Han Si

ΔNp63 Versatilely Regulates a Broad NF-κB Gene Program and Promotes Squamous Epithelial Proliferation, Migration, and Inflammation

Head and neck squamous cell carcinoma (HNSCC) and many other epithelial malignancies exhibit increased proliferation, invasion, and inflammation, concomitant with aberrant nuclear activation of TP53 and NF-κB family members ΔNp63, cRel, and RelA. However, the mechanisms of cross-talk by which these transcription factors coordinate gene expression and the malignant phenotype remain elusive. In this study, we showed that ΔNp63 regulates a cohort of genes involved in cell growth, survival, adhesion, and inflammation, which substantially overlaps with the NF-κB transcriptome. ΔNp63 with cRel and/or RelA are recruited to form novel binding complexes on p63 or NF-κB/Rel sites of multitarget gene promoters. Overexpressed ΔNp63- or TNF-α-induced NF-κB and inflammatory cytokine interleukin-8 (IL-8) reporter activation depended on RelA/cRel regulatory binding sites. Depletion of RelA or ΔNp63 by small interfering RNA (siRNA) significantly inhibited NF-κB-specific, or TNF-α-induced IL-8 reporter activation. ΔNp63 siRNA significantly inhibited proliferation, survival, and migration by HNSCC cells in vitro. Consistent with these data, an increase in nuclear ΔNp63, accompanied by increased proliferation (Ki-67) and adhesion (β4 integrin) markers, and induced inflammatory cell infiltration was observed throughout HNSCC specimens, when compared with the basilar pattern of protein expression and minimal inflammation seen in nonmalignant mucosa. Furthermore, overexpression of ΔNp63α in squamous epithelial cells in transgenic mice leads to increased suprabasilar cRel, Ki-67, and cytokine expression, together with epidermal hyperplasia and diffuse inflammation, similar to HNSCC. Our study reveals ΔNp63 as a master transcription factor that, in coordination with NF-κB/Rels, orchestrates a broad gene program promoting epidermal hyperplasia, inflammation, and the malignant phenotype of HNSCC.

Aberrant IKKα and IKKβ cooperatively activate NF-κB and induce EGFR/AP1 signaling to promote survival and migration of head and neck cancer

The inhibitor-κB kinase-nuclear factor-κB (IKK-NF-κB) and epidermal growth factor receptor-activator protein-1 (EGFR-AP1) pathways are often co-activated and promote malignant behavior, but the underlying basis for this relationship is unclear. Resistance to inhibitors of IKKβ or EGFR is observed in head and neck squamous cell carcinomas (HNSCC). Here, we reveal that both IKKα and β contribute to nuclear activation of canonical and alternate NF-κB/REL family transcription factors, and overexpression of signal components that enhance co-activation of the EGFR-AP1 pathway. We observed that IKKα and IKKβ exhibit increased protein expression, nuclear localization, and phosphorylation in HNSCC tissues and cell lines. Individually, IKK activity varied among different cell lines, but overexpression of both IKKs induced the strongest NF-κB activation. Conversely, siRNA knock down of both IKKs significantly decreased nuclear localization and phosphorylation of canonical RELA and IκBα and alternative p52 and RELB subunits. Knock down of both IKKs more effectively inhibited NF-κB activation, broadly modulated gene expression and suppressed cell proliferation and migration. Global expression profiling revealed that NF-κB, cytokine, inflammatory response and growth factor signaling are among the top pathways and networks regulated by IKKs. Importantly, IKKα and IKKβ together promoted the expression and activity of transforming growth factor α, EGFR and AP1 transcription factors cJun, JunB and Fra1. Knock down of AP1 subunits individually decreased 8/15 (53%) of IKK-targeted genes sampled and similarly inhibited cell proliferation and migration. Mutations of NF-κB and AP1-binding sites abolished or decreased IKK-induced interleukin-8 (IL-8) promoter activity. Compounds such as wedelactone with dual IKK inhibitory activity and geldanomycins that block IKKα/β and EGFR pathways were more active than IKKβ-specific inhibitors in suppressing NF-κB activation and proliferation and inducing cell death. We conclude that IKKα and IKKβ cooperatively activate NF-κB and EGFR/AP1 networks of signaling pathways and contribute to the malignant phenotype and the intrinsic or acquired therapeutic resistance of HNSCC.

