Germline P53 Mutations Research Articles

Uncommon Metachronicity: Breast and Endometrial Carcinomas in a Young Woman with No Familial Cancer History - A Case Report

Background: Worldwide, breast and endometrial carcinomas are the two most common cancers of women, but their coexistence in a woman <40 years of age is a rare event. The risk of developing a new second primary cancer is higher in cancer survivors compared to those without a history of cancer. The exact pathogenesis of two different primary malignancies is not clear. Still, the coexistence of breast and endometrial carcinoma may be due to germline mutations of p53, BRCA 1, and BRCA 2 genes, environmental, hereditary, and immune-mediated factors. Case Presentation: The present case report briefs the finding of a 37-year-old woman who presented to the Gynecological outpatient department with complaints of abnormal uterine bleeding and a solid irregular lump in the upper outer quadrant of the right breast and the right axilla. She had a history of modified radical mastectomy for left breast ductal carcinoma five years back. She was diagnosed as a case of metachronous endometrial carcinoma (stage IA) with a recurrence of invasive ductal carcinoma in the right breast with metastasis to the right axillary lymph node. Conclusion: The metachronous development of breast and endometrial carcinoma in young women is relatively rare. In young patients <40 years of age with gynecological or breast carcinoma, a thorough work-up, including genetic testing and follow-up, should be done for early diagnosis of metachronous or synchronous primary malignancies of other organs.

Publication trends in sarcoma research: A bibliometric analysis.

e23530 Background: Sarcomas are rare primary malignant tumors originating from connective tissue, worldwide research trends on this topic are unclear. We conducted a bibliometric analysis of the 100 most influential articles on sarcoma. Methods: We sourced the literature published in academic journals indexed by the Scopus Database. Using the key words “sarcoma,” “soft-tissue tumor,” “bone sarcoma,” “soft-tissue sarcoma,” “connective tissue malignancy.” A list of the 100 most-cited articles were extracted and sorted by number of citations, excluding articles related to Kaposi’s Sarcoma. The papers were checked independently by our authors for accuracy of topic. R was used to extract the publishing journal, year of publication, country of origin, affiliated institutions, authors, and total citations. Results: The total number of citations for 100 articles was 73,483. Citations for individual articles ranged from 453 to 3,064, with a median of 588 (IQR: 341).The United States contributed the most papers (n = 70), followed by France and the United Kingdom (n = 5). Most articles, 67, were published prior to the 21st century and only 3 were in the last 10 years. The most cited study was Malkin et al.’s Germ line p53 mutations in a familial syndrome of breast cancer, sarcomas, and other neoplasms with 3,064 citations. Of the top 10, 8 were published between 1978 and 1998. Journal of Clinical Oncology made the greatest number of contribution to the research on sarcoma with a total of 18 articles (citations = 12,166), followed by Cancer with 15 (citations = 9,052), and Proceedings of the National Academy of Sciences of the United States with 10 (citations = 8,431). The median Scopus CiteScore for publishing journals was 19.7, the highest being The Lancet (115.3) and The Iowa Orthopaedic Journal with the lowest score (1.5). Conclusions: Within the United States, the institutions producing the most literature were the NCI and Memorial Sloan Kettering Cancer Center. The majority of the 100 most cited studies are cited an average of 735 times, indicating that these sources are providing a substantial foundation for emerging research in the field. This begs the question of the quality and relevance of evidence in the literature, given the amount of citations. The Lancet produced literature with the highest journalistic impact, having a CiteScore of 115.3 based on Scopus metrics. While the majority of the 100 most cited pieces of literature (67%) on sarcomas were published prior the 21st century, a small portion, eight, were produced in the last decade. This information could indicate either a lull in the frequency of research being conducted, or an effort on the part of institutions to create more substantial investigations that provide more valuable contributions to the field of research in sarcomas. We hope to continue seeing the field of sarcoma expand with new advances in its management.

LncRNA H19 Suppresses Osteosarcomagenesis by Regulating snoRNAs and DNA Repair Protein Complexes.

