg) once a week. Moreover, human hepatocellular carcinoma (HepG2) cell line was used as an in-vitro model. In both in-vivo and in-vitro models, adiponectin completely blocked HCC-induced SULF2 elevation. The antitumor activity of adiponectin was confirmed by 80% increased the survival rate, 73% reduction in the average number of nodules per nodule-bearing liver and 46% reduction in serum AFP. In addition, adiponectin ameliorated HCC-induced expression of tumor invasion markers, MMP9, syndecan-1 and FGF-2. Moreover, adiponectin attenuated HCC-induced elevation of nfκb and TNF-α levels. Moreover, treatment of HepG2 cell line with adiponectin showed dose-dependent reduction of HepG2 cell viability and elevation of cellular cytotoxicity. Besides, Adiponectin yielded the same results in HepG2 cells in a dose-dependent manner. Adiponectin achieved both hepatoprotective and chemoprotective effects against HCC through blocking of SULF2.

Expression of Angiogenic Factors in Invasive Retinoblastoma Tumors Is Associated With Increase in Tumor Cells Expressing Stem Cell Marker Sox2.

Progression of retinoblastoma is associated with increased tumor angiogenesis. However, a clear relationship between the expression of angiogenic markers in specific regions of the tumor and tumor progression has not been established. This study investigates the association between angiogenic factors in retinoblastomas with choroidal and/or optic nerve invasion (high-risk/invasive retinoblastoma) and expression of Sox2, a stem cell marker. To investigate the association between the expression of angiogenic factors and markers of tumor invasiveness, such as the stem cell marker Sox2, in retinoblastoma tissues. Immunohistochemistry was used to evaluate coexpression of the angiogenic growth factors vascular endothelial growth factor A (VEGF-A), VEGF receptor 2 (VEGFR-2), and endoglin (CD105); markers of glial differentiation (vimentin and glial fibrillary acidic protein); and a neural stem cell marker (Sox2). Expression was assessed in nonneoplastic and neoplastic ocular tissues collected from enucleated eyes of patients with retinoblastoma. During qualitative data interpretation, evaluating pathologists were masked to patient grouping. Expression of VEGF-A and VEGFR-2 in noninvasive (non-high-risk feature) retinoblastoma tumors was lower than in the invasive, or high-risk feature tumors. Moreover, our data indicate that the tumor cells, and not the surrounding stroma, secrete VEGF-A and that angiogenesis is mostly localized to the iris. Finally, our data showed that the expression of the neural stem cell marker Sox2 is associated with eyes with increased VEGF-A expression and tumor invasiveness. Increased expression of angiogenic factors, with a concomitant increase in expression of the stem cell marker Sox2 observed in retinoblastoma tissues, may partially explain the aggressiveness of these tumors. The complex interaction of angiogenic and stem cell-related pathways in these tumors, especially in high-risk feature retinoblastoma, suggests that targeting tumor cells capable of secreting vasculogenic factors, as well as proangiogenic genes and signaling pathways, may be necessary for development of effective antimetastatic retinoblastoma drugs.

Phase II trial of everolimus and erlotinib in patients with platinum-resistant recurrent and/or metastatic head and neck squamous cell carcinoma

Enhanced phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway is one of the key adaptive changes accounting for epidermal growth factor receptor (EGFR) inhibitor-resistant growth in head and neck squamous cell carcinoma (HNSCC). We designed a phase II clinical trial of EGFR tyrosine kinase inhibitor (TKI), erlotinib, in association with the mTOR inhibitor, everolimus, based on the hypothesis that the downstream effects of Akt through inhibition of mTOR may enhance the effectiveness of the EGFR-TKI in patients with recurrent/metastatic HNSCC. Patients with histologically or cytologically confirmed platinum-resistant HNSCC received everolimus 5 mg and erlotinib 150 mg daily orally until disease progression, intolerable toxicity, investigator or patient decision. Cytokines and angiogenic factors profile, limited mutation analysis and p16 immunohistochemistry status were included in the biomarker analysis. Of the 35 assessable patients, 3 (8%) achieved partial response at 4 weeks, 1 confirmed at 12 weeks; overall response rate at 12 weeks was 2.8%. Twenty-seven (77%) patients achieved disease stabilization at 4 weeks, 11 (31%) confirmed at 12 weeks. Twelve-week progression-free survival (PFS) was 49%, median PFS 11.9 weeks and median overall survival (OS) 10.25 months. High neutrophil gelatinase lipocalin (P = 0.01) and vascular endothelial growth factor (VEGF) (P = 0.04) plasma levels were significantly associated with worse OS. The combination of erlotinib and everolimus did not show significant benefit in unselected patients with platinum-resistant metastatic HNSCC despite a manageable toxicity profile. Markers of tumor invasion and hypoxia identify a group of patients with particularly poor prognosis. NCT00942734.

