Head And Neck Cancer Cell Lines Research Articles

Abstract 4051: A mouse model of head and neck cancer with infrared-based molecular imaging

Abstract Introduction Head and neck cancer (HNC) is notoriously difficult to treat and the five-year survival rate has improved very little over the past thirty years. Head and neck squamous cell carcinomas (HNSCC) mouse models are invaluable tools for furthering mechanistic HNC research. Molecular imaging (MI) is a promising new modality that may improve current HNC mouse models. Visible florescent proteins (VFPs) have been widely used with animal models for tumor detection using MI. However, the low emission wavelength of VFP results in its overlapping with the auto-fluorescence of many tissues and causes insufficient penetration into deeper tissues, preventing the application of VFP to track local and metastatic tumor progressions. The recent development of infrared-fluorescent protein (IFP) with Biliverdin as a chromophore has greatly advanced in vivo molecular imaging of deep tissues in animals. Hypothesis The ultimate goal of this study is to develop a novel orthotopic mouse model of HNC with regional spreading and distant metastasis tumors that closely mimics human HNC and can be accurately detected by a noninvasive IFP-based molecular imaging (MI) system. Methods A stable human HNC cell line expressing IFP was established. This stable cell line was used for generating a mouse model with HNC. The tumor formation and tumor progression with this model were accurately monitored by a MI system. Results The IFP signal was clearly detected in our mouse model of HNC. Scanned imaging demonstrated a progression pattern of tumor growth in this model. The tumor can be detected earlier with molecular imaging than by conventional external caliper measurement. Unlike surgical measurement, the tumor can be quantified without disturbing the tumor environment. More important, our data suggest that local-regional metastasis tumors can be detected with this model. Conclusion This novel animal model represents an orthotopic human HNC model. The IFP signals penetrate tissues well and can be detected in the deep tissues with this model. Thus, our model has the potential to be used for monitoring tumor spreading and metastasis of advanced stage HNC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4051. doi:1538-7445.AM2012-4051

Abstract 1768: Enhanced cytotoxicity and apoptosis of cisplatin by p53-reactivating small molecule RITA in head and neck cancer

Abstract Overexpression of MDM2 inactivates the tumor suppressive function of p53 and a small-molecule reactivation of p53 and induction of tumor cell apoptosis (RITA) disrupts the MDM2-p53 interaction. We evaluated whether the restoration of p53 function by RITA enhances cytotoxicity and apoptosis of cisplatin in head-and-neck cancer (HNC). RITA was tested in four human HNC cell lines differing in TP53 status. In vitro and in vivo growth suppression, cell cycle arrest, apoptosis after RITA treatment individually or in combination with cisplatin were assessed. RITA induced prominent accumulation and reactivation of p53 in a wild-type TP53-bearing tumor cell line. RITA showed maximal growth suppression in tumor cells showing MDM2-dependent p53 degradation. RITA upregulated p21, BAX, and cleavage of caspase-3 but the p53-dependent apoptosis was blocked by pifithrin-μ. Strong induction of apoptosis rather than G2-phase arrest was observed along with increased dose of RITA. In combination therapy, RITA enhanced the growth inhibition and apoptosis of HNC by cisplatin in vitro and in vivo. Our data suggest that the restoration of p53 tumor suppressive function by RITA enhances cytotoxicity and apoptosis of cisplatin, which may offer an attractive strategy for treating HNC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1768. doi:1538-7445.AM2012-1768

S-Nitrosylation of mitogen activated protein kinase phosphatase-1 suppresses radiation-induced apoptosis

Radiotherapy is a key modality for head and neck cancer (HNC) treatment. Mitogen activated protein kinase phosphatase-1 (MKP-1) protein levels are elevated in various tumors and are negatively correlated with efficacy of chemo- or radio-therapy. However, the mechanisms underlying the moderate radiosensitivity of HNC and the increased MKP-1 protein levels are still dismal. Here we show that S-nitrosylation of MKP-1 on Cysteine 258 enhances MKP-1 protein stability, phosphatase activity, and MKP-1-mediated anti-apoptotic effect on HNC radiotherapy. Co-culturing MKP-1 transfected HNC cell lines with activated macrophages for mimicking the microenvironment of the irradiated cancer cells further confirms that S-nitrosylation-mediated increase of MKP-1 activity correlates with decrease of HNC radiosensitivity. Therefore, S-nitrosylation of MKP-1 presents a novel mechanism underlying the enhanced MKP-1 expression levels and MKP-1-mediated radio-resistance in head and neck cancer.