Osteosarcoma is one of the most frequent common primary malignant tumors in childhood and adolescence. Long non-coding RNAs (lncRNAs) have been reported to regulate the initiation and progression of tumors. However, the exact molecular mechanisms involving lncRNA in osteosarcomagenesis remain largely unknown. Li-Fraumeni syndrome (LFS) is a familial cancer syndrome caused by germline p53 mutation. We investigated the tumor suppressor function of lncRNA H19 in LFS-associated osteosarcoma. Analyzing H19-induced transcriptome alterations in LFS induced pluripotent stem cell (iPSC)-derived osteoblasts, we unexpectedly discovered a large group of snoRNAs whose expression was significantly affected by H19. We identified SNORA7A among the H19-suppressed snoRNAs. SNORA7A restoration impairs H19-mediated osteogenesis and tumor suppression, indicating an oncogenic role of SNORA7A. TCGA analysis indicated that SNORA7A expression is associated with activation of oncogenic signaling and poor survival in cancer patients. Using an optimized streptavidin-binding RNA aptamer designed from H19 lncRNA, we revealed that H19-tethered protein complexes include proteins critical for DNA damage response and repair, confirming H19's tumor suppressor role. In summary, our findings demonstrate a critical role of H19-modulated SNORA7A expression in LFS-associated osteosarcomas.

Association of Germline p53 Mutation With MLL Segmental Jumping Translocation in Treatment-Related Leukemia

AbstractSegmental jumping translocations are chromosomal abnormalities in treatment-related leukemias characterized by multiple copies of theABL and/or MLL oncogenes dispersed throughout the genome and extrachromosomally. Because gene amplification potential accompanies loss of wild-type p53, we examined the p53 gene in a case of treatment-related acute myeloid leukemia (t-AML) with MLLsegmental jumping translocation. The child was diagnosed with ganglioneuroma and embryonal rhabdomyosarcoma (ERMS) at 2 years of age. Therapy for ERMS included alkylating agents, DNA topoisomerase I and DNA topoisomerase II inhibitors, and local radiation. t-AML was diagnosed at 4 years of age. The complex karyotype of the t-AML showed structural and numerical abnormalities. Fluorescence in situ hybridization analysis showed multiple copies of the MLL gene, consistent with segmental jumping translocation. A genomic region including CD3ε, MLL, and a segment of band 11q24 was unrearranged and amplified by Southern blot analysis. There was no family history of a cancer predisposing syndrome, but single-strand conformation polymorphism (SSCP) analysis detected identical band shifts in the leukemia, ganglioneuroma, ERMS, and normal tissues, consistent with a germline p53 mutation, and there was loss of heterozygosity in the ERMS and the t-AML. Sequencing showed a CGA→TGA nonsense mutation at codon 306 in exon 8. The results of this analysis indicate that loss of wild-type p53 may be associated with genomic instability after DNA-damaging chemotherapy and radiation, manifest as a complex karyotype and gene amplification in some cases of t-AML.

A Mouse Homolog of a Human TP53 Germline Mutation Reveals a Lipolytic Activity of p53

SUMMARYThe physiological effects of the many germline mutations of TP53, encoding the tumor suppressor protein p53, are poorly understood. Here we report generating a p53 R178C knockin mouse modeling the human TP53 R181C mutation, which is notable for its prevalence and prior molecular characterization. Consistent with its weak cancer penetrance in humans, homozygous p53178C/C mice show a modest increase in tumorigenesis but, surprisingly, are lean with decreased body fat content. They display evidence of increased lipolysis and upregulation of fatty acid metabolism in their inguinal white adipose tissue (iWAT). Gene expression and chromatin immunoprecipitation sequencing (ChIP-seq) analyses show that the mutant p53 bound and trans-activated Beta-3-Adrenergic Receptor (ADRB3), a gene that is known to promote lipolysis and is associated with obesity. This study reveals that a germline mutation of p53 can affect fat metabolism, which has been implicated in cancer development.

GENE-03. CASE REPORT: CHOROID PLEXUS CARCINOMA TREATED WITH STEREOTACTIC RADIOSURGERY IN 2-YEAR-OLD WITH LI FRAUMENI SYNDROME

