TP53 Pathway Research Articles

Targeted sequencing of refractory myeloma reveals a high incidence of mutations in CRBN and Ras pathway genes

AbstractIn this study, targeted sequencing to screen 50 multidrug refractory multiple myeloma (rMM) patients was performed by using the Multiple Myeloma Mutation Panel. Patients were pretreated with both immunomodulatory drugs (IMiDs) and proteasome inhibitors (PIs), and 88%, 78%, and 68% were refractory to an IMiD, a PI, or both, respectively. The majority of patients had progressive (82%) or refractory (78%) disease immediately before sampling, with 43% being IMiD refractory and 46% being PI refractory in the most recent line of therapy. Compared with newly diagnosed MM, an increased prevalence of mutations in the Ras pathway genes KRAS, NRAS, and/or BRAF (72%), as well as TP53 (26%), CRBN (12%), and CRBN pathway genes (10%) was observed. Longitudinal analyses performed in 3 patients with CRBN mutations at time of IMiD resistance confirmed that these mutations were undetectable at earlier, IMiD-sensitive time points. Furthermore, the functional introduction of these mutations in MM cells conferred lenalidomide resistance in vitro. These data indicate a differential genetic landscape in rMM associated with drug response.

Gene-gene interactions in gastrointestinal cancer susceptibility.

Cancer arises from complex, multi-layer interactions between diverse genetic and environmental factors. Genetic studies have identified multiple loci associated with tumor susceptibility. However, little is known about how germline polymorphisms interact with one another and with somatic mutations within a tumor to mediate acquisition of cancer traits. Here, we survey recent studies showing gene-gene interactions, also known as epistases, affecting genetic susceptibility in colorectal, gastric and esophageal cancers. We also catalog epistasis types and cancer hallmarks with respect to the interacting genes. A total of 22 gene variation pairs displayed all levels of statistical epistasis, including synergistic, redundant, suppressive and co-suppressive interactions. Five genes primarily involved in base excision repair formed a linear topology in the interaction network, MUTYH-OGG1-XRCC1-PARP1-MMP2, and three genes in mTOR cell-proliferation pathway formed another linear network, PRKAG2-RPS6KB1-PIK3CA. Discrete pairwise epistasis was also found in nucleotide excision repair, detoxification, proliferation, TP53, TGF-β and other pathways. We propose that three modes of biological interaction underlie the molecular mechanisms for statistical epistasis. The direct binding, linear pathway and convergence modes can exhibit any level of statistical epistasis in susceptibility to gastrointestinal cancers, and this is likely true for other complex diseases as well. This review highlights the link between cancer hallmarks and susceptibility genes.

Abstract 4765: Impact of TP53 status and functional classification on molecular profiles in breast cancer subtypes

Abstract Breast cancer is a multifaceted disease with several clinical, pathological and molecular attributes contributing to disease prognosis or treatment outcome. Treatment measures in breast cancer are based on hormone/growth factor receptor -estrogen/progesterone receptor (ER/PR) or human epidermal growth factor receptor 2 (Her2) status. TP53 pathway inactivation in breast cancer is well-established. Although TP53's therapeutic relevance is well-recognized, it remains under-utilized in patient-management, since all TP53 mutants are treated equally in the diagnostic context. In reality, enormous heterogeneity exists in nature, type and functional impact of TP53 variants. Therefore, understanding the diversity of TP53 variants in breast cancer subtypes may enhance its diagnostic utility in this cancer. We utilized clinical NGS data, obtained using commercially available targeted panels, TruSeq Amplicon Cancer Panel (Illumina) and Ion AmpliSeq Cancer Hotspot Panel v2 (Thermofisher) to analyze tumor DNAs from cancer patients at the Advanced Molecular Diagnostic Laboratory (Princess Margaret Cancer Centre, Toronto, Canada). We focused on data from 105 advanced breast cancer patients. We consolidated several schemes proposed in the literature to classify TP53 variants, and evaluated patient molecular profiling and pathology data based on: (1) presence of TP53 variants; (b) coding effect; and (c) transcriptional activity. We further investigated whether TP53 variants were associated with reportable variant load, co-occurrence with other molecular changes and hormone/growth-factor receptor status. In our study group, 70.4% cases carried one or more variants. TP53 alterations were prevalent (40.9%) in our cohort, followed by PIK3CA variants (36.2%). 15/105 cases (14.3%) carried variants in both genes. Unlike in other cancer types, where missense TP53 variants predominate (e.g., colorectal, 72.6%), missense (49%) and nonsense/frameshift (42%) variants were similarly distributed in breast cancers. Gain-of-Function (GOF) and Loss-of-Function (LOF) TP53 variants were also equally distributed (32% vs. 33%). However, TP53mut PIK3CAmut breast cancer cases were more likely to carry missense and/or LOF variants (10/15 cases). TP53 variants were also associated with hormone/growth-factor receptor status. A greater proportion of ER- vs ER+, PR- vs PR+, and ER-PR-Her2- vs ER+PR+Her2- breast cancer cases carried missense GOF TP53 variants respectively when compared to missense LOF and variants of unknown significance taken together(80-85% vs 50-55%; p<0.0001). Finally, TP53mut cases were more likely to carry multiple variants in contrast to TP53wt cases (37.2% vs. 12.9%). Taken together, we define a stratification strategy for TP53 that takes into account the diversity of TP53 variants, and demonstrate its application to molecular profiling and clinico-pathological data in breast cancer. Citation Format: Swati Garg, Mahadeo A. Sukhai, Maksym Misyura, Mariam Thomas, Tong Zhang, Lillian L. Siu, Philippe L. Bedard, Tracy L. Stockley, Suzanne Kamel-Reid. Impact of TP53 status and functional classification on molecular profiles in breast cancer subtypes. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4765.

