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Genetic variants located within the 12p13.33/RAD52 locus have been associated with lung squamous cell carcinoma (LUSC). Here, within 5,947 UADT cancers and 7,789 controls from 9 different studies, we found rs10849605, a common intronic variant in RAD52, to be also associated with upper aerodigestive tract (UADT) squamous cell carcinoma cases (OR = 1.09, 95% CI: 1.04-1.15, p = 6x10(-4)). We additionally identified rs10849605 as a RAD52 cis-eQTL inUADT(p = 1x10(-3)) and LUSC (p = 9x10(-4)) tumours, with the UADT/LUSC risk allele correlated with increased RAD52 expression levels. The 12p13.33 locus, encompassing rs10849605/RAD52, was identified as a significant somatic focal copy number amplification in UADT(n = 374, q-value = 0.075) and LUSC (n = 464, q-value = 0.007) tumors and correlated with higher RAD52 tumor expression levels (p = 6x10(-48) and p = 3x10(-29) in UADT and LUSC, respectively). In combination, these results implicate increased RAD52 expression in both genetic susceptibility and tumorigenesis of UADT and LUSC tumors.

MK5 (MAPK-activated protein kinase 5) or PRAK (p38-regulated and -activated kinase) are alternative names for a serine/threonine protein kinase downstream to ERK3/4 and p38 MAPK. A previous gene targeting approach for MK5/PRAK (termed here MK5/PRAK-Δex8) revealed a seemingly tumor-suppressive role of MK5/PRAK in DMBA-induced one step skin carcinogenesis and Ras-induced transformation. Here we demonstrate that an alternative targeting strategy of MK5/PRAK (termed MK5/PRAK-Δex6) increased neither tumor incidence in the one step skin carcinogenesis model, nor Ras-induced transformation in primary cells. Interestingly, due to the targeting strategies and exon skipping both knockouts do not completely abolish the generation of MK5/PRAK protein, but express MK5/PRAK deletion mutants with different biochemical properties depending on the exon targeted: Targeting of exon 6 leads to expression of an unstable cytoplasmic catalytically inactive MK5/PRAK-Δex6 mutant while targeting of exon 8 results in a more stable nuclear MK5/PRAK-Δex8 mutant with residual catalytic activity. The different properties of the MK5/PRAK deletion mutants could be responsible for the observed discrepancy between the knockout strains and challenge the role of MK5/PRAK in p53-dependent tumor suppression. Further MK5/PRAK knockout and knock-in mouse strains will be necessary to assign a physiological function to MK5/PRAK in this model organism.

Genetic variants in nicotinic acetylcholine receptor and alcohol metabolism genes have been associated with propensity to smoke tobacco and drink alcohol, respectively, and also implicated in genetic susceptibility to head and neck cancer. In addition to smoking and alcohol, tobacco chewing is an important oral cancer risk factor in India. It is not known if these genetic variants influence propensity or oral cancer susceptibility in the context of this distinct etiology.We examined 639 oral and pharyngeal cancer cases and 791 controls from two case-control studies conducted in India. We investigated six variants known to influence nicotine addiction or alcohol metabolism, including rs16969968 (CHRNA5), rs578776 (CHRNA3), rs1229984 (ADH1B), rs698 (ADH1C), rs1573496 (ADH7), and rs4767364 (ALDH2).The CHRN variants were associated with the number of chewing events per day, including in those who chewed tobacco but never smoked (P = 0.003, P = 0.01 for rs16969968 and rs578776 respectively). Presence of the variant allele contributed to approximately 13% difference in chewing frequency compared to non-carriers. While no association was observed between rs16969968 and oral cancer risk (OR = 1.01, 95% CI = 0.83- 1.22), rs578776 was modestly associated with a 16% decreased risk of oral cancer (OR = 0.84, 95% CI = 0.72- 0.98). There was little evidence for association between polymorphisms in genes encoding alcohol metabolism and oral cancer in this population.The association between rs16969968 and number of chewing events implies that the effect on smoking propensity conferred by this gene variant extends to the use of smokeless tobacco.