CASE REPORT: The child was diagnosed with choroid plexus carcinoma (CPC) at age 8 months. The tumor was positive for TP53 genomic alteration. Germline genetic test sent at age 10 months returned positive for heterozygous TP53 pathogenic mutation. She underwent surgical resection, chemotherapy, and repeat resection at 13 months. At 31 months, she had three small nodules concerning for recurrent disease surrounding the large surgical cavity. Repeat surgical resection was declined by her family. She therefore underwent stereotactic radiosurgery (SRS) to each lesion to a dose of 24 Gy in 1 fraction. Anesthesia was required at the time of planning CT simulation and at the time of treatment. At one year since the procedure, she has demonstrated no further recurrent disease and no evidence of secondary malignancy at this time. DISCUSSION: Retrospective studies approximate rates of p53 mutations in children with CPC at 40%. Radiation is avoided—unless absolutely necessary—in children less than 3 years old because of the risks of neurocognitive delay, endocrine and growth toxicities, and the risk of secondary malignancies. Patients with Li Fraumeni syndrome have further dramatic increase in rates of developing secondary malignancies after exposure to ionizing radiation. SRS provides dose escalation while minimizing ionizing radiation dose to normal surrounding tissue. CONCLUSIONS: Any child diagnosed with CPC should undergo genetic testing for p53 germline mutation because of high pre-test probability. In selected cases, SRS provides an alternative to surgery as local control for limited CPC. SRS minimizes the dose of ionizing radiation delivered to normal tissues in patients with p53 mutation, or in young patients in whom less-focused radiation techniques may cause increased negative neurocognitive effects. Because of the short treatment course, SRS also minimizes the number of days under anesthesia required for young patients, thereby minimizing risk incurred by repeated anesthetic requirements.

P53, stem cell biology and childhood blastomas

Childhood blastomas, unlike adult cancers, originate from developing organs in which molecular and cellular features exhibit differentiation arrest and embryonic characteristics. Conventional cancer therapies, which rely on the generalized cytotoxic effect on rapidly dividing cells, may damage delicate organs in young children, leading to multiple late effects. Deep understanding of the biology of embryonal cancers is crucial in reshaping the cancer treatment paradigm for children. p53 plays a major physiological role in embryonic development, by controlling cell proliferation, differentiation and responses to cellular stress. Tumor suppressor function of p53 is commonly lost in adult cancers through genetic alterations. However, both somatic and germline p53 mutations are rare in childhood blastomas, suggesting that in these cancers, p53 may be inactivated through other mechanisms than mutation. In this review, we summarize current knowledge about p53 pathway inactivation in childhood blastomas (specifically neuroblastoma, retinoblastoma and Wilms' tumor) through various upstream mechanisms. Laboratory evidence and clinical trials of targeted therapies specific to exploiting p53 upstream regulators are discussed. Despite the low rate of inherent TP53 mutations, p53 pathway inactivation is a common denominator in childhood blastomas. Exploiting p53 and its regulators is likely to translate into more effective targeted therapies with minimal late effects for children. (see Video Abstract, Supplemental Digital Content 1, http://links.lww.com/COON/A23).

An EBNA3C-deleted Epstein-Barr virus (EBV) mutant causes B-cell lymphomas with delayed onset in a cord blood-humanized mouse model.

EBV causes human B-cell lymphomas and transforms B cells in vitro. EBNA3C, an EBV protein expressed in latently-infected cells, is required for EBV transformation of B cells in vitro. While EBNA3C undoubtedly plays a key role in allowing EBV to successfully infect B cells, many EBV+ lymphomas do not express this protein, suggesting that cellular mutations and/or signaling pathways may obviate the need for EBNA3C in vivo under certain conditions. EBNA3C collaborates with EBNA3A to repress expression of the CDKN2A-encoded tumor suppressors, p16 and p14, and EBNA3C-deleted EBV transforms B cells containing a p16 germline mutation in vitro. Here we have examined the phenotype of an EBNAC-deleted virus (Δ3C EBV) in a cord blood-humanized mouse model (CBH). We found that the Δ3C virus induced fewer lymphomas (occurring with a delayed onset) in comparison to the wild-type (WT) control virus, although a subset (10/26) of Δ3C-infected CBH mice eventually developed invasive diffuse large B cell lymphomas with type III latency. Both WT and Δ3C viruses induced B-cell lymphomas with restricted B-cell populations and heterogeneous T-cell infiltration. In comparison to WT-infected tumors, Δ3C-infected tumors had greatly increased p16 levels, and RNA-seq analysis revealed a decrease in E2F target gene expression. However, we found that Δ3C-infected tumors expressed c-Myc and cyclin E at similar levels compared to WT-infected tumors, allowing cells to at least partially bypass p16-mediated cell cycle inhibition. The anti-apoptotic proteins, BCL2 and IRF4, were expressed in Δ3C-infected tumors, likely helping cells avoid c-Myc-induced apoptosis. Unexpectedly, Δ3C-infected tumors had increased T-cell infiltration, increased expression of T-cell chemokines (CCL5, CCL20 and CCL22) and enhanced type I interferon response in comparison to WT tumors. Together, these results reveal that EBNA3C contributes to, but is not essential for, EBV-induced lymphomagenesis in CBH mice, and suggest potentially important immunologic roles of EBNA3C in vivo.