Abstract 4173: The oncopig cancer model as a validated model for human hepatocellular carcinoma

Abstract Human hepatocellular carcinoma (HCC) is the fifth most common cancer globally and is the third most common cause of cancer-related deaths worldwide, accounting for more than 600,000 deaths each year. While the information gained from human clinical trials may enhance the efficacy of HCC treatment, such trials provide limited capability for investigation into the processes contributing to procedural effectiveness and disease progression. Immunological, genetic and physiological differences between rodents and humans represent major limitations for the use of rodent based cancer models in HCC studies. Pigs represent an attractive model given that they share many genetic, anatomical, and physiological similarities with humans. In a previous study, we demonstrated the functionality of a porcine inducible model of cancer (Oncopig; Schook et. al., 2015). In order to develop and validate the Oncopig HCC model, we established primary (control) hepatocyte cultures from 3 individual Oncopigs and transformed them with Cre recombinase (HepCre). Within a week the activated cells began to proliferate, taking on transformed morphologic characteristics. Transcriptional analysis confirmed both the primary and transformed cells expressed hepatocyte specific marker genes (albumin, HNF4 and G6P). The Cre treated cell lines also expressed oncogenic P53R167H and KrasG12D. Each of the three transformed cell lines were then injected into the livers of SCID mice to verify tumorigenicity. SCID mice (9/9) developed tumors that recapitulated several morphological similarities with human HCC including cellular arrangement and marked pleomorphism. The primary hepatocytes expressed cytokeratin and the HepCre cell lines expressed both cytokeratin and vimentin, indicative of an epithelial-mesenchymal transition, which is a common tumor progression signal observed in human HCC and other highly proliferative carcinomas. Alpha-fetoprotein is often found in the serum of HCC patients and was measurable in the medium of porcine HepCre cells but not primary hepatocytes. Oncopig HCC cells were polygonal with eosinophilic granular cytoplasm as observed in human HCC. Principal component analysis of gene expression data showed clear separation between the primary hepatocytes and HepCre cells. Differential expression analysis revealed upregulation of proto-oncogenes and downregulation of tumor supressor genes in the HepCre cells, as well as enrichment of several pathways that are concordantly dysregulated in human HCC, including the TP53 pathway, fatty acid metabolism pathway and complement and coagulation cascade pathway. These data demonstrate the utility of the Oncopig model in terms of relevance to human HCC and therefore aid in improved detection, treatment, biomarker discovery for liver tumors and other unmet clinical needs. Reference Schook et al. A Genetic Porcine Model of Cancer. PLoS One. 2015;10: e0128864 Citation Format: Kwame A. Darfour-Oduro, Arun K. De, Laurie A. Rund, Ron C. Gaba, Charles E. Ray, Regina M. Schwind, Kyle M. Schachtschneider, Kuldeep B. Singh, Lawrence B. Schook. The oncopig cancer model as a validated model for human hepatocellular carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4173.