Molecular Chaperones as a Common Set of Proteins That Regulate the Invasion Phenotype of Head and Neck Cancer

The goal of this study was to establish a common set of molecules that regulate cell invasion in head and neck cancer (HNC). Five invasive sublines derived from HNC cell lines were established using the Matrigel selection method. Proteomic technology, MetaCore algorithm, and reverse transcriptase-PCR methods were used to search for molecules that contribute to the invasion phenotype. Cellular functional analyses and clinical association studies were applied to examine the significance of the molecules. Fifty-two proteins were identified in more than two of the four independent proteomic experiments, including 10 (19%) molecular chaperones. Seven chaperones were confirmed to be differentially expressed in five sublines, Hsp90α, Hsp90β, Hsp90-B1/Gp96, Hsp70-A5/Grp78, and HYOU1, that upregulate, whereas Hsp60 and glucosidase-α neutral AB (GANAB) downregulate. Four molecules were further investigated. In all cell lines, knockdown of Hsp60 or GANAB and silencing of Gp96 or Grp78 considerably enhanced or reduced cell migration and invasion, respectively. Clinical association studies consistently revealed that low levels of Hsp60 or GANAB and high levels of Gp96 or Grp78 are significantly associated with advanced cancer (P < 0.001 to P = 0.047, respectively, for the four molecules) and poor survival (P < 0.001 to P = 0.025, respectively, for the four molecules). Our study defined molecular chaperones as a common set of proteins that regulate the invasion phenotype of HNC. Loss of the tumor suppression function of Hsp60 or GANAB and acquisition of the oncogenic function of Gp96 or Grp78 contribute to aggressive cancers. These molecules may serve as prognostic markers and targets for cancer drug development.

Abstract 2358: Molecular chaperones as a common set of proteins that regulate the invasion phenotype of head and neck cancer

Abstract Purpose: The goal of this study was to establish a common set of molecules that regulate cell invasion in head and neck cancer (HNC). Experimental design: Five invasive sublines derived from HNC cell lines were established using the Matrigel selection method. Proteomic technology, MetaCore™ algorithm, and RT-PCR methods were used to search for molecules that contribute to the invasion phenotype. Cellular functional analyses and clinical association studies were applied to examine the significance of the molecules. Results: Fifty-two proteins were identified in more than two of the four independent proteomic experiments, including 10 (19%) molecular chaperones. Seven chaperones were confirmed to be differentially expressed in five sublines: Hsp90α, Hsp90β, Hsp90-B1/Gp96, Hsp70-A5/Grp78, HYOU1 up-regulation, while Hsp60 and GANAB down-regulation. Four molecules with the highest differential expressions (&amp;gt; 4-fold) were further investigated. In all cell lines, knockdown of Hsp60 or GANAB and silencing of Gp96 or Grp78 considerably enhanced or reduced cell migration and invasion, respectively. Clinical association studies consistently revealed that low levels of Hsp60 or GANAB and high levels of Gp96 or Grp78 are significantly associated with advanced cancer (P&amp;lt;0.001 to P = 0.047, respectively, for the four molecules) and poor survival (P&amp;lt;0.001 to P=0.025, respectively, for the four molecules). Conclusion: In conclusion, our study defined molecular chaperones as a common set of proteins that regulate the invasion phenotype of head and neck cancer. Loss of the tumor suppression function of Hsp60 or GANAB and acquisition of the oncogenic function of Gp96 or Grp78 contribute to aggressive cancers. These molecules may serve as prognostic markers and targets for cancer drug development. 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 2358. doi:10.1158/1538-7445.AM2011-2358

Abstract 3967: Identification of metadherin as a novel target of miR-375 in head and neck cancer