Gene and protein expression of glucose transporter 1 and glucose transporter 3 in human laryngeal cancer-the relationship with regulatory hypoxia-inducible factor-1α expression, tumor invasiveness, and patient prognosis.

Increased glucose uptake mediated by glucose transporters and reliance on glycolysis are common features of malignant cells. Hypoxia-inducible factor-1α supports the adaptation of hypoxic cells by inducing genes related to glucose metabolism. The contribution of glucose transporter (GLUT) and hypoxia-inducible factor-1α (HIF-1α) activity to tumor behavior and their prognostic value in head and neck cancers remains unclear. The aim of this study was to examine the predictive value of GLUT1, GLUT3, and HIF-1α messenger RNA (mRNA)/protein expression as markers of tumor aggressiveness and prognosis in laryngeal cancer. The level of hypoxia/metabolic marker genes was determined in 106 squamous cell laryngeal cancer (SCC) and 73 noncancerous matched mucosa (NCM) controls using quantitative real-time PCR. The related protein levels were analyzed by Western blot. Positive expression of SLC2A1, SLC2A3, and HIF-1α genes was noted in 83.9, 82.1, and 71.7 % of SCC specimens and in 34.4, 59.4, and 62.5 % of laryngeal cancer samples. Higher levels of mRNA/protein for GLUT1 and HIF-1α were noted in SCC compared to NCM (p < 0.05). SLC2A1 was found to have a positive relationship with grade, tumor front grading (TFG) score, and depth and mode of invasion (p < 0.05). SLC2A3 was related to grade and invasion type (p < 0.05). There were also relationships of HIF-1α with pTNM, TFG scale, invasion depth and mode, tumor recurrences, and overall survival (p < 0.05). In addition, more advanced tumors were found to be more likely to demonstrate positive expression of these proteins. In conclusion, the hypoxia/metabolic markers studied could be used as molecular markers of tumor invasiveness in laryngeal cancer.

O1.06 * NOVEL MARKERS OF MENINGIOMA AGGRESSIVENESS - A STUDY OF MENINGIOMA VERSUS PERITUMORAL NERVOUS TISSUE

Meningiomas constitute approximately one-third of primary CNS tumors and classified by WHO in meningioma (grade I), with many histologic variants, atypical meningioma (grade II) and anaplastic meningioma (grade III). Most of them have a long survival, 1-3% are anaplastic with poor outcome and high recurrence rate and 15-20% of meningiomas are atypical with an intermediate and variable behavior. At present, only histologic grading is a reliable prognostic factor and we lack markers to identify among atypical meningiomas the most aggressive ones. For the first time in meningiomas, we propose to evaluate Lycopersicon esculentum lectin (LEA, specific for N-acetyl-D-glucosamine and N-acetyl-polylactosamine oligomers) and Aquaporin-4 (AQP4) as new factors involved in tumor invasion. LEA was chosen because it was localized in endothelial cell glycocalix in blood brain barrier (BBB) and it has been demonstrated that altered glycosilation of neoplastic cells induce vascular infiltration of cancer cells through endothelium. AQP4 is the most abundant water channel localized in the glial endfoots of the CNS and has been correlated with peritumoral oedema, that in meningioma can result as a marker of tumor invasion. The present study was performed in 20 grade I, 10 atypical and 5 anaplastic meningiomas in which a part of nervous tissue adjacent the meningioma was present on the surgical specimen. We observed a huge increase of AQP4 expression in the peritumoral nervous tissue in meningioma compared to normal nervous tissue. A thin staining in the leptomeninge and the muscolaris mucosae of the biggest vessels in tumor periphery was noticed, while tumor cells were always negative. Also in atypical meningioma AQP4 clearly define tumor boundaries. So, a minimum infiltration of nervous tissue, poorly identifiable with HE staining, could be simply visible using AQP4 staining. As for the anaplastic meningioma, which shows infiltration of nervous tissue, AQP4 has been localized also in astrocytes trapped into the tumor. In this aggressive meningioma, we found a strong LEA staining in tumor cells membrane, while grade I meningioma showed a less intense positivity. LEA expression was localized in meningothelial tumor cells and tumor vessels, as in the nervous tissue only in the endothelial cells. These observations show a role of AQP4 in peritumoral oedema and point to AQP4 as a useful marker to identify nervous tissue infiltration extension and thus meningioma aggressiveness. Moreover, LEA staining localized both in meningioma cell membranes and in the endothelial cells of the tumor vessels indicates that LEA-affinity oligomers could be implicated in adhesion and invasiveness of tumor cells. Both the glycosidic residues stained by LEA and AQP4 could be potential therapeutic targets.