Mutant p53 gain of function induces HER2 over-expression in cancer cells

BackgroundHER2 over-expression is related with a poor prognosis in patients with invasive breast cancer tumors. Clinical associations have reported that somatic mutations of p53 more frequently detected in cases of sporadic breast cancer of the HER2 subtypes, besides a high percentage of HER2-amplifying tumors carry germline mutations of p53. The mechanisms responsible for the acquisition of oncogenic functions of p53 mutant proteins (mtp53), known as Gain of Function (GOF), over HER2 expression have not been reported. The objective of this study was to evaluate a possible relationship between p53 mutants and HER2 regulation.MethodsHER2 expression (transcription and protein), as well as HER2 protein stabilization have been evaluated after inducing or silencing of p53 mutants’ expression in cell lines. Finally, we evaluated the interaction of the p53 mutants over the HER2 receptor promoter.ResultsHigher HER2 expression in cell lines harboring endogenous mtp53 compared with wt or null expression of p53 cell lines. Transfection of p53 mutants (R248Q and R273C) in cell lines increased the expression of HER2. Silencing of p53 mutants, decrease HER2 expression. The p53 mutants R248Q and R273C significantly increase the luciferase activity on the HER2 promoter, and both mutants also promote acetylation of H3 and H4 histones binding in it.ConclusionsThese findings show for the first time that p53 mutants induce over-expression of HER2 at transcriptional level of the HER2 protein. Our results could have clinical implications in breast cancer and other types of cancer where HER2 is over-expressed and used as a therapy target.

Abstract PR05: Familial cancer patient specific iPSCs based study of a potential oncogenic factor, sFRP2, in osteosarcoma

Abstract Osteosarcoma (OS), despite being a serious pediatric cancer with the lowest survival rate, is not well studied due to its rare occurrence. For this reason, studying OS in a patient-specific manner is more challenging. In an attempt to establish a method to study OS in a patient-specific manner, we previously generated Li-Fraumeni syndrome (LFS---a germline P53 mutation disease) patient specific iPSCs, and demonstrated that the LFS-iPSC derived mesenchymal stem cells (MSCs) and osteoblasts (OBs) can recapitulate tumorigenic phenotypes in vitro and in vivo (Lee, Kim, et al., Cell 2015). Taking advantage of this new platform, we focused on finding potential therapeutic targets. We found SFRP2 (secreted frizzled related protein 2) to be highly expressed in LFS iPSC-derived MSCs and OBs by global RNA-seq gene expression analysis. Using global transcript profiling of ectopic overexpression (OE) of sFRP2 in wild-type MSCs, we discovered that sFRP2 regulates osteoblast differentiation, cell adhesion, cell proliferation, and apoptosis-related gene sets. Further functional assays revealed that sFRP2 OE contributes to the generation of abnormal osteoblast cells, which are the cause of OS, by suppressing osteogenic alkaline phosphatase activity, mineralization, and osteogenic BMP and WNT signaling. Furthermore, ectopic OE of sFRP2 is associated with LFS mediated tumorigenesis. sFRP2 OE in MSCs and OBs facilitates cell proliferation through cell cycle alteration and increases anchorage independency. In in ovo and in vivo xenograft assays (subcutaneous and intratibial injections), the tumor size is significantly increased in sFRP2 OE group and decreased in knockdown sFRP2 group. Interestingly, we also found that sFRP2 OE augments angiogenesis in vitro and in vivo, suggesting sFRP2 has a role as a paracrine factor. In the future, we will analyze sFRP2 expression levels in human OS tissue samples to determine whether sFRP2 OE has clinical relevance. Conclusively, this study demonstrates that P53 mutation-mediated osteosarcoma exerts oncogenicity through sFRP2 overexpression and that targeting of sFRP2 is a potential therapeutic strategy for OS treatment. This abstract is also being presented as Poster A22. Citation Format: Huen Suk Kim, Ye X. Yuan, Jeffrey M. Bernitz, Andreia Gomes, Christopher Schaniel, Dung-Fang Lee, Ihor R. Lemischka. Familial cancer patient specific iPSCs based study of a potential oncogenic factor, sFRP2, in osteosarcoma [abstract]. In: Proceedings of the AACR Conference on Advances in Sarcomas: From Basic Science to Clinical Translation; May 16-19, 2017; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(2_Suppl):Abstract nr PR05.