TNF-α modulates genome-wide redistribution of ΔNp63α/TAp73 and NF-κB cREL interactive binding on TP53 and AP-1 motifs to promote an oncogenic gene program in squamous cancer.

The Cancer Genome Atlas (TCGA) network study of 12 cancer types (PanCancer 12) revealed frequent mutation of TP53, and amplification and expression of related TP63 isoform ΔNp63 in squamous cancers. Further, aberrant expression of inflammatory genes and TP53/p63/p73 targets were detected in the PanCancer 12 project, reminiscent of gene programs comodulated by cREL/ΔNp63/TAp73 transcription factors we uncovered in head and neck squamous cell carcinomas (HNSCCs). However, how inflammatory gene signatures and cREL/p63/p73 targets are comodulated genome wide is unclear. Here, we examined how the inflammatory factor tumor necrosis factor-α (TNF-α) broadly modulates redistribution of cREL with ΔNp63α/TAp73 complexes and signatures genome wide in the HNSCC model UM-SCC46 using chromatin immunoprecipitation sequencing (ChIP-seq). TNF-α enhanced genome-wide co-occupancy of cREL with ΔNp63α on TP53/p63 sites, while unexpectedly promoting redistribution of TAp73 from TP53 to activator protein-1 (AP-1) sites. cREL, ΔNp63α and TAp73 binding and oligomerization on NF-κB-, TP53- or AP-1-specific sequences were independently validated by ChIP-qPCR (quantitative PCR), oligonucleotide-binding assays and analytical ultracentrifugation. Function of the binding activity was confirmed using TP53-, AP-1- and NF-κB-specific REs or p21, SERPINE1 and IL-6 promoter luciferase reporter activities. Concurrently, TNF-α regulated a broad gene network with cobinding activities for cREL, ΔNp63α and TAp73 observed upon array profiling and reverse transcription–PCR. Overlapping target gene signatures were observed in squamous cancer subsets and in inflamed skin of transgenic mice overexpressing ΔNp63α. Furthermore, multiple target genes identified in this study were linked to TP63 and TP73 activity and increased gene expression in large squamous cancer samples from PanCancer 12 TCGA by CircleMap. PARADIGM inferred pathway analysis revealed the network connection of TP63 and NF-κB complexes through an AP-1 hub, further supporting our findings. Thus, inflammatory cytokine TNF-α mediates genome-wide redistribution of the cREL/p63/p73, and AP-1 interactome, to diminish TAp73 tumor suppressor function and reciprocally activate NF-κB and AP-1 gene programs implicated in malignancy.

Abstract 4804: High throughput whole exome DNA and transcriptome RNA sequencing to identify genetic drivers and alterations in HPV-negative and HPV-positive HNSCC cell lines