Laminin‐5 gamma 2 chain expression is associated with intensity of tumor budding and density of stromal myofibroblasts in oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is one of the most prevalent cancers worldwide. Laminin-5 gamma 2 chain (laminin-5 γ2) is a protein associated to a migratory phenotype in epithelial neoplastic cells. Stromal myofibroblasts also play a significant role in tumor invasion, due to its ability to modify the extracellular matrix. Tumor budding is a morphologic marker of tumor invasion. The aim of this study was to evaluate the expression of laminin-5 γ2 in OSCC and its association with intensity of tumor budding and density of stromal myofibroblasts. Paraffin-embedded archival samples of 57 OSCC patients were evaluated. Immunohistochemistry was employed to detect laminin-5 γ2, alpha smooth muscle actin (marker of stromal myofibroblasts), and multicytokeratin (to identify OSCC cells in tumor budding evaluation). Laminin-5 γ2 expression and its association with intensity of tumor budding and density of stromal myofibroblasts were analyzed. Association among intensity of tumor budding and density of stromal myofibroblasts was also evaluated. Higher laminin-5 γ2 expression was associated with high-intensity tumor budding (P < 0.05) and with higher density of stromal myofibroblasts (P < 0.05). Moreover, high-intensity tumor budding was associated with higher density of stromal myofibroblasts (P < 0.05). In OSCC, higher laminin-5 γ2 expression is associated with high-intensity tumor budding and with higher density of stromal myofibroblasts, suggesting that this expression is related to the establishment of an invasive phenotype of neoplastic cells and a permissive environment for tumor invasion in this neoplasia.

Significance of serum vascular endothelial growth factor in chronic liver disease and hepatocellular carcinoma: An exploratory study

Background and Objectives: Vascular endothelial growth factor (VEGF) is the most potent directly acting angiogenic growth factor that plays an important role in inducing tumor-associated angiogenesis. We evaluated the clinical significance of the circulating VEGF in patients with hepatocelluar carcinoma (HCC) and chronic liver disease (CLD). Patients and Methods: The study included 65 patients [20 with chronic viral hepatitis (CVH), 20 with liver cirrhosis (LC) and 25 with HCC] and 15 age- and gender- matched healthy subjects as controls. For each studied subject, detection of hepatitis viral markers, and assessment of liver function tests, α-fetoprotein (AFP) and VEGF were performed. Results: There was a significantly higher VEGF level in the sera of HCC patients as compared to other groups (p&lt; 0.001). Serum VEGF level was significantly associated with portal vein tumor thrombosis (p&lt; 0.01), but was not related to tumor size. There was no significant difference between the serum VEGF levels in either LC or CVH groups when compared to the controls. Moreover, no significant difference was detected between the different Child-Pugh classes among LC patients. Furthermore, no correlation was found between the level of serum VEGF and AFP, serum albumin, aminotransferases or prothrombin time (PT) in all the studied groups. Conclusions: Serum VEGF may be useful as a tumor marker for diagnosis of HCC and as a prognostic marker for tumor invasion. Large studies with greater numbers of patients and controls is required to support our conclusion.