Pancreatic cancer screening in high-risk individuals with germline genetic mutations

Pancreatic cancer (PC) is a deadly disease that is most commonly diagnosed at an incurable stage. Different high-risk genetic variants and cancer syndromes increase the lifetime risk of developing PC. This study aims to assess the yield of initial PC screening in patients with high-risk germline mutations. Asymptomatic adults underwent PC screening by EUS, magnetic resonance imaging, or CT during a 10-year period and were retrospectively identified. High-risk individuals were defined as carrying germline mutations in BRCA1, BRCA2, p53 (Li-Fraumeni), STK11 (Peutz-Jeghers), MSH2 (Lynch), ATM (ataxia-telangiectasia), or APC (familial adenomatous polyposis). Patients without germline mutations were excluded. In total, 86 patients met the study criteria. The median age was 48.5 years (interquartile range, 40-58), 79.1% (68) were women, and 43.0% (37) had a family history of PC. The genetic mutations were BRCA2 (50,58.1%), BRCA1 (14, 16.3%), p53 (12, 14.0%), STK11 (5, 5.8%), MSH2 (3, 3.5%), ATM (1, 1.2%), and APC (1,1.2%). Screening detected a pancreatic abnormality (PA) in 26.7% (23/86), including cysts (11, 47.8%), hyperechoic strands and foci (10, 43.5%), and mild pancreatic duct dilation (2, 8.7%). Patients older than 60 years were more likely to have a PA detected (P= .043). EUS detected more PAs than magnetic resonance imaging or CT. Nocases of PC were diagnosed by screening or during follow-up (median, 29.8 months; interquartile range, 21.7-43.5). Unless indicated otherwise by family or personal history, PC screening under the age of 50 is low yield. Linear EUS may be the preferred modality for initial PC screening.

Combined Loss of EAF2 and p53 Induces Prostate Carcinogenesis in Male Mice.

Mutations in the p53 tumor suppressor are frequent in patients with castration-resistant prostate cancer but less so in patients with localized disease, and patients who have Li-Fraumeni with germline p53 mutations do not have an increased incidence of prostate cancer, suggesting that additional molecular and/or genetic changes are required for p53 to promote prostate carcinogenesis. ELL-associated factor 2 (EAF2) is a tumor suppressor that is frequently downregulated in advanced prostate cancer. Previous studies have suggested that p53 binds to EAF2, providing a potential mechanism for their functional interactions. In this study, we tested whether p53 and EAF2 could functionally interact in prostate cancer cells and whether concurrent inactivation of p53 and EAF2 could promote prostate carcinogenesis in a murine knockout model. Endogenous p53 coprecipitated with EAF2 in prostate cancer cells, and deletion mutagenesis indicated that this interaction was mediated through the C terminus of EAF2 and the DNA binding domain of p53. Concurrent knockdown of p53 and EAF2 induced an increase in proliferation and migration in cultured prostate cancer cells, and conventional p53 and EAF2 knockout mice developed prostate cancer. In human prostate cancer specimens, concurrent p53 nuclear staining and EAF2 downregulation was associated with high Gleason score. These findings suggest that EAF2 and p53 functionally interact in prostate tumor suppression and that simultaneous inactivation of EAF2 and p53 can drive prostate carcinogenesis.