Abstract Purpose: The objective of this study is to investigate the significance of miR-375 and its target metadherin in head and neck cancer (HNC) patients. Methods and Results: Expression of miR-375 was measured by quantitative RT-PCR and observed to be significantly under expressed in both head and neck squamous cell (HNSCC) and nasopharyngeal carcinoma (NPC) samples by factors of 5.0 and 4.4, respectively. One mechanism of miR-375 silencing appears to be promoter methylation. Using a combination of experimentally-observed transcript changes after miR-375 re-expression in HNC cell lines, with in silico prediction algorithms, metadherin was identified as a novel target of miR-375. Luciferase reporter assays confirmed an inhibitory interaction between miR-375 and the 3’UTR of metadherin. Functional characterizations of miR-375 and metadherin were then conducted using FaDu (hypopharyngeal squamous cell) and C666-1 (nasopharyngeal) cancer cell lines. In vitro re-expression of miR-375 as well as knock-down of metadherin using siRNA markedly decreased cell viability and clonogenic survival (30% – 45%), cell migration (50% – 80%), and in vivo tumour forming ability in both cancer models. In vitro cell invasion was also significantly decreased by miR-375 over-expression and metadherin knock-down in FaDu cells (&amp;gt;90%). Western blot analyses demonstrated that metadherin knock-down resulted in reduced phosphorylation of PKB, indicating that the PI(3)K pathway may be involved in mediating these effects. Using qRT-PCR on 100 NPC and 22 HNSCC archival samples, there was an inverse relationship between metadherin over-expression with miR-375 under-expression, corroborating the relevance of this axis in primary human HNC tissues. Furthermore, NPC patients whose tumours expressed high levels of metadherin (defined as &amp;gt; median) experienced significantly lower overall and disease-free survival (5-year OS of 82%, DFS of 73%) compared to patients with low (&amp;lt; median) expression levels (5-year OS of 66%, DFS of 56%). Specifically, this reduced survival was most significantly related to distant metastases, wherein high metadherin expression was significantly associated with distant relapse (5-year distant relapse rates: 26% vs. 5% for high vs. low expressors, respectively). Conclusions: We report that the miR-375:metadherin control mechanism may represent a novel oncogenic pathway driving human HNC progression, potentially mediated in part by the PI(3)K cascade. Most importantly, metadherin over-expression was a significant predictor of distant metastases, a major clinical challenge in HNC given the improvements in loco-regional control as a consequence of technical advances in radiation therapy delivery. Hence, targeting this pathway could be a potentially novel therapeutic strategy to improve outcome for HNC patients. 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 3967. doi:10.1158/1538-7445.AM2011-3967

Abstract A165: Determining drug inhibition profiles and opportunities for synergistic combination therapies in head and neck cancer cell lines

Abstract Background: Metastatic head and neck cancer (HNC) remains poorly treatable and novel therapies are needed. We explored drug inhibition profiles in a large panel of head and neck cancer cell lines in order to select better therapies in the future and explore synergistic combinations. Methods: We measured the IC50 values of 23 clinically relevant small molecule inhibitors in 30 HNC cell lines by viability (resazurin). Hierarchical cluster analysis of single agent profiles was performed. Two cell lines were treated with combinations of two inhibitors to evaluate synergy via combinatorial index calculation (Chou). Signaling pathways were interrogated by immunoblot. Results: Single agents with a high degree of activity included bortezomib (lowest IC50 for SCC74: 15pM), dasatinib (HN13: 36pM), NVP-BEZ235 (SCC74: 50pM). However cluster analysis did not show consistent groupings, but similar agents clustered together. In both cell lines investigated for synergy — combinations of MET inhibitors with agents blocking GSK3-ß, Proteasome, src, EGFR, and PKC-β, as well as combinations of src inhibition with PI3K, HDAC, IGF1R, or PKC-β inhibition showed strong synergy by CI values. In SCC58 (non-HPV) inhibitors of EGFR, MET, IGF1-R, and HIF1 synergized with each other or with src or PKC-ß inhibition. In OSCC3 (HPV(+) MET inhibition synergized with everolimus (mTOR) or NVP-BEZ235 (PI3K/mTOR). Specific combinations with high degrees of synergy include: SU11274 (MET)/Erlotinib (EGFR): SCC58 CI50=0.29; OSCC3: CI50=0.53; PHA665752 (MET)/Bortezomib (Proteasome): OSCC3 CI50=0.41; IGF1-R-Inh./HIF1-Inh.: SCC58 CI50=0.33; IGF1-R-Inh./Enzastaurin (PKC-s): SCC58 CI50=0.15; Bortezomib (Proteasome)/Enzastaurin (PKC-ß): SCC58 CI50=0.38; Enzastaurin(PKC-ßs)/IGF-R-Inh.: OSCC3 CI50=0.72; Enzastaurin (PKC-ß):/SU11274 (MET): OSCC3 CI50=0.47; SB216763(GSK3-ß)/SU11274: OSCC3 CI50=0.52; Dasatinib (src)/Wortmannin (PI3K): OSCC3 CI50=0.64; SAHA/Dasatinib(src): OSCC3 CI50=0.58; SB216763 (GSK3-ß)/SU11274 (MET): OSCC3 CI50=0.5. Immunoblotting showed varying effects on several downstream signaling cascades. E.g. after combined treatment with PF-2341066 and Enzastaurin p-ERK signaling was reduced significantly more than after treatment with either agent alone. Conclusion: HNC cell lines show differential sensitivity to targeted agents and do not readily cluster based on drug inhibition. Nevertheless remarkable sensitivity in multiple cell lines was observed for certain inhibitors targeting src, the proteasome, and PI3K. Furthermore synergy was noted combining inhibitors of established targets in HNC (MET, EGFR, IGFR, and HIF1). Furthermore HPV-infected cell lines appear to be sensitive to combined inhibition of MET and the downstream PI3K/Akt/mTOR pathway. We are in the process of investigating the underlying etiology of specific drug sensitivities. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A165.