Head and neck squamous cell carcinoma (HNSCC) is among the top cancer types with high frequencies of genetic alterations, including mutation and copy number variation (CNV). The Cancer Genome Atlas (TCGA) has profiled over 279 HNSCC tumors and generated a comprehensive genomic characterization of HNSCC. This has led to an urgent need for a panel of head and neck cell line models with genomic alterations representative of those found by TCGA. We performed whole exome DNA sequencing (exome DNA-seq) and transcriptome RNA sequencing (RNA-seq) on 15 HPV negative and 11 HPV positive HNSCC lines, which were compared with three normal human oral mucosa lines and 8 matched blood samples. Exome DNA- and RNA-seq were performed on the ABI SOLiD platform with an average depth of 87X and 44X respectively. Using an in-house analysis pipeline, we determined the CNVs and single nucleotide variants (SNV) obtained from DNA-seq to be able to compare with the genomic alterations found in TCGA, and also to cross-validate these with the SNVs identified in our RNA-seq. We identified chromosome losses in 3p, 5q, 8p, 9p and 18q and gains in 3q, 7p and 11q in a significant portion of cell lines with software CONTRA (COpy Number Targeted Resequencing Analysis), which are consistent with previous karyotype and TCGA CNV studies. Integrative analysis between CNV by exome-seq and gene expression by RNA-seq of these cell lines revealed a significant positive correlation in multiple oncogenes including PIK3CA, TP63, CCND1, FADD, BIRC2 and YAP1, which is in concordance with TCGA results. We established a workflow to determine deleterious mutations and somatic mutations using software ANNOVAR, in combination with functional prediction tool Mutation Assessor, and Sanger Institute9s somatic mutation database, COSMIC, in order to characterize legacy and newer HNSCC lines without and with matched samples. We identified a median of 1588 potentially deleterious and/or somatic mutations for each cell line. The most recurrently mutated genes in TCGA with a functional impact are also frequently mutated in cell lines, including TP53, FAT1 and NOTCH1, etc. Many of the genomic alterations identified converge on the networks we previously defined in HNSCC, including the PI3K/AKT/mTOR, NFκB, and RAS/MAPK pathways. Our findings suggest that these cell lines can serve as HNSCC models for mechanistic and therapeutic studies, and thereby provide a valuable resource for the wider biomedical research community. Supported by NIDCD intramural projects ZIA-DC-000016, 73 and 74. Citation Format: Hui Cheng, Xinping Yang, Han Si, Anthony Saleh, Jamie Coupar, Robert L. Ferris, Wendell G. Yarbrough, Mark E. Prince, Thomas E. Carey, Carter Van Waes, Zhong Chen. High throughput whole exome DNA and transcriptome RNA sequencing to identify genetic drivers and alterations in HPV-negative and HPV-positive HNSCC cell lines. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4804. doi:10.1158/1538-7445.AM2015-4804

Abstract 4968: TNF-alpha promotes genome-wide replacement of TAp73 chromatin occupancy by cREL and DeltaNp63

cREL, DeltaNp63, and TAp73, members of NFkappaB and TP53 transcription factor (TF) families, together regulate broad biological processes critical for inflammation and cancer survival. However, the underlying mechanisms explaining their cross-regulation of transcriptome and interactive signal network have not been revealed. Here, we demonstrate that inflammatory cytokine TNF-alpha induces a dynamic, genome-wide increase in cREL and DeltaNp63 chromatin occupancy after replacing TAp73 binding activity, using ChIP-seq and fitting the data into a negative binomial distribution model integrated in CisGenome. Globally, most of the binding peaks were localized in intergenic and intronic areas, with 4.2% and 5.6% peaks significantly enriched into promoter regions (3000 bp upstream of transcription starting sites) of cREL and DeltaNp63 bound DNAs in response to TNF-alpha respectively, and co-localized peaks for any of the two TFs are prevalent within a distance of ∼200 bp. De novo motif search using Gibbs Sampler identified TP53 and novel binding motifs. A total of 188 differentially expressed genes are associated with the TNF-alpha-induced chromatin occupancy shift by the three TFs. These genes are enriched in cancer-related transcription factors and TP53 signaling pathways. Our data reveal a novel mechanism of genome-wide replacement of TAp73 by cREL and DeltaNp63, supporting cross-regulation and an interactive network between NFkappaB and TP53 families. 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 4968. doi:10.1158/1538-7445.AM2011-4968

Abstract 4374: Chromosome 3q22-29 amplification is linked to increased expression of multiple genes in key pathways deregulated in head and neck cancer tumors and cell lines