Abstract LB-169: Targeted tumor prevention in Li-Fraumeni syndrome

Abstract Li-Fraumeni syndrome (LFS) is a rare, dominant cancer predisposition disorder. The majority of LFS cases are brought about by germline p53 mutations. Individuals affected develop a spectrum of cancers occurring since childhood or early adulthood (Mai et al, 2012). Tumors associated with LFS include but are not limited to sarcomas, lymphomas and leukemias, breast cancers, brain tumors, etc (Olivier et al, 2010; Mai et al, 2012). p53 is a tumor suppressor gene that regulates many cellular processes and prevents cancer formation, growth and dissemination. When inactivated by mutation, it not only loses its tumor suppressive function, but also acquires oncogenic properties. Mutations of p53 promote destabilization of its three-dimensional structure, which can cause p53 to partially unfold, aggregate and accumulate in cells (Xu et al, 2011). We developed a peptide, ReACp53, to arrest p53 clumping and accumulation, chaperone p53 to a functional state and promote its degradation, hence arresting tumor proliferation (Soragni et al, 2016). ReACp53 can also target p63 and p73 (Zheng et al, 2016). It has shown good tolerability and a favorable safety profile in vivo. In LFS patients and LFS animal models, p53 is undetectable in normal tissues. However, the protein starts to accumulate in tumors and pre-cancerous lesions (Lang et al, 2004; Soragni et al, unpublished data). As p53 accumulation is correlated with aggregation (Xu et al, 2011; Soragni et al, 2016), we postulated that, by periodically administering ReACp53, we could target this initial event early on, and prevent p53 accumulation and/or push the p53 aggregates-ridden cells into apoptosis (Soragni et al, 2016). Here, we tested ReACp53 as a tumor preventive agent, by administering it twice a week from weaning to LFS mice carrying the aggregation-prone R172H mutation (R175H in humans). ReACp53 delayed cancer onset resulting in a significant increase in overall survival (OS). The homozygous cohort (n=16/group) showed a 50% increase in median survival in ReACp53-treated mice (median OS 15.79 vs 23.93 weeks for vehicle vs ReACp53, p=0.005; HR=2.6). Heterozygous mice (n=8/group) showed a 36% increase in OS with a median of 58.86 vs 80.50 weeks for vehicle vs ReACp53 (p=0.03, HR=2.6). No long-term side effect was observed upon ReACp53 administration. The tumor spectrum reported differed between the two groups. Activation of a functional p53 response in mouse cells and LFS patient cells treated with ReACp53 is undergoing. Overall, our study supports ReACp53 therapy as a potential approach to tumor prevention in LFS. Citation Format: Alice Soragni, Nhan Phan, David Eisenberg. Targeted tumor prevention in Li-Fraumeni syndrome [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 LB-169. doi:10.1158/1538-7445.AM2017-LB-169

P11.13 Metachronous spinal primitive neuroectodermal tumor after treatment of a hemispheric anaplastic oligodendroglioma through p53 germline mutation

P11.13 Metachronous spinal primitive neuroectodermal tumor after treatment of a hemispheric anaplastic oligodendroglioma through p53 germline mutation

Abstract A45: Neat1 is a p53-inducible lincRNA important for pancreatic cancer suppression

Abstract p53’s role as a critical tumor suppressor is well illustrated by the fact that it is found mutated in more than half of all human cancers. In addition, the highly penetrant cancer predisposition observed in Li-Fraumeni patients with germline mutations in p53 and in p53 null mice further highlight the importance of p53 in tumor suppression. p53 is a transcription factor, able to sense stress signals such as DNA damage and hyperproliferative signals, promoting in turn the expression of a network of target genes to induce anti-proliferative responses such as cell cycle arrest or apoptosis. Although studies of genes in the p53 network have largely focused on protein-coding genes, in recent years the role of p53-regulated non-coding (nc) RNAs in p53 biology has been increasingly recognized. To identify new p53-regulated ncRNAs that could contribute to tumor suppression, we leveraged chromatin immunoprecipitation (ChIP)-sequencing and RNA-sequencing (RNA-seq) data we generated previously in p53 wild-type and p53-/- mouse embryonic fibroblasts (MEFs). This analysis uncovered the lincRNA Neat1 as a p53 target gene. We showed that Neat1 is induced by different DNA damaging agents in a p53-dependent manner in different mouse and human cell types, including pancreatic ductal cells. Although we found that Neat1 is not involved in DNA damage-induced p53-dependent cell cycle arrest or apoptosis responses, we found that it does play a role in p53-dependent transformation suppression in both oncogene-expressing fibroblasts and pancreatic cancer cells. Furthermore, using oncogene-expressing fibroblasts, we showed that Neat1 loss promotes cancer in vivo in a subcutaneous xenograft model. Preliminary evidence suggests also that Neat1 loss promotes acinar to ductal metaplasia, bolstering its importance in pancreatic cancer suppression. These findings highlight the importance of Neat1 for p53 tumor suppressor function. Detailed analyses of Neat1’s mechanism of action will provide insights in its function as a tumor suppressor. Citation Format: Stephano S. Mello, Carolyn Sinow, Pawel K. Mazur, Hannes Vogel, John Rinn, Laura D. Attardi.{Authors}. Neat1 is a p53-inducible lincRNA important for pancreatic cancer suppression. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2016 May 12-15; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2016;76(24 Suppl):Abstract nr A45.