Sheddase Activity of Tumor Necrosis Factor-α Converting Enzyme Is Increased and Prognostically Valuable in Head and Neck Cancer

Tumor necrosis factor alpha converting enzyme (TACE) is a sheddase overexpressed in cancers that generates cancer cell growth and survival factors, and is implicated in carcinogenesis and tumor growth. This indicates that TACE could be a potentially important cancer biomarker. Unexpectedly, TACE expression in cancer tissues does not correlate with cancer stage or invasiveness. Although TACE sheddase activity is a more direct and potentially better indicator of TACE biology and might be a better cancer biomarker than TACE expression, it has not been studied in cancer tissues. In the present study, we developed a reliable specific assay for quantification of TACE sheddase activity, investigated TACE activity and TACE protein expression in head and neck cancer (HNC) tissues, and examined the correlation of the results with HNC clinical stages and likelihood to recur. We found that HNC cell lines and tissues contained remarkably higher quantities of TACE activity and TACE protein than normal keratinocytes or oral mucosa. siRNA silencing of TACE resulted in the inhibition of release of the tumorogenic factors amphiregulin and transforming growth factor alpha, and tumor protective factors tumor necrosis factor receptors from HNC cells. Importantly, TACE activity, but not TACE protein expression, was significantly higher in large, T3/T4, primary tumors relative to small, T1/T2, primary tumors, and especially in primary tumors likely to recur relative to those unlikely to recur. These data show that increased TACE activity in cancer is biologically and clinically relevant, and indicate that TACE activity could be a significant biomarker of cancer prognosis.

Whole-Cell Impedance Analysis for Highly and Poorly Metastatic Cancer Cells

A micro electrical impedance spectroscopy (muEIS) system has been developed and implemented to analyze highly and poorly metastatic head and neck cancer (HNC) cell lines with single-cell resolution. The microsystem has arrays of 16 impedance analysis sites, each of which is capable of capturing a single cell and analyzing its whole-cell electrical impedance spectrum. This muEIS system was used to obtain the electrical impedance spectra of the poorly metastatic HNC cell line 686 LN and the highly metastatic HNC cell line 686 LN-M4e over a frequency range of 40 Hz - 10 MHz. The 686 LN cells had higher impedance phase compared to that of 686 LN-M4e cells at frequencies between 50 kHz and 2 MHz. This result demonstrates that the metastatic state of HNC cells can be distinguished using the developed muEIS system. This system is expected to serve as a powerful tool for future detection and quantification of cancer cells from various tumor stages.

Glucose-regulated protein 78 regulates multiple malignant phenotypes in head and neck cancer and may serve as a molecular target of therapeutic intervention

Glucose-regulated protein 78 (Grp78) is an endoplasmic reticulum chaperone protein and is overexpressed in various cancers. However, it is unclear how significance of this molecule play an active role contributing to the oncogenic effect of head and neck cancer (HNC). To investigate the potential function of Grp78, six HNC cell lines were used. We found that Grp78 is highly expressed in all six cell lines and many of the proteins were localized in the periphery regions, implying other function of this molecule aside from endoplasmic reticulum stress response. Knockdown of Grp78 by small interfering RNA significantly reduced cell growth and colony formation to 53% to 12% compared with that of controls in all six HNC cell lines. Using in vitro wound healing and Matrigel invasion assays, we found that cell migration and invasive ability were also inhibited to 23% to 2% in all these cell lines tested. In vivo xenograft studies showed that administration of Grp78-small interfering RNA plasmid into HNC xenografts significantly inhibited both tumor growth in situ (>60% inhibition at day 34) and liver metastasis (>90% inhibition at day 20). Our study showed that Grp78 actively regulates multiple malignant phenotypes, including cell growth, migration, and invasion. Because knockdown Grp78 expression succeeds in the reduction of tumor growth and metastatic potential, this molecule may serve as a molecular target of therapeutic intervention for HNC.