Transforming growth factor β1 is not a reliable biomarker for valvular fibrosis but could be a potential serum marker for invasiveness of prolactinomas (pilot study)

Transforming growth factor β1 (TGFβ1) signaling pathway is crucial for both human fibrogenesis and tumorigenesis. This study aimed to investigate the usefulness of TGFβ1 and matrix metalloproteinase 2 (MMP2) as potential circulating markers for fibrotic valvular heart disease (FVHD) and invasiveness as well as of Fetuin A as a marker for calcification in patients with prolactinomas. The study population consisted of 147 subjects divided into four groups: 30 dopamine agonist (DA)-treated prolactinoma patients with proven FVHD and three control groups with normal echocardiograms: 43 DA-treated patients, 26 naïve patients, and 48 healthy subjects. We observed significantly higher serum TGFβ1 levels in all three patient groups than in the healthy subjects (21.4 ± 8.86 vs 19.1 ± 9.03 vs 20.7±11.5 vs 15.8 ± 7.2 ng/ml; P=0.032). Moreover, TGFβ1 levels were significantly higher in patients with macroprolactinomas and invasive prolactinomas than in those with microprolactinomas and noninvasive tumors respectively. In addition, a strong positive linear relationship between TGFβ1 levels and invasiveness score (ρ=0.924; P<0.001) and a moderate correlation between TGFβ1 levels and tumor volume (r=0.546; P<0.002) were observed in patients with invasive prolactinomas. By contrast, prolactin (PRL) levels exhibited a better correlation with tumor volume (r=0.721; P<0.001) than with invasiveness score (ρ=0.436; P<0.020). No significant difference was observed in Fetuin A levels between patients with FVHD and healthy controls. Results concerning MMP2 were unclear. TGFβ1, MMP2, and Fetuin A are not reliable biomarkers for valvular fibrosis and calcification in DA-treated patients with prolactinomas, but TGFβ1 may represent a useful serum marker for tumor invasiveness. The simultaneous determination of TGFβ1 and PRL levels could improve the noninvasive assessment of prolactinoma behavior.

Phase II trial of everolimus and erlotinib in patients with platinum-resistant recurrent and/or metastatic head and neck squamous cell carcinoma.

6067 Background: Head and neck squamous cell carcinoma (HNSCC) is characterized by epidermal growth factor receptor (EGFR) overexpression but treatments targeting EGFR have met with limited clinical success. We hypothesized that everolimus (EV), an mTOR inhibitor, potentiates the activity of erlotinib (ER), an EGFR tyrosine kinase inhibitor, in platinum-resistant recurrent and/or metastatic HNSCC. Methods: This single-arm two-stage phase II clinical trial evaluated EV 5 mg and ER 150 mg orally daily in patients (pts) with platinum-resistant recurrent and/or metastatic HNSCC. Primary endpoints were response rate and 12-week progression-free survival (PFS). Plasma biomarkers by multianalyte profiling (Luminex Corp) and ELISA at baseline (20 pts), 4 weeks (18 pts) and 12 weeks (16 pts) as well as p16 expression in baseline tumor tissue (29 pts) were assessed in correlation with clinical results. Results: Of the 36 evaluable pts (median age 61 years, 29 males, 19 with oropharynx primaries, 15 with ≥2 lines of chemotherapy, 17 with prior cetuximab), 3 (8%) achieved partial response at 4 weeks, one of which was confirmed at 12 weeks, while 27 (75%) pts achieved disease stabilization at 4 weeks, 11 of which were confirmed at 12 weeks. The 12-week PFS was 49%, median PFS was 11.9 weeks and median overall survival (OS) was 10.25 months. Grade 3 toxicities included mucositis (17%), fatigue (14%), skin (8%), diarrhea (8%), and 5 pts withdrew from trial secondary to toxicity. High neutrophil gelatinase-associated lipocalin(NGAL) levels at baseline were associated with worse OS (HR 4.59; 95%CI 1.36, 15.46; p=0.014) and at 4 weeks with worse PFS (HR=12.29; 95% CI 1.26, 119.87; p=0.03) and OS (HR=6.21; 95% CI 0.95, 40.62; p=0.057). High carbonic anhydrase-9 (CA-9) levels at 4 weeks were associated with worse OS (HR=1.18; 95% CI 1.001, 1.41; p=0.049), while decreased CA-9 levels at 4 weeks were associated with better PFS (p=0.77). Conclusions: Efficacy for the combination ofEV and ER is reasonable however plasma markers of tumor invasion and hypoxia are associated with worse outcomes, and further biomarker analysis may define subsets of pts with significant therapeutic benefit. Clinical trial information: NCT00942734.