Abstract 3170: Analysis of copy number variations in meningioma samples using microarrays revealed 22q deletions more frequent in higher grade tumors

Abstract Background: Meningiomas represent one of the most common intracranial tumors. They are generally thought to progress from low to high-grade lesions. However, the molecular mechanisms underlying their pathogenesis remain still uncertain. We suppose the existence of a correlation between the parameters that will help to predict more precisely their biological behavior and response to therapy. Identification of meningioma molecular subgroups may have significant potential to improve clinical management, through molecular disease risk stratification strategies and the identification of patients who could benefit from targeted molecular therapeutics. Methods and patients: Formalin-fixed paraffin embedded tumor samples were obtained from 30 patients (10 patients in each meningioma grade) and 5 healthy controls (dura mater). Comprehensive clinical-pathological data were mined. There were 7 males and 23 females; mean age was 56.7 years, range 33 - 100 years. Total DNA was purified from FFPE samples after pathological verification using proteinase K treatment followed by QIAmp DNA FFPE Tissue Kit (Qiagen). Microarray analysis was performed using the OncoScan FFPE Assay Kit (Affymetrix), raw data were obtained using Chromosome Analysis Suite (Affymetrix) in default manner. Subsequently, the data were analyzed using Nexus Express software (Biodiscovery). 10 samples were excluded from analyses because of poor quality of results. Results: In total, 35 OncoScan arrays were performed for copy number variants (CNV) analyses in meningioma and control samples. Our results confirm that del(22q) and del(1p) are the most common (44%, resp. 24% of cases) deletions in meningiomas. Del(22q) was present in 75% grade 2 menigiomas cases in contrast to 25% in grade 1 patients. Additionally, monosomy of chromosome 6 (12%), 8 (8%), 14 (20%) and 18 (12%) were observed. Surprisingly, chromosomal gains and LOH were found only in small portion of cases (8%). Finally, mutation in TP53 (c.817C>T/A, c.637C>T) and PIK3CA (c.3140A>G) genes were found in 12% of cases. Only common CNV variants were found in healthy control samples. Conclusion: We have identified the CNV profiles in meningioma patients allowing for better knowledge of pathological pathways and tumor progression. Del(22q) is frequently present in higher grade tumors probably altering NF2 a TP53 pathways. However, it will require further validation. Acknowledgment: This work was financially supported by Ministry of Health of the Czech Republic, grant nr. 15-29021A, IGA UP LF 2015_010 and NPU LO1304. Citation Format: Josef Srovnal, Vladimir Balik, Jiri Ehrmann, Miroslav Vaverka, Lumir Hrabalek, Katerina Staffova, Marian Hajduch. Analysis of copy number variations in meningioma samples using microarrays revealed 22q deletions more frequent in higher grade tumors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3170.

Genetic Variants in the p14ARF/MDM2/TP53 Pathway Are Associated with the Prognosis of Esophageal Squamous Cell Carcinoma Patients Treated with Radical Resection.

The p14ARF/MDM2/ TP53 pathway is known to play an important role in tumor progression by cell cycle control, although the association between this pathway and the prognosis of esophageal squamous cell carcinoma (ESCC) is unclear. In this study, we explored the association between genetic variants in the p14ARF/MDM2/TP53 pathway and prognosis in ESCC patients with radical resection. 124 ESCC patients with radical resection were included in this retrospective study and genotyped using the MassArray method. According to multivariate Cox hazard analysis and multiple testing, the TC/CC genotype of p14ARF rs3814960 was shown to be strongly related to a decreased overall survival (OS) (HR = 2.77, 95% CI: 1.33–5.75, P = 0.006, Pc = 0.030) and disease-free survival (DFS) (HR = 2.45, 95% CI: 1.30–4.61, P = 0.005, Pc = 0.025). Moreover, patients with the DEL/A +AA genotype of MDM2 rs34886328 had a notably increased OS (HR = 0.27, 95% CI: 0.13–0.56, P = 4.7×10−4, Pc = 0.003) and DFS (HR = 0.22, 95% CI: 0.11–0.43, P = 1.1×10−5, Pc = 6.6×10−5). We also found that these two SNPs had a cumulative effect on the prognosis of ESCC, with the OS (P < 0.001) and DFS (P < 0.001) being shortest for patients carrying both of these unfavorable genotypes. In conclusion, genetic variants of the p14ARF/MDM2/TP53 pathway are significantly related to OS and DFS, and may be predictors of the prognosis of ESCC after surgery. We speculate the individuals with the TC/CC genotype of p14ARF rs3814960 and/or the DEL/DEL genotype of MDMD2 rs34886328 should have more aggressive treatment and may greatly benefit from early prediction and prevention of an unfavorable prognosis by genotyping before the initiation of therapy. These findings should be further validated in a larger population.

A novel TP53 pathway influences the HGS-mediated exosome formation in colorectal cancer.