As part of the Cancer Genome Atlas (TCGA) Network, our comprehensive genomic analysis of 279 head and neck squamous cell carcinomas (HNSCCs) found frequent chromosomal copy number variation (CNV) and mutations of potential biologic and therapeutic importance. This underscored an urgent need to identify cell line models that harbor genomic alterations representative of HNSCC. We performed whole exome-DNA and transcriptome RNA sequencing on 15 human papillomavirus HPV(−) and 11 HPV(+) HNSCC cell lines. HNSCC lines harbored chromosome gains (3q, 5p, 7p, 8q, 11q) and losses (3p, 5q, 8p, 9p, 18q), consistent with those found in HNSCC tumors by TCGA and previous karyotype studies. Integrative genome-wide analysis of CNV with gene expression uncovered over 1500 genes that display significant correlation between CNV and gene expression in both TCGA tumors and cell lines. Ingenuity Pathway Analysis revealed multiple genes that converge on key pathways and functions deregulated in HNSCC, including PI3K/AKT/mTOR, NF-κB, RAS/MAPK, TP53, death receptor signaling, inflammation, and differentiation. Intriguingly, 103 genes displaying significant amplification and increased expression were predominately located on chromosome 3q22-29. These genes encode components involved in the PI3K/AKT/mTOR, Hippo, TGF-beta and Wnt/beta-catenin pathways, cell cycle, translational and post-translational regulation, and mitochondrial biosynthesis. Fisher’s exact test and survival analysis showed significant co-occurrence and worse overall survival of 3q26.3 amplification and TP53 mutation in HNSCC patients from TCGA datasets; 3q26.3 encompasses 53 genes including PIK3CA, PLD1, ACTL6A and SOX2. HNSCC cell lines also harbor common mutations found in TCGA, such as TP53, FAT1 and NOTCH1, and novel and rare tumor suppressor genes, such as MYH9. Our findings suggest that these cell lines could serve as models for mechanistic studies and pharmacologic screening, and investigation of genomic and expression alterations as potential biomarkers for precision diagnosis and prognosis of HNSCC. (Supported by NIDCD/NIH intramural projects ZIA-DC-000073, ZIA-DC-000074. This study utilized the high-performance computational capabilities of the Biowulf Linux cluster at the National Institutes of Health, Bethesda, MD. (http://biowulf.nih.gov).) Citation Format: Hui Cheng, Xinping Yang, Han Si, Anthony Saleh, Jamie Coupar, Robert L. Ferris, Wendell G. Yarbrough, Mark E. Prince, Thomas E. Carey, Carter Van Waes, Zhong Chen. Chromosome 3q22-29 amplification is linked to increased expression of multiple genes in key pathways deregulated in head and neck cancer tumors and cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4374. doi:10.1158/1538-7445.AM2017-4374

TNF-α modulates genome-wide redistribution ofΔNp63α/TAp73 and NF-κB c-REL interactive binding on TP53and AP-1 motifs to promote an oncogenic gene program in squamouscancer

The Cancer Genome Atlas (TCGA) network study of 12 cancer types (PanCancer 12) revealed frequent mutation of TP53, and amplification and expression of related TP63 isoform ΔNp63 in squamous cancers. Further, aberrant expression of inflammatory genes and TP53/p63/p73 targets were detected in the PanCancer 12 project, reminiscent of gene programs comodulated by cREL/ΔNp63/TAp73 transcription factors we uncovered in head and neck squamous cell carcinomas (HNSCCs). However, how inflammatory gene signatures and cREL/p63/p73 targets are comodulated genome wide is unclear. Here, we examined how the inflammatory factor tumor necrosis factor-α (TNF-α) broadly modulates redistribution of cREL with ΔNp63α/TAp73 complexes and signatures genome wide in the HNSCC model UM-SCC46 using chromatin immunoprecipitation sequencing (ChIP-seq). TNF-α enhanced genome-wide co-occupancy of cREL with ΔNp63α on TP53/p63 sites, while unexpectedly promoting redistribution of TAp73 from TP53 to activator protein-1 (AP-1) sites. cREL, ΔNp63α and TAp73 binding and oligomerization on NF-κB-, TP53- or AP-1-specific sequences were independently validated by ChIP-qPCR (quantitative PCR), oligonucleotide-binding assays and analytical ultracentrifugation. Function of the binding activity was confirmed using TP53-, AP-1- and NF-κB-specific REs or p21, SERPINE1 and IL-6 promoter luciferase reporter activities. Concurrently, TNF-α regulated a broad gene network with cobinding activities for cREL, ΔNp63α and TAp73 observed upon array profiling and reverse transcription–PCR. Overlapping target gene signatures were observed in squamous cancer subsets and in inflamed skin of transgenic mice overexpressing ΔNp63α. Furthermore, multiple target genes identified in this study were linked to TP63 and TP73 activity and increased gene expression in large squamous cancer samples from PanCancer 12 TCGA by CircleMap. PARADIGM inferred pathway analysis revealed the network connection of TP63 and NF-κB complexes through an AP-1 hub, further supporting our findings. Thus, inflammatory cytokine TNF-α mediates genome-wide redistribution of the cREL/p63/p73, and AP-1 interactome, to diminish TAp73 tumor suppressor function and reciprocally activate NF-κB and AP-1 gene programs implicated in malignancy.