DNA Methylation: Its Role in Cancer Development and Therapy

Ferroptosis is a newly defined form of cell death induced by iron-dependent accumulation of lethal lipid peroxidation. Ferroptosis represent a therapeutic strategy to suppress therapy-resistant cancer cells with more property of epithelial-mesenchymal transition (EMT). However, epigenetic reprogramming of EMT has been rarely studied in the context of ferroptosis susceptibility. Therefore, we examined the therapeutic potentiality of EMT epigenetic reprogramming in promoting ferroptosis in head and neck cancer (HNC) cells. The effects of ferroptosis inducers and EMT inhibition or induction were tested in HNC cell lines and mouse tumor xenograft models. These effects were analyzed concerning cell viability and death, lipid reactive oxygen species and iron production, labile iron pool, glutathione contents, NAD/NADH levels, and mRNA/protein expression. Cell density and the expression levels of E-cadherin, vimentin, and ZEB1 were associated with the different susceptibility to ferroptosis inducers. CDH1 silencing or ZEB1 overexpression increased the susceptibility to ferroptosis, whereas CDH overexpression or ZEB1 silencing decreased the susceptibility, in vitro and in vivo. Histone deacetylase SIRT1 gene silencing or pharmacological inhibition by EX-527 suppressed EMT and consequently decreased ferroptosis, whereas SIRT inducers, resveratrol and SRT1720, increased ferroptosis. MiR-200 family inhibitors induced EMT and increased ferroptosis susceptibility. In HNC cells with low expression of E-cadherin, the treatment of 5-azacitidine diminished the hypermethylation of CDH1, resulting in increased E-cadherin expression and decreased ferroptosis susceptibility. Our data suggest that epigenetic reprogramming of EMT contributes to promoting ferroptosis in HNC cells.

Wnt signaling controls radiosensitivity via cyclooxygenase‐2‐mediated Ku expression in head and neck cancer

It has been proposed that Wnt signaling pathway may be a key radioprotective mechanism in irradiated cancer cells; however, the specific radioresistance mechanisms remain not to be fully clarified. Here we elucidate a novel signaling pathway of radioresistance in head and neck cancer (HNC) cell lines involving interactions among the Wnt signaling pathway, cyclooxygenase-2 (COX-2) and Ku expression. Activation of the Wnt signaling pathway by (2'Z,3'E)-6-bromoindirubin-3'-oxime (BIO) resulted in beta-catenin cytoplasmic accumulation and translocation to the nucleus, upregulated Ku expression and increased radioresistance in the COX-2-expressing HNC cell line. In contrast, Wnt singaling activation by BIO had no effects on Ku expression and radiosensitivity in a HNC cell line negative for COX-2. Interactions between Wnt singaling and Ku were indirectly regulated by COX-2. Blockage of COX-2 signaling led to the suppression of beta-catenin-induced Ku expression, and to consequent recovery of the radiosensitivity in HNC cells. Our results conclusively suggest that beta-catenin plays a pivotal role in the regulation of Ku expression via the proposed COX-2 intracellular pathway, thus supporting a novel radioresistance mechanism of HNC.

Enhanced Gene Transfer and Oncolysis of Head and Neck Cancer and Melanoma Cells by Fiber Chimeric Oncolytic Adenoviruses