Tumor-derived exosomes are important for cell-cell communication. However, the role of TP53 in the control of exosome production in colorectal cancer (CRC) is controversial and unclear. The features of exosomes secreted from HCT116 TP53-wild type (WT), TP53-knockout (KO) and constructed TP53 (R273H)-mutant (MT) cells were assessed. The exosomes from the MT and KO cells exhibited significantly reduced sizes compared with the WT cells. A comprehensive proteomic analysis of exosomal proteins was performed using the isobaric tag for relative and absolute quantitation (iTRAQ)-2D-LC-MS/MS strategy. A total of 3437 protein groups with ≥2 matched peptides were identified. Specifically, hepatocyte growth factor-regulated tyrosine kinase substrate (HGS) was consistently down-regulated in the exosomes from the MT and KO cells. Functional studies demonstrated that low HGS levels were responsible for the decreased exosome size. TP53 regulated HGS expression and thus HGS-dependent exosome formation. Furthermore, the HGS expression was gradually increased concomitant with CRC carcinogenesis and was an independent poor prognostic factor. In conclusion, a novel HGS-dependent TP53 mechanism in exosome formation was identified in CRC. HGS may serve as a novel prognostic biomarker and a candidate target for therapeutic interventions.

A novel TRIM family member, Trim69, regulates zebrafish development through p53-mediated apoptosis.

Trim69 contains the hallmark domains of a tripartite motif (TRIM) protein, including a Ring-finger domain, B-box domain, and coiled-coil domain. Trim69 is structurally and evolutionarily conserved in zebrafish, mouse, rat, human, and chimpanzee. The role of this protein is unclear, however, so we investigated its function in zebrafish development. Trim69 is extensively expressed in zebrafish adults and developing embryos-particularly in the testis, brain, ovary, and heart-and its expression decreases in a time- and stage-dependent manner. Loss of trim69 in zebrafish induces apoptosis and activates apoptosis-related processes; indeed, the tp53 pathway was up-regulated in response to the knockdown. Expression of human trim69 rescued the apoptotic phenotype, while overexpression of trim69 does not increase cellular apoptosis. Taken together, our results suggest that trim69 participates in tp53-mediated apoptosis during zebrafish development. Mol. Reprod. Dev. 83: 442-454, 2016. © 2016 Wiley Periodicals, Inc.

Tumor-Suppressor Functions of the TP53 Pathway.

The fundamental biological importance of the Tp53 gene family is highlighted by its evolutionary conservation for more than one billion years dating back to the earliest multicellular organisms. The TP53 protein provides essential functions in the cellular response to diverse stresses and safeguards maintenance of genomic integrity, and this is manifest in its critical role in tumor suppression. The importance of Tp53 in tumor prevention is exemplified in human cancer where it is the most frequently detected genetic alteration. This is confirmed in animal models, in which a defective Tp53 gene leads inexorably to cancer development, whereas reinstatement of TP53 function results in regression of established tumors that had been initiated by loss of TP53. Remarkably, despite extensive investigation, the specific mechanisms by which TP53 acts as a tumor suppressor are yet to be fully defined. We review the history and current standing of efforts to understand these mechanisms and how they complement each other in tumor suppression.

Increased mitochondrial fission promotes autophagy and hepatocellular carcinoma cell survival through the ROS-modulated coordinated regulation of the NFKB and TP53 pathways

ABSTRACTMitochondrial morphology is dynamically remodeled by fusion and fission in cells, and dysregulation of this process is closely implicated in tumorigenesis. However, the mechanism by which mitochondrial dynamics influence cancer cell survival is considerably less clear, especially in hepatocellular carcinoma (HCC). In this study, we systematically investigated the alteration of mitochondrial dynamics and its functional role in the regulation of autophagy and HCC cell survival. Furthermore, the underlying molecular mechanisms and therapeutic application were explored in depth. Mitochondrial fission was frequently upregulated in HCC tissues mainly due to an elevated expression ratio of DNM1L to MFN1, which significantly contributed to poor prognosis of HCC patients. Increased mitochondrial fission by forced expression of DNM1L or knockdown of MFN1 promoted the survival of HCC cells both in vitro and in vivo mainly by facilitating autophagy and inhibiting mitochondria-dependent apoptosis. We further demonstrated that the survival-promoting role of increased mitochondrial fission was mediated via elevated ROS production and subsequent activation of AKT, which facilitated MDM2-mediated TP53 degradation, and NFKBIA- and IKK-mediated transcriptional activity of NFKB in HCC cells. Also, a crosstalk between TP53 and NFKB pathways was involved in the regulation of mitochondrial fission-mediated cell survival. Moreover, treatment with mitochondrial division inhibitor-1 significantly suppressed tumor growth in an in vivo xenograft nude mice model. Our findings demonstrate that increased mitochondrial fission plays a critical role in regulation of HCC cell survival, which provides a strong evidence for this process as drug target in HCC treatment.