TNF-|[alpha]| modulates genome-wide redistribution of |[Delta]|Np63|[alpha]||[sol]|TAp73 and NF-|[kappa]|B cREL interactive binding on TP53 and AP-1 motifs to promote an oncogenic gene program in squamous cancer

The Cancer Genome Atlas (TCGA) network study of 12 cancer types (PanCancer 12) revealed frequent mutation of TP53, and amplification and expression of related TP63 isoform ΔNp63 in squamous cancers. Further, aberrant expression of inflammatory genes and TP53/p63/p73 targets were detected in the PanCancer 12 project, reminiscent of gene programs comodulated by cREL/ΔNp63/TAp73 transcription factors we uncovered in head and neck squamous cell carcinomas (HNSCCs). However, how inflammatory gene signatures and cREL/p63/p73 targets are comodulated genome wide is unclear. Here, we examined how the inflammatory factor tumor necrosis factor-α (TNF-α) broadly modulates redistribution of cREL with ΔNp63α/TAp73 complexes and signatures genome wide in the HNSCC model UM-SCC46 using chromatin immunoprecipitation sequencing (ChIP-seq). TNF-α enhanced genome-wide co-occupancy of cREL with ΔNp63α on TP53/p63 sites, while unexpectedly promoting redistribution of TAp73 from TP53 to activator protein-1 (AP-1) sites. cREL, ΔNp63α and TAp73 binding and oligomerization on NF-κB-, TP53- or AP-1-specific sequences were independently validated by ChIP-qPCR (quantitative PCR), oligonucleotide-binding assays and analytical ultracentrifugation. Function of the binding activity was confirmed using TP53-, AP-1- and NF-κB-specific REs or p21, SERPINE1 and IL-6 promoter luciferase reporter activities. Concurrently, TNF-α regulated a broad gene network with cobinding activities for cREL, ΔNp63α and TAp73 observed upon array profiling and reverse transcription–PCR. Overlapping target gene signatures were observed in squamous cancer subsets and in inflamed skin of transgenic mice overexpressing ΔNp63α. Furthermore, multiple target genes identified in this study were linked to TP63 and TP73 activity and increased gene expression in large squamous cancer samples from PanCancer 12 TCGA by CircleMap. PARADIGM inferred pathway analysis revealed the network connection of TP63 and NF-κB complexes through an AP-1 hub, further supporting our findings. Thus, inflammatory cytokine TNF-α mediates genome-wide redistribution of the cREL/p63/p73, and AP-1 interactome, to diminish TAp73 tumor suppressor function and reciprocally activate NF-κB and AP-1 gene programs implicated in malignancy.

Abstract 3435: Genome-wide RNA and DNA high throughput sequencing reveals proinflammatory and death gene signatures in head and neck squamous cell carcinoma lines with different HPV status