The purpose of this study was to evaluate a fiber knob replacement strategy to improve infectivity and efficacy of Ad5 fiber chimeric oncolytic viruses for treatment of melanoma and head and neck cancers (HNC). Adenoviral receptors and transduction levels were used to determine the level of infectivity of fiber-modified, green fluorescent protein-expressing, replication-deficient viruses in a panel of melanoma and HNC cell lines in vitro. Virus yield and cytotoxicity assays were used to determine the tumor specificity and virus replication-mediated cytotoxicity of the fiber-modified oncolytic viruses in the same panel of melanoma and HNC in vitro. Xenograft tumor models were used to assess the antitumor activity of those fiber-modified chimeric viruses compared with the parental virus. Marker gene expression following gene transfer of the fiber chimeric vectors in melanoma and HNC cell lines was approximately 10-fold higher than that obtained with parental Ad5 vector. The fiber chimeric oncolytic variants mediated killing of melanoma and HNC cells that was 2- to 576-fold better than with the parental virus. In addition, fiber chimeric variants produced 2- to 7-fold more progeny virus in tumor cells than the parental virus. Moreover, a high multiplicity of infection was needed for the fiber chimeric viruses to produce cytotoxicity in normal cells. A significantly stronger antitumor response and survival advantage were shown in the tested melanoma and HNC xenograft models following i.t. injections. In vitro and in vivo studies showed the improved transduction, replication, cytotoxicity, antitumor efficacy, and survival advantage in melanoma and HNC tumor models, suggesting a potential use of these oncolytic agents for the treatment of melanoma and HNCs.

545. Adenovirus Mediated Gene Transfer and Oncolysis In Vitro in Head and Neck Cancer and Melanoma Cells Enhanced by Polycations

Oncolytic adenoviruses (OAVs) designed to selectively replicate and kill cancer cells are rapidly emerging as promising agents for oncolytic virotherapy. We have generated and evaluated several OAVs both in vitro and in vivo in clinical trials. In several pre-clinical and clinical studies, OAVs as monotherapy have shown limited efficacy for treatment of head and neck cancer (HNC) and melanoma. This limited efficacy may be due to low or no expression of coxsackievirus and adenovirus receptors (CAR) on the surface of cancer cells. In addition, OAV-mediated gene transfer into cancer cells may be limited by electrostatic repulsion between the net negatively charged viral capsid and negatively charged cell surface. Removal of negatively charged molecules from cell surface or neutralization of negative charge on adenovirus capsid has been demonstrated to significantly improve Ad5-mediated gene transfer. Here, the effect of polycations such as polybrene, protamine and cytopure on Ad5 mediated gene transfer into cancer cells was investigated. Pretreatment of Ad5 marker vector (Ad5-GFP) with the polycations increased transgene expression up to 65-fold in several HNC cell lines. Similarly, in several melanoma cell lines Ad5-mediated gene expression was enhanced by up to 121-fold. In addition, treatment of an OAV with polycations has improved the virus yields by more than 100-fold in both HNC and melanoma cell lines. Moreover, pretreatment of virus with polycations partially protected the virus in vitro from neutralizing antibodies. These findings may have important implications for the treatment of head and neck cancers and melanoma using oncolytic adenoviral vectors.

538. Fiber Chimeric Oncolytic Adenoviral Vectors Mediate Efficient Gene Transfer and Oncolysis of Head and Neck Cancer and Melanoma Cells In Vitro and In Vivo

Current treatment modalities for melanoma and head and neck cancers (HNC) are not completely effective and there is an urgent need for development of novel therapeutic strategies. Oncolytic adenoviruses (OAVs) designed to selectively replicate and kill cancer cells are rapidly emerging as promising agents for oncolytic virotherapy. However, low expression levels of the adenovirus receptors, CAR and integrins in melanoma and HNC cells have imposed significant hurdles to the use of adenovirus type 5 (Ad5)-based vectors. To improve gene transfer and mediate efficient oncolysis of these cells, fiber chimeric vectors in which the fiber knob was replaced with the fiber knob domain of Ad35 were generated. Marker gene expression mediated by the fiber chimeric vectors in melanoma and HNC cell lines was approximately 10-fold higher than that obtained with parental Ad5 virus. The fiber chimeric structure was then built into an oncolytic adenovirus, in which the expression of the E1a gene is placed under the control of E2F-1 promoter. The cytotoxicity mediated by fiber chimeric vectors in melanoma and HNC cells was 100-fold better than that mediated by the parental Ad5 vector. In addition, the fiber chimeric structures improved the virus yields as much as 100-fold in several melanoma and HNC cells. These results correlated with low levels of CAR and high levels of CD46 on the surface of the cancer cell lines. The results of in vitro transduction, cytotoxicity and virus yields assays correlated with the results of in vivo anti-tumor efficacy studies in FaDu xenograft tumor model in nude mice. These findings have important implications for the potential treatment of melanoma and HNC using fiber chimeric oncolytic adenovirus.