Characterization of TP53 and PI3K signaling pathways as molecular targets in gynecologic malignancies.

Recent developments in genomic analysis have unveiled the key signaling pathways in human cancer. However, only a limited number of molecular-targeted drugs are applicable for clinical use in gynecologic malignancies. TP53 signaling and phosphatidylinositol 3 kinase pathways are frequently mutated in cancer, and have received much attention as molecular targets in human cancers. In this review, we mainly focus on the functions of these pathways, and discuss the molecular-targeted drugs under clinical trials. The molecular-targeted drugs described in this review include dual phosphatidylinositol 3 kinase/mTOR inhibitors (NVP-BEZ235, DS-7423, SAR245409), an mTOR inhibitor (everolimus), an MEK inhibitor (pimasertib), an autophagy inhibitor (chloroquine), a cyclin-dependent kinases 4/6 inhibitor (PD0332991), and a poly (ADP-ribose) polymerase inhibitor (olaparib).

Pathway Implications of Aberrant Global Methylation in Adrenocortical Cancer.

ContextAdrenocortical carcinomas (ACC) are a rare tumor type with a poor five-year survival rate and limited treatment options.ObjectiveUnderstanding of the molecular pathogenesis of this disease has been aided by genomic analyses highlighting alterations in TP53, WNT, and IGF signaling pathways. Further elucidation is needed to reveal therapeutically actionable targets in ACC.DesignIn this study, global DNA methylation levels were assessed by the Infinium HumanMethylation450 BeadChip Array on 18 ACC tumors and 6 normal adrenal tissues. A new, non-linear correlation approach, the discretization method, assessed the relationship between DNA methylation/gene expression across ACC tumors.ResultsThis correlation analysis revealed epigenetic regulation of genes known to modulate TP53, WNT, and IGF signaling, as well as silencing of the tumor suppressor MARCKS, previously unreported in ACC.ConclusionsDNA methylation may regulate genes known to play a role in ACC pathogenesis as well as known tumor suppressors.

Transglutaminase 2 contributes to a TP53-induced autophagy program to prevent oncogenic transformation

Genetic alterations which impair the function of the TP53 signaling pathway in TP53 wild-type human tumors remain elusive. To identify new components of this pathway, we performed a screen for genes whose loss-of-function debilitated TP53 signaling and enabled oncogenic transformation of human mammary epithelial cells. We identified transglutaminase 2 (TGM2) as a putative tumor suppressor in the TP53 pathway. TGM2 suppressed colony formation in soft agar and tumor formation in a xenograft mouse model. The depletion of growth supplements induced both TGM2 expression and autophagy in a TP53-dependent manner, and TGM2 promoted autophagic flux by enhancing autophagic protein degradation and autolysosome clearance. Reduced expression of both CDKN1A, which regulates the cell cycle downstream of TP53, and TGM2 synergized to promote oncogenic transformation. Our findings suggest that TGM2-mediated autophagy and CDKN1A-mediated cell cycle arrest are two important barriers in the TP53 pathway that prevent oncogenic transformation.

Abstract B49: The role of KCNK9 and TP53 on the racial disparity in biologically aggressive breast cancer subtype in The Cancer Genome Atlas (TCGA)