Osteonectin Expression in Surrounding Stroma of Craniopharyngiomas

Craniopharyngioma is an epithelial tumor of the sellar region with a high survival rate but a high rate of recurrence, especially in children. Hypothalamic involvement, tumor recurrence, and multiple treatments result in clinical deterioration and impaired quality of life. Using immunohistochemistry, we investigated the expression pattern of osteonectin, a marker of tumor invasion and aggressive behavior, in 43 cases of craniopharyngioma. We observed a positive correlation of osteonectin expression in connective-type stromal tissue surrounding the epithelial tumor cells of craniopharyngioma with the extent of central nervous system infiltration and recurrence rate (P < .001). Given the previous success of chemotherapeutic agents that target the tumor microenvironment, our findings on osteonectin expression in stroma of craniopharyngiomas might, hopefully, be a guide to find newer prognostic markers capable of estimating the risk of progression or recurrence. They may also aid in the development of therapeutics that target tumor microenvironment to improve patient outcome.

Abstract 1411: Pathogenesis and treatment of rhabdomyosarcoma in a susceptible murine model.

Abstract Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma of childhood. While the establishment of standard multi-modal therapy has led to improved survival, new therapeutic approaches are desperately needed to improve outcomes in patients with metastatic and recurrent disease. Our objective was to develop a highly penetrant mouse model of RMS to study the molecular pathways of the disease and to evaluate novel anticancer agents for the treatment of RMS. The hedgehog (Hh) pathway has been demonstrated to be activated in embryonal RMS. Communication between Hh and PI3K/AKT/mTOR pathways provides a rationale for the study of mTOR inhibitors such as rapamycin in RMS therapy. We developed a novel RMS mouse model by breeding Ptch6-7 knockouts onto a tumor susceptible SKH1 genetic background. We employed these animals (n=20/group) to study the effects of mTOR inhibitor, rapamycin, on the growth and progression of RMS tumors. We found that 42% of the Ptch1+/−SKH1 mice receiving no treatment developed RMS. These tumors were confirmed to have embryonal RMS morphology. These animals spontaneously develop RMS with embryonal histology predominantly in the hind limbs. The tumors developed at approximately 20 weeks of age; thereafter they grew very prolifically. Molecular studies demonstrated high expression of both Myogenin and Myf5. The tumors also manifested Hh pathway activation. The RMS bearing animals were divided into two groups with a third group of Ptch+/+ mice acting as a wild-type control group. Group-I animals receiving no treatment served as positive controls. The increase in tumor size in these animals was compared with the tumor growth of animals treated with rapamycin. The treatment resulted in a significant decrease in the rate of tumor growth as compared to the control group (p&amp;lt;0.05). This treatment also reduced the expression of AKT and Hh pathways, which was consistent with the tumor inhibitory response seen on a gross level. Decreases in markers of tumor invasiveness and VEGF expression were also noted. We conclude that the Ptch1+/− Skh1 mouse represents a novel murine model for aggressive embryonal RMS that recapitulates the pathobiology of human disease. Rapamycin effectively inhibits the growth of Ptch1+/− Skh1 RMS tumors providing insight into the mechanism underlying RMS tumorigenesis. mTOR inhibition with possible Hh pathway cross talk inhibition represents a promising target in the treatment of RMS in this murine model. Citation Format: Samer Zaid-Kaylani, Joseph G. Pressey, David R. Kelly, Mohammad Athar. Pathogenesis and treatment of rhabdomyosarcoma in a susceptible murine model. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1411. doi:10.1158/1538-7445.AM2013-1411