Expression profile of E-cadherin, estrogen receptors, and P53 in early-onset gastric cancers.

Early‐onset gastric cancer (EOGC) is predominant in females, diffuse histology, and hereditary pattern. Germline mutation of CDH1 and p53 has been reported previously and female dominance was speculated to be associated with estrogen and its receptors. Expression of E‐cadherin, estrogen receptor α (ER α), estrogen receptor β (ER β), and p53 in EOGC remains unclear, which was the focus of this study, to assess clinical significance of their expression in EOGC. The expression of E‐cadherin, ER α, ER β, and p53 in tumors and normal tissues from surgically resected EOGCs was assessed by immunohistochemistry (n = 139) and Western blot (n = 7) methods, respectively. The expression in tumor tissues was significantly higher for ER α, ER β, and p53, but lower for E‐cadherin, compared to uninvolved mucosa. Positive staining of ER β and p53 was more frequently observed in younger patients with advanced TNM stages. For E‐cadherin, significant correlation was observed between the immunopositivity and TNM stages IA+IB. P53‐negative patients had significantly better outcomes than p53‐positive patients. Significant association between expression of E‐cadherin and histologic types was found in familial, but not in sporadic, EOGC. In conclusion, our results demonstrated E‐cadherin may have a role in initiation of EOGC and positive ER β and p53 expression may partially explained early‐onset and tumor progression of EOGC.

Juvenile Granulosa Cell Tumor of the Ovary: A Clinicopathologic Study

Study ObjectiveTo report on the clinical characteristics and outcome of pediatric patients with juvenile granulosa cell tumor (JGCT) of the ovary. Design, Setting, Participants, Interventions, and Main Outcome MeasuresPatients with histopathologically confirmed ovarian JGCT diagnosed between 1990 and 2016 were identified. Data on the clinical presentation, surgical management, oncologic management, laboratory investigation, follow-up, and outcome were collected. Tumors were staged according to the International Federation of Gynecology and Obstetrics criteria. ResultsEight patients were diagnosed with ovarian JGCT during the study period. The median age at presentation was 3 years (range, 0.7-14 years). Precocious puberty was the presenting symptom in all five prepubertal children; abdominal distension due to mass effect was the presenting symptom in three children older than 9 years of age. In patients who had preoperative serologic testing, estradiol (n = 3) and inhibin (n = 3) levels were elevated. Five patients had stage I disease, and three had stage III. All stage I patients underwent salpingo-oophorectomy as the only treatment. Stage III patients received adjuvant chemotherapy. After a median follow-up of 6.2 years, six patients (75%) were alive without evidence of disease. One stage I patient with germline p53 mutation and phosphatase and tensin homolog mutation, died because of subsequent liposarcoma. One patient with stage IIIB disease developed recurrence detected according to an elevated inhibin serum level, and died due to progressive disease despite receiving multiple chemotherapy regimens. ConclusionJuvenile granulosa cell tumor has a favorable prognosis in patients with stage I disease after surgical resection alone. Adjuvant chemotherapy might be indicated in patients with higher-stage tumors.

Abstract B07: Gain of function activities of p53R245W in a conditional knock-in mouse model