Abstract Introduction: Compared to non-Hispanic (nH) white, nH black women are more likely to present with biologically aggressive estrogen and progesterone receptor (ER/PR)-negative (ERN) and triple negative (TN) breast cancer subtypes which are associated with worse prognosis (black:white disparity). These subtypes also exhibit greater genetic instability, with TP53 being the single most recurrently mutated gene. KCNK9 is a maternally imprinted and functionally mono-allelic 2-pore domain potassium channel proto-oncogene located at chromosomal region 8q24.3, encoding TASK-3. KCNK9 overexpression may arise via loss of methylation (LOM) or copy number amplification (CNA). We examined KCNK9 and TP53 as potential mediators of the black:white racial disparity in aggressive breast cancer subtypes. Methods: Using publically available TCGA data on 1040 women with invasive breast cancer, we examined: associations of KCNK9 LOM (HM450 beta values; highest third loss vs. others), amplification (putative CNA from GISTIC; any vs. neutral), and mRNA overexpression (RNA-Seq from RSEM; z-score &amp;gt; +1 SD vs. others) with ERN and TN subtype, and overall survival (OS) and disease-free survival (DFS). General TP53 pathway function was examined using mRNA expression z-scores for six related genes (TP53 MDM2 MDM4 CDKN2A CDKN2B TP53BP1). Prevalence risk ratios (RRs) with 95% confidence intervals (CIs) were estimated via logistic regression with model-based standardization (predictive margins) and hazard ratios (HRs) using Cox proportional hazards models, and all models were adjusted at baseline for age as a continuous variable and stage as categorical variable. In mediation analysis, we estimated the proportionate reduction in the racial disparity in aggressive subtypes after accounting for differences in KCNK9 and TP53 pathway biomarkers by comparison of rescaled coefficients using the method of Karlson, Holm and Breen in adjusted models. Death from any cause was considered an event for all-cause mortality (i.e. OS), and recurrence or progression of disease was considered an event for DFS. Results: KCNK9 LOM, amplification and overexpression were each significantly associated with ERN and TN subtype and black race (all p&amp;lt;0.05). In mutually adjusted models including TP53 pathway markers and race, KCNK9 status significantly predicted outcome for both subtypes suggesting the importance of KCNK9 as a molecular driver of aggressive subtype (ERN: LOM RR, 3.69; 95% CI, 2.28-5.98; p&amp;lt;0.001. Amplification RR, 1.40; 95% CI, 0.98-1.99; p=0.064. Overexpression RR, 1.33; 95% CI, 1.07-1.67; p=0.011) (TN: LOM RR, 3.37; 95% CI, 1.99-5.70; p&amp;lt;0.001. Amplification RR, 1.82; 95% CI, 1.12-2.95; p=0.016. Overexpression RR, 1.41; 95% CI, 1.11-1.80; p=0.005). In baseline adjusted Cox models, amplification and overexpression were associated with greater OS, and overexpression was marginally associated with DFS. However, in mutually adjusted Cox models including subtype and TP53 pathway markers, no KCNK9 markers remained independent predictors of mortality or recurrence; however, TN subtype and CDKN2A overexpression remained significant predictors for both (TN: OS HR, 8.96; 95% CI, 2.65-30.28; p&amp;lt;0.001. DFS HR, 8.58; 95% CI, 2.24-32.8; p=0.002) (CDKN2A: OS HR, 0.29; 95% CI, 0.09-0.97; p=0.045. DFS HR, 0.21; 95% CI, 0.06-0.76; p=0.017). ERN and TN subtypes were more common among black patients (black:white disparity: ERN RR, 2.16; 95% CI, 1.6-2.8; p&amp;lt;0.001. TN RR, 2.54; 95% CI, 1.8-3.53; p&amp;lt;0.001). The racial disparity in ERN and TN subtypes was mediated by molecular pathways as follows: KCNK9 variables accounted for about one-third, TP53 pathway variables accounted for one-half, and together they accounted for more than two-thirds of the disparity (all p&amp;lt;0.05). Conclusions: KCNK9 appears to be a molecular driver of aggressive breast tumor subtype, and in combination with aberrant TP53 pathway overexpression, largely accounts for observed black:white disparity in more biologically aggressive breast cancer subtypes. Citation Format: Keith A. Dookeran, Jacob K. Kresovich, Maria Argos, Garth H. Rauscher. The role of KCNK9 and TP53 on the racial disparity in biologically aggressive breast cancer subtype in The Cancer Genome Atlas (TCGA). [abstract]. In: Proceedings of the Eighth AACR Conference on The Science of Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; Nov 13-16, 2015; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2016;25(3 Suppl):Abstract nr B49.