Keratins are structural marker proteins with tissue specific expression; however, recent reports indicate their involvement in cancer progression. Previous study from our lab revealed deregulation of many genes related to structural molecular integrity including KRT76. Here we evaluate the role of KRT76 downregulation in oral precancer and cancer development.We evaluated KRT76 expression by qRT-PCR in normal and tumor tissues of the oral cavity. We also analyzed K76 expression by immunohistochemistry in normal, oral precancerous lesion (OPL), oral squamous cell carcinoma (OSCC) and in hamster model of oral carcinogenesis. Further, functional implication of KRT76 loss was confirmed using KRT76-knockout (KO) mice.We observed a strong association of reduced K76 expression with increased risk of OPL and OSCC development. The buccal epithelium of DMBA treated hamsters showed a similar trend. Oral cavity of KRT76-KO mice showed preneoplastic changes in the gingivobuccal epithelium while no pathological changes were observed in KRT76 negative tissues such as tongue.The present study demonstrates loss of KRT76 in oral carcinogenesis. The KRT76-KO mice data underlines the potential of KRT76 being an early event although this loss is not sufficient to drive the development of oral cancers. Thus, future studies to investigate the contributing role of KRT76 in light of other tumor driving events are warranted.

Identifying oral cancer lesions associated with high risk of relapse and predicting clinical outcome remain challenging questions in clinical practice. Genomic alterations may add prognostic information and indicate biological aggressiveness thereby emphasizing the need for genome-wide profiling of oral cancers. High-resolution array comparative genomic hybridization was performed to delineate the genomic alterations in clinically annotated primary gingivo-buccal complex and tongue cancers (n = 60). The specific genomic alterations so identified were evaluated for their potential clinical relevance. Copy-number changes were observed on chromosomal arms with most frequent gains on 3q (60%), 5p (50%), 7p (50%), 8q (73%), 11q13 (47%), 14q11.2 (47%), and 19p13.3 (58%) and losses on 3p14.2 (55%) and 8p (83%). Univariate statistical analysis with correction for multiple testing revealed chromosomal gain of region 11q22.1-q22.2 and losses of 17p13.3 and 11q23-q25 to be associated with loco-regional recurrence (P = 0.004, P = 0.003, and P = 0.0003) and shorter survival (P = 0.009, P = 0.003, and P 0.0001) respectively. The gain of 11q22 and loss of 11q23-q25 were validated by interphase fluorescent in situ hybridization (I-FISH). This study identifies a tractable number of genomic alterations with few underlying genes that may potentially be utilized as biological markers for prognosis and treatment decisions in oral cancers.

SB202190, a widely used inhibitor of p38 MAPKα and β, was recently described to induce autophagic vacuoles and cell death in colon and ovarian cancer cells lines and, therefore, this effect was supposed to be specific for transformed cells and to open therapeutic options. Here, we demonstrate that SB202190 and the structurally related inhibitor SB203580 induce pro-autophagic gene expression and vacuole formation in various cancer and non-cancer cell lines of human, rat, mouse and hamster origin. This effect seems to induce defective autophagy leading to the accumulation of acidic vacuoles, p62 protein and lipid conjugated LC3. Using further p38 inhibitors we show that p38 MAPK inhibition is not sufficient for the autophagic response. In line with these results, expression of a SB202190-resistant mutant of p38α, which significantly increases activity of the p38 pathway under inhibitory conditions, does not block SB202190-dependent vacuole formation, indicating that lack of p38α activity is not necessary for this effect. Obviously, the induction of autophagic vacuole formation by SB203580 and SB202190 is due to off-target effects of these inhibitors on post-translational protein modifications, such as phosphorylation of the MAPKs ERK1/2 and JNK1/2, ribosomal protein S6, and PKB/Akt. Interestingly, the PI3K-inhibitor wortmannin induces transient vacuole formation indicating that the PI3K-PKB/Akt-mTOR pathway is essential for preventing autophagy and that cross-inhibition of this pathway by SB202190 could be the reason for the early part of the effect observed.