The Clinical Significance of the CD163+ and CD68+ Macrophages in Patients with Hepatocellular Carcinoma

Our previous study has found that the abundance of peritumoral CD68+ macrophages was associated with poor prognosis in hepatocellular carcinoma (HCC) after resection. However, CD68 staining could not discriminate the protumoral or tumoricidal subpopulations from pan-macrophages. CD163 is a marker of alternatively activated macrophages. In this study, the clinical significance of CD163+ cells in tumors and peritumoral liver tissues was evaluated in a cohort of 295 patients with HCC after curative resection. We found that the density of CD163+ cells was well correlated with that of CD68+ cells in both tumors and peritumoral liver tissues but was much more. Immunostaining on consecutive sections and flow cytometry assay on surgical resected specimens further supported the findings that the CD163+ cells was more abundant than CD68+ cells. The density of peritumoral CD68+ cells was associated with poor recurrence-free survival (RFS) and poor overall survival (OS) (P = 0.004 and P = 0.001, respectively), whereas the CD163+ cells have no prognostic values either in tumors or in peritumoral liver tissues. In another cohort of 107 HCC patients, preoperative plasma concentration of soluble form of CD163 (sCD163) was associated with active hepatitis-related factors but not associated with the markers of tumor invasion. In conclusion, both the CD163+ cells local infiltration and plasma sCD163 were of limited significance in HCC, and they were more likely markers related to active hepatitis rather than tumor progression.

Efficacy of Aerosolized Celecoxib Encapsulated Nanostructured Lipid Carrier in Non-small Cell Lung Cancer in Combination with Docetaxel

Evaluation of in-vivo anticancer activity of aerosolized Celecoxib encapsulated Nanolipidcarriers (Cxb-NLC) as a single therapeutic agent and combined with intravenously administered Docetaxel (Doc) against non-small cell lung cancer. Cxb-NLC were prepared by high-pressure homogenization and were characterized for its physicochemical characteristics. Metastatic A549 tumor model in Nu/Nu mice was used to evaluate response of aerosolized Cxb-NLC & Doc. Isolated lung tumor samples were analyzed for: a) DNA fragmentation and cleaved caspase-3 by immunohistochemistry, b) apoptotic and angiogenic protein markers by western blot, c) global proteomic alterations by an isobaric labeling quantitative proteomic method and d) toxicity studies of NLC. The particle size of Cxb-NLC was 217 ± 20nm, while entrapment efficiency was more than 90%. Cxb-NLC and Doc alone and in combination showed 25 ± 4%, 37 ± 5%, and 67 ± 4% reduction in tumor size respectively compared to control. Proteomic analysis with combination treatment further revealed significantly decreased expression of multiple pro-survival and pro-metastasis proteins as well as tumor invasion markers and the expression of S100 family proteins, such as S100A6 and S100P were decreased by 2.5 and 1.6 fold. Combination therapy with Cxb-NLC and Doc showed significant reduction in tumor growth which was further confirmed by proteomic analysis.

Characteristic Immunophenotype of Solid Subtype Component in Lung Adenocarcinoma

Lung adenocarcinomas represent a morphologically heterogeneous tumor composed of an admixture of different histologic subtypes (lepidic, papillary, acinar, and solid subtype). The presence of a solid subtype component is reported to be associated with a poorer prognosis. The aim of this study was to evaluate the characteristic immunophenotype of the solid subtype component compared with the immunophenotypes of other components. We analyzed the clinicopathological characteristics of stage I adenocarcinoma patients with predominant solid subtype disease. Furthermore, we immunostained adenocarcinomas with predominant lepidic, papillary, acinar, and solid subtype components (n=23 each) for 10 molecular markers of tumor invasiveness and scored the results. Patients showing predominance of the solid subtype component (solid subtype adenocarcinoma) had a poorer prognosis than those showing predominance of the lepidic, papillary, or acinar component. Lymphovascular invasion was more often detected in solid subtype tumors than in others. The solid subtype component showed a significantly stronger staining intensity of laminin-5 expression than the lepidic, papillary, and acinar components (P<0.001, P<0.001, and P=0.016, respectively). The fibronectin and vimentin expression levels were also significantly higher in the solid subtype component than in other components. This immunostaining character was validated by using mixed-subtype adenocarcinomas containing all four components in the same tumor. This study concluded that the solid subtype component in lung adenocarcinomas exhibit the invasive immunophenotype, including increased laminin-5 expression, compared with the other components, which may be associated with a poorer prognosis.