Abstract IA26: Oncogenic mechanisms in diffuse intrinsic pontine gliomas

Abstract Diffuse intrinsic pontine glioma (DIPG) is a devastating disease with no effective cures. DIPG arises in the brainstem and is found almost exclusively in children. Mutations in canonical cancer signalling pathways including the RTK/RAS/PI3K pathway, the Tp53 pathway, and the CDK4/6 G1 checkpoint occur in DIPGs, targeting different effectors within these pathways and resulting in significant genetic heterogeneity between tumors. However, genome-wide studies provided abundant evidence that unique selective pressures drive HGG in children compared to adults, identifying novel oncogenic mutations connecting tumorigenesis and chromatin regulation as well as developmental signaling pathways. Recurrent mutations of the BMP receptor ACVR1/ALK2 are found in 25-30% of DIPGs, and are further restricted to the youngest patients, highlighting critical connections between development and gliomagenesis. The molecular hallmark of DIPGs is frequent recurrent somatic mutation in histone H3. Mutually exclusive somatic heterozygous H3F3A or HIST1H3B mutations encoding a K27M variant of histone H3.3 or H3.1, respectively, are found in nearly 80 percent of DIPGs. These recurrent mutations provide clear evidence of an important role for chromatin regulation in DIPG disease pathogenesis, and present a unifying molecular mechanism driving DIPG. We have evaluated the contribution of H3.3 K27M mutation to DIPG tumor growth in vitro to gain insights into molecular mechanisms of oncogenic activity, and to assess the potential of histone H3 mutation as a therapeutic sensitivity in this incurable brain tumor. Citation Format: Andre B. Silveira, Lawryn H. Kasper, Jon D. Larson, Xiaoyan Zhu, Alexander K. Diaz, Suzanne J. Baker. Oncogenic mechanisms in diffuse intrinsic pontine gliomas. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Pediatric Cancer Research: From Mechanisms and Models to Treatment and Survivorship; 2015 Nov 9-12; Fort Lauderdale, FL. Philadelphia (PA): AACR; Cancer Res 2016;76(5 Suppl):Abstract nr IA26.

Understanding the Osteosarcoma Pathobiology: A Comparative Oncology Approach.

Osteosarcoma is an aggressive primary bone tumor in humans and is among the most common cancer afflicting dogs. Despite surgical advancements and intensification of chemo- and targeted therapies, the survival outcome for osteosarcoma patients is, as of yet, suboptimal. The presence of metastatic disease at diagnosis or its recurrence after initial therapy is a major factor for the poor outcomes. It is thought that most human and canine patients have at least microscopic metastatic lesions at diagnosis. Osteosarcoma in dogs occurs naturally with greater frequency and shares many biological and clinical similarities with osteosarcoma in humans. From a genetic perspective, osteosarcoma in both humans and dogs is characterized by complex karyotypes with highly variable structural and numerical chromosomal aberrations. Similar molecular abnormalities have been observed in human and canine osteosarcoma. For instance, loss of TP53 and RB regulated pathways are common. While there are several oncogenes that are commonly amplified in both humans and dogs, such as MYC and RAS, no commonly activated proto-oncogene has been identified that could form the basis for targeted therapies. It remains possible that recurrent aberrant gene expression changes due to gene amplification or epigenetic alterations could be uncovered and these could be used for developing new, targeted therapies. However, the remarkably high genomic complexity of osteosarcoma has precluded their definitive identification. Several advantageous murine models of osteosarcoma have been generated. These include spontaneous and genetically engineered mouse models, including a model based on forward genetics and transposon mutagenesis allowing new genes and genetic pathways to be implicated in osteosarcoma development. The proposition of this review is that careful comparative genomic studies between human, canine and mouse models of osteosarcoma may help identify commonly affected and targetable pathways for alternative therapies for osteosarcoma patients. Translational research may be found through a path that begins in mouse models, and then moves through canine patients, and then human patients.

Actionable targets in patients with cisplatin-resistant advanced germ cell tumors.

473 Background: Approximately 30% of patients with advanced germ cell tumor (aGCT) will progress after first-line chemotherapy. Nearly half of these patients will die of progressive GCT. We describe potentially actionable mutations in a cohort of patients with platinum-resistant aGCT through targeted sequencing. Methods: 76 patients with cisplatin-resistant (CR) disease were sequenced using the Memorial Sloan Kettering Integrated Mutation Profiling of Actionable Cancer Targets (MSK-IMPACT) assay that examines 341 cancer-related genes. Patients were categorized as CR if they met any of the following criteria: 1) incomplete response to first-line cisplatin-based chemotherapy; 2) nonteratomatous tumor progression after standard chemotherapy; 3) nonteratomatous GCT identified at postchemo surgery. We grouped all somatic mutations into core signal transduction pathways or canonical cell functions to identify potential precise targets for therapy. Results: The majority of patients had nonseminoma histology (n = 64, 84%). International Germ Cell Cancer Collaborative Group risk was good, intermediate, and poor in 34%, 13%, and 53% of patients. 17 patients died of disease. In total, 51 potentially actionable alterations were identified in 36/76 (47%) patients. In the TP53 pathway, 7 MDM2 amplifications and 4 MYCN amplifications that may sensitize to nutlin-3 inhibitors were identified. Within the receptor tyrosine kinase pathway, 3 KIT mutations, 1 KDR amplification, and 1 MET amplification were seen that may sensitize to tyrosine kinase inhibitors. Eleven KRAS mutations, 3 NRAS mutations, 3 BRAF mutations, and 2 RAC1 mutations were see among the RAS pathway with preclinical data suggesting efficacy towards respective inhibitors. Actionable targets were also among the PI3-K, WNT, and cell cycle pathways. Potential targets with chromatin modifying or tumor suppressor functions were also seen. Conclusions: We describe actionable alterations that may guide treatment selection in a significant proportion of patients with CR aGCT. Targeted sequencing of these patients may allow us to enrich future clinical trials with patients whose tumors harbor alterations in the drug target of interest.