Abstract Missense mutations of the p53 tumor suppressor gene are present in more than 50% of human tumors. Emerging evidence suggests that p53 missense mutations in human cancer patients are associated with the poor prognosis and drug resistance due to their gain-of-function (GOF) properties, rather than just the loss of normal p53 function. Interestingly, these missense mutations have been shown to contribute to tumor progression by different mechanisms: interactions with Smad and SREBPs, modulation of EGFR/integrin signaling, and inhibition of p63 and p73 tumor suppressors. Combined, these studies suggest that the GOF of p53 missense mutations may be dependent on cell contexts and mutation types. Of particular interest, the p53R248W mutation is one of the most frequent missense mutation among human cancer patients; yet to date a mouse model that faithfully mimics this mutation does not exist. Additionally, the majority of human cancers evolve from somatic inactivation of p53 and current mouse models do not mimic somatic mutation of p53. Thus, we generated a conditional p53WMR245/+ knock-in mouse model, which normally expresses wild type p53, but produces p53R245W by Cre-mediated deletion of wild type p53 sequences. Germ line mutation of p53R245W/+ can rescue the early embryonic lethality of Mdm2 and Mdm4 null alleles like p53 null. Preliminary analyses of p53R245W/+ cohorts indicate differences in tumor spectrum in comparison to p53+/- and p53R172H/+ mice. Somatic mutation of p53R245W has also been performed to examine germ line versus somatic differences in tumor development. This study will help us understand the in vivo contributions of different p53 missense mutations to tumorigenesis, as well as the genetic alterations that occur during tumor evolution initiated by both germ line and somatic p53 mutations. Citation Format: Shunbin Xiong, Yun Zhang, Guillermina Lozano. Gain of function activities of p53R245W in a conditional knock-in mouse model. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Metastasis; 2015 Nov 30-Dec 3; Austin, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(7 Suppl):Abstract nr B07.

Abstract P3-03-03: A mouse model of sporadic breast tumor with a conditional P53 mutation

Abstract Missense mutations in the tumor suppressor gene p53 are present in more than 50% of human tumors. In particular, the arginine-to-histidine mutation at codon 175 (p53R175H) has been found in more than 4% of human breast cancers. p53 mutations fall into two general categories: germline mutations that are associated with hereditary tumors, and somatic mutations that cause sporadic tumors. Mouse models for cancers induced by p53 germline mutations have been established and characterized. However, hitherto there are no accurate animal models for sporadic tumors induced by p53 somatic missense mutations, although this mechanism of inactivating p53 occurs in a large fraction of human cancers. We have generated a mouse allele, p53WM, that carries coding sequences for both wild type (WT) and R172H mutant p53 (corresponding to R175H in humans) at the p53 endogenous locus. The coding sequence for WT p53 is flanked by loxP sites and therefore can be deleted by Cre recombinase. In the absence of Cre, following radiation, the p53WM/+ mice have the similar ability as the p53 WT mice to activate p53, as demonstrated by transcriptional activation of p53 targets and induction of apoptosis. In addition, like WT mouse embryonic fibroblasts (MEFs), the p53WM/+ MEFs also underwent cell cycle arrest following radiation. After crossing p53WM/+ mice with Zp3-Cre mice which express the Cre recombinase in oocytes, the coding sequence for WT p53 was removed, and the mutant p53R172H allele was regenerated as determined by sequencing, western blot analysis and real time reverse transcription PCR. To investigate whether a somatic p53R172H mutation in mammary epithelium can induce breast tumors, adenoviruses expressing Cre (Ad-Cre) were injected into the mammary duct of p53WM/+ mice. Limiting the adenovirus dose allowed us to induce the p53R172H mutation in 1 of 20, 100 or 1000 cells, generating a mutant p53 surrounded by normal cells which more accurately mimics the clinical situation of sporadic tumors. Meanwhile, p53WM/+; K14-Cre and p53WM/+; WAP-Cre mice have been generated, in which p53R172H mutation are induced through Cre transgene specifically in the mammary epithelial cells. Lastly, to examine how somatic p53 mutation cooperates with an oncogene in mammary tumor induction, MMTV-neu; p53WM/+ mice were also subjected to the mammary intraductal Ad-Cre injection. We are currently monitoring mice and will further investigate the molecular mechanisms of tumorigenesis by studying the loss of heterozygosity, and the co-evolution of tumor epithelial cells and their microenvironment. This mouse model is more likely to share the underlying molecular pathology with human sporadic tumors. Therefore, it will be more predictive of human responses to drugs, and thus a more valuable tool in preclinical testing. Citation Format: Zhang Y, Xiong S, Pant V, El-Naggar A, Lozano G. A mouse model of sporadic breast tumor with a conditional P53 mutation. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P3-03-03.

Phenotypic and Histopathological Tumor Characteristics According to CDKN2A Mutation Status among Affected Members of Melanoma Families

Phenotypic and Histopathological Tumor Characteristics According to CDKN2A Mutation Status among Affected Members of Melanoma Families