Abstract 1462: Anti-metastasis effect of novel ALK5 inhibitor in both breast cancer cells in vitro and 4T1 xenograft mice in vivo

Abstract We have synthesized novel ALK5 inhibitors and investigated their effects of on breast cancer cells as well as tumor bearing MMTV/cNeu transgenic mice and 4T1 orthopathologically xenograft balbC mice. The treatment of ALK5 inhibitors into breast tumor bearing MMTV/cNeu transgenic mice and orthotopically injected mice inhibited the numbers of nodules in the lung compared to that of untreated animal. The level of ß-casein in the lung of the breast tumor bearing MMTV/cNeu transgenic mice and 4T1 orthotopically xenograft mice were decreased by ALK5 inhibitor treatment. Zymogragh showed that the levels of MMP2 and MMP9 were decreased in the lung and breast tumor of ALK5 inhibitors treated breast tumor bearing MMTV/cNeu transgenic mice and 4T1 orthotopically xenograft mice. The action mechanisms of new ALK5 inhibitors were investigated in both breast cancer cells and breast cancer mice model. TGFß signaling, phosphorylated smad2/3 based from the western blot analysis in vitro and in vivo as well as confocal microscopic analysis in vitro was decreased by ALK5 inhibitors. And also, new ALK5 inhibitors inhibited the TGFß action on wound healing and cell invasion based on wound healing assay and invasion assay. This data strongly argue that ALK5 inhibitors have anti-metastasis effects via inhibiting ALK5 activity in vivo and in vitro. In order to gain further insight into the mechanism of ALK5 inhibitor, we have carried out experiment to examine the effects of ALK5 inhibitor in human breast cancer MDA-MB-231 cell. TGFß stimulated mesenchymal markers of epithelial mesenchymal transition (EMT), such as Snail, TWIST, N-cadherin and these were inhibited by ALK5 inhibitor concomitant treatment. Also, TGFß increased tumor invasion markers such as MMP2, and MMP9 which were inhibited by ALK5 inhibitor concomitant treatment. In summary, ALK5 inhibitors have anti-metastatic effect on human breast cancer cells as well as tumor bearing MMTV/cNeu transgenic mice. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1462. doi:10.1158/1538-7445.AM2011-1462

Molecular markers of tumor invasiveness in ameloblastoma: An update

The aim of the present article was to review the current new knowledge on the molecular markers of tumor invasion in ameloblastoma. In this review, tumor molecular markers were identified and allocated to the following six groups according to their functions: (I) Markers involved in extracellular matrix degradation, (II) Molecular markers involved in cell adhesion lost, (III) Molecular markers involved in bone remodeling, (IV) Cytokines involved in angiogenesis, (V) Molecular markers related with the function of tumor stromal cells on the invasion of ameloblastoma, and (VI) Molecular markers involved in cell proliferation related with invasion.In general, the location of markers within the tumor and not their quantitative assessments as such is emphasized. Data showed that the correlation among molecular markers of invasive relevance is still not quite clear. Results on markers of tumor invasion and metastatic potential appeared to be too premature for a statement regarding the instinct invasive nature of ameloblastoma. The unraveling of specific new details concerning these mechanisms, whereby the expression and relationships among the molecules are mediated, may provide an opportunity to afford efficient prevention and develop new treatment therapies.