Frequent somatic mutations of chromatin remodeling genes in metastatic cutaneous squamous cell carcinoma

Exome and targeted sequencing studies have identified potential driver mutations for a variety of tumor types. Cutaneous squamous cell carcinoma (cSCC) is one of the most highly mutated cancers but is typically correlated with high survival rates and low rates of metastasis. Nevertheless, metastatic cSCC is a significant health threat; up to 8800 individuals are estimated to die yearly from this disease. As it is difficult to predict which cSCCs are more likely to metastasize, and because there are no targeted therapies specifically designated for metastatic cSCC, we performed exome and targeted sequencing of 18 metastatic and 10 primary cSCCs to identify mutations with potential therapeutic benefit. Genes previously shown to be mutated in primary cSCC as well as aggressive tumors such as TP53 and NOTCH pathway genes were mutated at high rates in both metastatic and primary tumors. We compared our results to published sequencing results of an additional 223 primary tumors and 68 aggressive cSCCs. We identified several genes showing higher mutation frequencies in metastatic cSCC relative to primary tumors including the chromatin remodeling gene KMT2D, AKT/PI3K pathway gene PIK3CG, and the classic skin tumor suppressor TP53. These studies uncover potential pathways important in metastatic cSCC that may lead to new therapeutic strategies and understanding of the biology leading to more aggressive tumor behavior.

MicroRNAs as Therapeutic Targets and Colorectal Cancer Therapeutics.

The diagnosis and treatment of colorectal cancer (CRC) have improved greatly over recent years; however, CRC is still one of the most common cancers and a major cause of cancer death worldwide. Several recently developed drugs and treatment strategies are currently in clinical trials; however, there is still a compelling need for novel, highly efficacious therapies. MicroRNAs (miRNAs) are short non-coding RNAs consisting of 20-25 nucleotides that regulate post-transcriptional gene expression by binding to the 3'-untranslated region of mRNAs. miRNAs are known to regulate cancer pathways and to be expressed aberrantly in cancer. Since their initial discovery, a large number of miRNAs have been identified as oncogenes, whereas others function as tumor suppressors. Furthermore, signaling pathways that are important in CRC (e.g. the WNT, MAPK, TGF-β, TP53 and PI3K pathways) are regulated by miRNAs. A single miRNA can simultaneously regulate several target genes and pathways, indicating the therapeutic potential of miRNAs in CRC. However, significant obstacles remain to be overcome, such as an efficient miRNA delivery system, and the assessment of safety and side effects. Thus, miRNA therapy is still developing and possesses great potential for the treatment of CRC. In this chapter, we focus on miRNAs related to CRC and summarize previous studies that emphasize the therapeutic aspects of miRNAs in CRC.

Renal Cell Carcinomas in Vinylidene Chloride-exposed Male B6C3F1 Mice Are Characterized by Oxidative Stress and TP53 Pathway Dysregulation.

Vinylidene chloride (VDC) has been widely used in the production of plastics and flame retardants. Exposure of B6C3F1 mice to VDC in the 2-year National Toxicology Program carcinogenicity bioassay resulted in a dose-dependent increases in renal cell hyperplasia, renal cell adenoma, and renal cell carcinomas (RCCs). Among those differentially expressed genes from controls and RCC of VDC-exposed mice, there was an overrepresentation of genes from pathways associated with chronic xenobiotic and oxidative stress as well as c-Myc overexpression and dysregulation of TP53 cell cycle checkpoint and DNA damage repair pathways in RCC. Trend analysis comparing RCC, VDC-exposed kidney, and chamber control kidney showed a conservation of pathway dysregulation in terms of overrepresentation of xenobiotic and oxidative stress, and DNA damage and cell cycle checkpoint pathways in both VDC-exposed kidney and RCC, suggesting that these mechanisms play a role in the pathogenesis of RCC in VDC-exposed mice.