The genomic landscape of retinoblastoma: a review

Retinoblastoma is a paediatric ocular tumour that continues to reveal much about the genetic basis of cancer development. Study of genomic aberrations in retinoblastoma tumours has exposed important mechanisms of cancer development and identified oncogenes and tumour suppressors that offer potential points of therapeutic intervention. The recent development of next-generation genomic technologies has allowed further refinement of the genomic landscape of retinoblastoma at high resolution. In a relatively short period of time, a wealth of genetic and epigenetic data has emerged on a small number of tumour samples. These data highlight the inherent molecular complexity of this cancer despite the fact that most retinoblastomas are initiated by the inactivation of a single tumour suppressor gene. This review outlines the current understanding of the genomic, genetic and epigenetic changes in retinoblastoma, highlighting recent genome-wide analyses that have identified exciting candidate genes worthy of further validation as potential prognostic and therapeutic targets

Molecular analysis distinguishes metastatic disease from second cancers in patients with retinoblastoma

The pediatric ocular tumor retinoblastoma readily metastasizes, but these lesions can masquerade as histologically similar pediatric small round blue cell tumors. Since 98% of retinoblastomas have RB1 mutations and a characteristic genomic copy number “signature”, genetic analysis is an appealing adjunct to histopathology to distinguish retinoblastoma metastasis from second primary cancer in retinoblastoma patients. Here, we describe such an approach in two retinoblastoma cases. In patient one, allele-specific (AS)-PCR for a somatic nonsense mutation confirmed that a temple mass was metastatic retinoblastoma. In a second patient, a rib mass shared somatic copy number gains and losses with the primary tumor. For definitive diagnosis, however, an RB1 mutation was needed, but heterozygous promoter→exon 11 deletion was the only RB1 mutation detected in the primary tumor. We used a novel application of inverse PCR to identify the deletion breakpoint. Subsequently, AS-PCR designed for the breakpoint confirmed that the rib mass was metastatic retinoblastoma. These cases demonstrate that personalized molecular testing can confirm retinoblastoma metastases and rule out a second primary cancer, thereby helping to direct the clinical management

Achieving optimal cancer outcomes in East Africa through multidisciplinary partnership: a case study of the Kenyan National Retinoblastoma Strategy group.

BACKGROUND: Strategic, interdisciplinary partnerships are essential to addressing the complex drivers of health inequities that result in survival disparities worldwide. Take for example the aggressive early childhood eye cancer retinoblastoma, where survival reaches 97 % in resource-rich countries, but is as low 30 % in some resource-limited nations, where 92 % of the burden lies. This suggests a need for a multifaceted approach to achieve a tangible and sustainable increase in survival. METHODS: We assembled the history the Kenyan National Retinoblastoma Strategy (KNRbS), using information documented in NGO reports, grant applications, news articles, meeting agendas and summaries. We evaluated the KNRbS using the principles found in the guide for transboundary research partnerships developed by the Swiss Commission for Research Partnerships with Developing Countries. RESULTS: A nationally co-ordinated approach drawing input and expertise from multiple disciplines and sectors presented opportunities to optimise cure of children with retinoblastoma. Annual meetings were key to achieving the over 40 major outputs of the group's efforts, related to Awareness, Medical Care, Family Support and Resource Mobilization. Three features were found to be critical to the KNRbS success: multidisciplinarity, consistency and flexibility. CONCLUSION: The KNRbS has achieved a number of key outputs with limited financial investment. As a partnership, the KNRbS meets most of the criteria identified for success. Challenges remain in securing the long-term sustainability of its achievements. Elements of the Kenyan National Retinoblastoma Strategy may be useful to other developing countries struggling with limited survival of retinoblastoma and other cancers or rare diseases

Regulation of p14ARF expression by miR-24: a potential mechanism compromising the p53 response during retinoblastoma development