Preoperative Serum IL-6 Levels: Clinical Importance in Gastric Carcinoma with Lymph Node Metastasis

Aim: The interleukin-6 (IL-6) pathway is one of the mechanisms that link inflammation and angiogenesis. The purpose of this study was to investigate the preoperative serum levels of IL-6 in gastric carcinoma with lymph node metastasis, and to correlate them with disease status. Method: A total of 100 patients who underwent gastrectomy were enrolled in this study. Serum levels of IL-6 were assessed enzyme linked immunosorbent assay (ELISA). Histological findings included tumor size, depth of tumor invasion, lymph node (LN) metastasis, and TNM stage. Result:Increases in cancer invasion and staging are generally associated with increases in preoperative serum IL-6 levels. IL-6 levels were correlated with invasion depth (p<0.001), LN metastasis (p<0.001), and TNM stage. The presence of peritoneal seeding metastasis is associated with IL-6 levels (p<0.001). Conclusion: Preoperative serum IL-6 levels might be markers of tumor invasion, LN metastasis, and TNM stage. These results suggest that IL-6 levels are potential molecular markers that predict lymphatic involvement in gastric carcinoma.

The Significance of Fibroblast Growth Factor Receptor 2 Expression in Differentiation of Hepatocellular Carcinoma

Fibroblast growth factor receptors (FGFRs) have been reported to be involved in the progression of many cancers. The aim of this study is to clarify the significance of FGFR2 expression in the differentiation of hepatocellular carcinoma (HCC). One nodule-in-nodule HCC sample was obtained from a patient to analyze the different expression in well- to moderately differentiated HCC and poorly differentiated HCC using microarray technique. The expression of FGFR2 in 46 patients with surgically resected HCC was immunohistochemically examined and analyzed in relation to their clinicopathological factors. Fgfr2 was 4.7 times up-regulated in poorly differentiated HCC from a nodule-in-nodule sample. The high expression group was 16 cases and the low expression group was 30 cases. The high FGFR2 expression correlated significantly with a poor histological differentiation, a higher incidence of portal vein and a high level of alpha-fetoprotein. The overall survival rates and the disease-free survival rates in high expression were significantly worse than those in low. In conclusion, a high FGFR2 expression plays an important role in poor differentiation, portal vein invasion, high alpha-fetoprotein production, and poor prognosis. These data suggest that FGFR2 may be a potentially useful biological marker of tumor invasiveness in HCC as well as a novel molecular target for HCC.

Abstract 1448: Novel ALK5 inhibitor as an anti-metastasis agent in both breast cancer cells in vitro and MMTV/cNeu transgenic mice in vivo

Abstract We have synthesized novel ALK5 inhibitors and investigated their effects of on breast cancer cells as well as tumor bearing MMTV/cNeu transgenic mice and orthotopically injected mice. The treatment of ALK5 inhibitors into breast tumor bearing MMTV/cNeu transgenic mice and orthotopically injected mice inhibited lung metastasis compared to that of untreated animal. The level of ß-casein in the lung of the breast tumor bearing MMTV/cNeu transgenic mice and orthotopically injected mice were decreased by ALK5 inhibitor treatment. The levels of CXCR4, MMP2 and MMP9 were decreased in the lung and breast tumor of ALK5 inhibitors treated breast tumor bearing MMTV/cNeu transgenic mice and orthotopically injected mice. The action mechanisms of new ALK5 inhibitors were investigated in both breast cancer cells and breast cancer mice model. TGFß signaling, phosphorylated smad2/3 based from the western blot analysis in vitro and in vivo as well as confocal microscopic analysis in vitro was decreased by ALK5 inhibitors. And also, results from wound healing assay and invasion assay showed strong inhibition of TGFß action by new ALK5 inhibitors. This data strongly argue that ALK5 inhibitors have anti-metastasis effects via inhibiting ALK5 activity in vivo and in vitro. In order to gain further insight into the mechanism of ALK5 inhibitor, we have carried out experiment to examine the effects of ALK5 inhibitor in human breast cancer MDA-MB-231 cell. TGFß stimulated mesenchymal markers of epithelial mesenchymal transition (EMT), such as Snail, TWIST, N-cadherin and these were inhibited by ALK5 inhibitor concomitant treatment. Also, TGFß increased tumor invasion markers such as MMP2, and MMP9 which were inhibited by ALK5 inhibitor concomitant treatment. In addition, ALK5 inhibitors recover the level of E-cadherin, SnoN, Ski which were downregulated by TGFß treatment. In summary, ALK5 inhibitors have anti-metastatic effect on human breast cancer cells as well as tumor bearing MMTV/cNeu transgenic mice. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1448.