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Our laboratory previously discovered that aberrant NF-κB signaling and activation plays a critical role in promoting the oncogenic progression of head and neck squamous cell carcinoma (HNSCC). The Cancer Genome Atlas (TCGA) project, which investigated 279 HNSCC tissue specimens, uncovered significant genomic alterations of key molecules involved in NF-κB and death signaling, including amplification of FADD and BIRC2/3, mutation of caspase-8, and altered RIPKs in HPV(-), or deletion of TRAF3 in HPV(+) HNSCC tissues. Using PRISM (Protein Interactions by Structural Matching), we modeled protein-protein interaction complexes three dimensionally and identified ∼30 proteins that potentially interact with these genetically altered molecules involved in NF-κB and death pathways. Among more than 20 major cancer types that TCGA project has investigated, we found that HNSCC ranks among the top cancers harboring alterations of these genes, which are present in more than 70% of HNSCC tumors. To further investigate and identify models for functional studies of these genetic and phenotypic alterations, we performed whole transcriptome (RNA-seq) and genome-wide exome DNA sequencing (exome DNA-seq) in 16 HPV(-) and 8 HPV(+) HNSCC lines, and compared them with three normal human oral mucosa lines and 8 matched blood samples. RNA-seq or exome DNA-seq was performed using total RNA depleted of rRNA, or exome captured genomic DNA. The samples were multiplexed, amplified, and sequenced with ABI SOLiD sequencer. Quality control of the output data was performed using FastQC, and the reads were mapped to the human reference genome (Hg19). The data were further analyzed with LifeScope2.5 and an in-house developed computation program of publicly available software and R package. RNA-seq had an average of 168 million mapped reads per sample with an estimate mean read depth of ∼44X coverage, and exome DNA-seq had an average of ∼87X coverage in each cell line sequenced. We identified amplifications, overexpression and mutations among the molecules involved in the NF-κB and death pathways in HPV positive or negative cell lines, which mimic the genetic and phenotypic defects identified in the HNSCC TCGA project. Our data are consistent with the finding from the TCGA project, and support the conclusion that genetic and expression alterations of the molecules involved in NF-κB and the death pathways are the important events leading HNSCC oncogenesis. Molecular mechanisms and targeted therapies can be further investigated in these HNSCC lines having the defined genetic and phenotypic defects associated with different HPV status. Supported by NIDCD intramural projects ZIA-DC-000016, 73 and 74. Citation Format: Xinping Yang, Hui Cheng, Han Si, Anthony Saleh, Emine Guven Maiorov, Jamie Coupar, Ozlem Keskin, Attila Gursoy, Ruth Nussinov, Robert L. Ferris, Wendell G. Yarbrough, Mark E. Prince, Thomas E. Carey, Carter Van Waes, Zhong Chen. Genome-wide RNA and DNA high throughput sequencing reveals proinflammatory and death gene signatures in head and neck squamous cell carcinoma lines with different HPV status. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3435. doi:10.1158/1538-7445.AM2014-3435

Abstract 3434: Genomic exome DNA sequencing identifies top driver genetic alterations in head and neck cancer cell lines of different HPV status

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Head and neck squamous cell carcinoma (HNSCC) is among the top cancer types with high frequencies of genetic alterations, including mutation and copy number variation (CNV). The Cancer Genome Atlas (TCGA) has characterized 279 HNSCC tumors to provide a landscape of comprehensive genomic alterations. To identify cell line models which mimic HNSCC subtypes of genetic alterations, we have conducted genome-wide exome DNA sequencing (exome DNA-seq) and whole transcriptome sequencing (RNA-seq) on 16 HPV negative and 8 HPV positive HNSCC lines, in comparison with three normal human oral mucosa lines and 8 matched blood samples. Exome DNA-seq and RNA-seq were performed using an ABI SOLiD sequencer, and the data were analyzed with LifeScope2.5 and an in-house developed computation program. An average of 87X coverage in each cell line was achieved by exome DNA-seq. Deleterious mutations that negatively affect protein function were identified in each cell line using ANNOVAR software, in combination with multiple functional prediction tools, such as SIFT, PolyPhen-2, Likelihood Ratio Test (LRT), MutationTaster and MutationAssessor. The top deleterious mutations include TP53 in 42% of cell lines (56% in HPV- and 13% in HPV+ cell lines), and FAT1 and NOTCH1 mutations in 83% and 42% cell lines, respectively. These mutations are cross-validated through RNA-seq, which has an average of 168 million mapped reads with an estimated mean of 44X coverage per sample. In addition, we identified 261 putative CNV calls using the CoNIFER (copy number inference from exome reads) analysis. Among these putative CNV calls, 12 of them occur inside of the over 30 significant somatic copy number alteration (SCNA) sites identified in TCGA, including amplification of FADD, BIRC2/3, YAP1 and TP63. Our sequencing results identified the HNSCC cell lines with specific genetic alterations, which resemble many defects found from tumor specimens in TCGA project. These cell lines provided experimental models, which are amenable for future investigation of molecular mechanisms and targeted therapy, and could benefit preclinical drug evaluation and provide rationales for future clinical trials with genetic testing. Supported by NIDCD intramural projects ZIA-DC-000016, 73 and 74. Citation Format: Hui Cheng, Xinping Yang, Han Si, Anthony Saleh, Jamie Coupar, Robert L. Ferris, Wendell G. Yarbrough, Mark E. Prince, Thomas E. Carey, Carter Van Waes, Zhong Chen. Genomic exome DNA sequencing identifies top driver genetic alterations in head and neck cancer cell lines of different HPV status. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3434. doi:10.1158/1538-7445.AM2014-3434