<p>Abstract</p> <p>Background</p> <p>Most human cancers show inactivation of both pRB- and p53-pathways. While retinoblastomas are initiated by loss of the <it>RB1 </it>tumor suppressor gene, <it>TP53 </it>mutations have not been found. High expression of the p53-antagonist MDM2 in human retinoblastomas may compromise p53 tumor surveillance so that <it>TP53 </it>mutations are not selected for in retinoblastoma tumorigenesis. We previously showed that p14^ARFprotein, which activates p53 by inhibiting MDM2, is low in retinoblastomas despite high mRNA expression.</p> <p>Methods</p> <p>In human fetal retinas, adult retinas, and retinoblastoma cells, we determined endogenous <it>p14^ARF</it>mRNA, ARF protein, and miR-24 expression, while integrity of p53 signalling in WERI-Rb1 cells was tested using an adenovirus vector expressing p14^ARF. To study p14^ARFbiogenesis, retinoblastoma cells were treated with the proteasome inhibitor, MG132, and siRNA against miR-24.</p> <p>Results</p> <p>In human retinoblastoma cell lines, <it>p14^ARF</it>mRNA was disproportionally high relative to the level of p14^ARFprotein expression, suggesting a perturbation of p14^ARFregulation. When p14^ARFwas over-expressed by an adenovirus vector, expression of p53 and downstream targets increased and cell growth was inhibited indicating an intact p14^ARF-p53 axis. To investigate the discrepancy between <it>p14^ARF</it>mRNA and protein in retinoblastoma, we examined p14^ARFbiogenesis. The proteasome inhibitor, MG132, did not cause p14^ARFaccumulation, although p14^ARFnormally is degraded by proteasomes. miR-24, a microRNA that represses p14^ARFexpression, is expressed in retinoblastoma cell lines and correlates with lower protein expression when compared to other cell lines with high <it>p14^ARF</it>mRNA. Transient over-expression of siRNA against miR-24 led to elevated p14^ARFprotein in retinoblastoma cells.</p> <p>Conclusions</p> <p>In retinoblastoma cells where high levels of <it>p14^ARF</it>mRNA are not accompanied by high p14^ARFprotein, we found a correlation between miR-24 expression and low p14^ARFprotein. p14^ARFprotein levels were restored without change in mRNA abundance upon miR-24 inhibition suggesting that miR-24 could functionally repress expression, effectively blocking p53 tumor surveillance. During retinal tumorigenesis, miR-24 may intrinsically compromise the p53 response to <it>RB1 </it>loss.</p

Characterisation of retinoblastomas without RB1 mutations: genomic, gene expression, and clinical studies

SummaryBackgroundRetinoblastoma is the childhood retinal cancer that defined tumour-suppressor genes. Previous work shows that mutation of both alleles of the RB1 retinoblastoma suppressor gene initiates disease. We aimed to characterise non-familial retinoblastoma tumours with no detectable RB1 mutations.MethodsOf 1068 unilateral non-familial retinoblastoma tumours, we compared those with no evidence of RB1 mutations (RB1+/+) with tumours carrying a mutation in both alleles (RB1−/−). We analysed genomic copy number, RB1 gene expression and protein function, retinal gene expression, histological features, and clinical data.FindingsNo RB1 mutations (RB1+/+) were reported in 29 (2·7%) of 1068 unilateral retinoblastoma tumours. 15 of the 29 RB1+/+ tumours had high-level MYCN oncogene amplification (28–121 copies; RB1+/+MYCNA), whereas none of 93 RB1−/− primary tumours tested showed MYCN amplification (p<0·0001). RB1+/+MYCNA tumours expressed functional RB1 protein, had fewer overall genomic copy-number changes in genes characteristic of retinoblastoma than did RB1−/− tumours, and showed distinct aggressive histological features. MYCN amplification was the sole copy-number change in one RB1+/+MYCNA retinoblastoma. One additional MYCNA tumour was discovered after the initial frequencies were determined, and this is included in further analyses. Median age at diagnosis of the 17 children with RB1+/+MYCNA tumours was 4·5 months (IQR 3·5–10), compared with 24 months (15–37) for 79 children with non-familial unilateral RB1−/− retinoblastoma.InterpretationAmplification of the MYCN oncogene might initiate retinoblastoma in the presence of non-mutated RB1 genes. These unilateral RB1+/+MYCNA retinoblastomas are characterised by distinct histological features, only a few of the genomic copy-number changes that are characteristic of retinoblastoma, and very early age of diagnosis.FundingNational Cancer Institute–National Institutes of Health, Canadian Institutes of Health Research, German Research Foundation, Canadian Retinoblastoma Society, Hyland Foundation, Toronto Netralaya and Doctors Lions Clubs, Ontario Ministry of Health and Long Term Care, UK-Essen, and Foundations Avanti-STR and KiKa

Retinoblastoma seeds: Impact on American Joint Committee on Cancer clinical staging

Aim To investigate whether the American Joint Committee on Cancer (AJCC) clinical category cT2b needs to be subclassified by the type and distribution of retinoblastoma (RB) seeding. Methods Multicentre, international registry-based data were collected from RB centres enrolled between January 2001 and December 2013. 1054 RB eyes with vitreous or subretinal seeds from 18 ophthalmic oncology centres, in 13 countries within six continents were analysed. Local treatment failure was defined as the use of secondary enucleation or external beam radiation therapy (EBRT) and was estimated with the Kaplan-Meier method. Results Clinical category cT2b included 1054 eyes. Median age at presentation was 16.0 months. Of these, 428 (40.6%) eyes were salvaged, and 430 (40.8%) were treated with primary and 196 (18.6%) with secondary enucleation. Of the 592 eyes that had complete data for globe salvage analysis, the distribution of seeds was focal in 143 (24.2%) and diffuse in 449 (75.8%). The 5-year Kaplan-Meier cumulative globe-salvage (without EBRT) was 78% and 49% for eyes with focal and diffuse RB seeding, respectively. Cox proportional hazards regression analysis confirmed a higher local treatment failure risk with diffuse seeds as compared with focal seeds (hazard rate: 2.8; p<0.001). There was insufficient evidence to prove or disprove an association between vitreous seed type and local treatment failure risk(p=0.06). Conclusion This international, multicentre, registry-based analysis of RB eyes affirmed that eyes with diffuse intraocular distribution of RB seeds at diagnosis had a higher risk of local treatment failure when compared with focal seeds. Subclassification of AJCC RB category cT2b into focal vs diffuse seeds will improve prognostication for eye salvage.Fil: Tomar, Ankit Singh. New York Eye Cancer Center; Estados UnidosFil: Finger, Paul T.. New York Eye Cancer Center; Estados UnidosFil: Gallie, Brenda. University Of Toronto. Hospital For Sick Children; CanadáFil: Kivelä, Tero. University of Helsinki; Finlandia. Helsinki University Hospital; FinlandiaFil: Mallipatna, Ashwin. University Of Toronto. Hospital For Sick Children; Canadá. Narayana Nethralaya; IndiaFil: Zhang, Chengyue. Beijing Children's Hospital; ChinaFil: Zhao, Junyang. Beijing Children's Hospital; ChinaFil: Wilson, Matthew. University of Tennessee; Estados UnidosFil: Brennan, Rachel. St Jude Children's Research Hospital; Estados UnidosFil: Burges, Michala. University of Tennessee; Estados UnidosFil: Kim, Jonathan. Keck Medical School of the University of Southern California; Estados UnidosFil: Berry, Jesse L.. Children's Hospital Los Angeles; Estados UnidosFil: Jubran, Rima. Childrens Hospital Society of Los Angeles; Estados UnidosFil: Khetan, Vikas. Vitreo Retinal Services; IndiaFil: Ganeshan, Suganeswari. Vitreo Retinal Services; IndiaFil: Yarovoy, Andrey. Fyodorov Eye Microsurgery Federal State Institution; RusiaFil: Yarovaya, Vera. Fyodorov Eye Microsurgery Federal State Institution; RusiaFil: Kotova, Elena. Fyodorov Eye Microsurgery Federal State Institution; RusiaFil: Volodin, Denis. Fyodorov Eye Microsurgery Federal State Institution; RusiaFil: Yousef, Yacoub. King Hussein Cancer Center; JordaniaFil: Nummi, Kalle. University of Helsinki; Finlandia. Helsinki University Hospital; FinlandiaFil: Ushakova, Tatiana L.. N.N. Blokhin Russian Cancer Research Center; Rusia. Russian Academy of Postgraduate Medical Education; RusiaFil: Yugay, Olga V.. N.N. Blokhin Russian Cancer Research Center; RusiaFil: Polyakov, Vladimir G. N.N. Blokhin Russian Cancer Research Center; Rusia. Russian Academy of Sciences; RusiaFil: Ramirez Ortiz, Marco Antonio. Hospital Infantil de Mexico Federico Gomez; MéxicoFil: Esparza Aguiar, Elizabeth. Hospital Infantil de Mexico Federico Gomez; MéxicoFil: Chantada, Guillermo Luis. Gobierno de la Ciudad de Buenos Aires. Hospital de Pediatría "Juan P. Garrahan"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Schaiquevich, Paula Susana. Gobierno de la Ciudad de Buenos Aires. Hospital de Pediatría "Juan P. Garrahan"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Fandiño, Adriana Cristina. Gobierno de la Ciudad de Buenos Aires. Hospital de Pediatría "Juan P. Garrahan"; ArgentinaFil: Yam, Jason C.. The Chinese University of Hong Kong Faculty of Medicine; Hong Kon

Cdh11 Acts as a Tumor Suppressor in a Murine Retinoblastoma Model by Facilitating Tumor Cell Death

CDH11 gene copy number and expression are frequently lost in human retinoblastomas and in retinoblastomas arising in TAg-RB mice. To determine the effect of Cdh11 loss in tumorigenesis, we crossed Cdh11 null mice with TAg-RB mice. Loss of Cdh11 had no gross morphological effect on the developing retina of Cdh11 knockout mice, but led to larger retinal volumes in mice crossed with TAg-RB mice (p = 0.01). Mice null for Cdh11 presented with fewer TAg-positive cells at postnatal day 8 (PND8) (p = 0.01) and had fewer multifocal tumors at PND28 (p = 0.016), compared to mice with normal Cdh11 alleles. However, tumor growth was faster in Cdh11-null mice between PND8 and PND84 (p = 0.003). In tumors of Cdh11-null mice, cell death was decreased 5- to 10-fold (p<0.03 for all markers), while proliferation in vivo remained unaffected (p = 0.121). Activated caspase-3 was significantly decreased and β-catenin expression increased in Cdh11 knockdown experiments in vitro. These data suggest that Cdh11 displays tumor suppressor properties in vivo and in vitro in murine retinoblastoma through promotion of cell death

Proceedings of the Association for Research in Vision and Ophthalmology and Champalimaud Foundation Ocular Oncogenesis and Oncology Conference

Ophthalmic researc

Global Retinoblastoma Presentation and Analysis by National Income Level.

Importance: Early diagnosis of retinoblastoma, the most common intraocular cancer, can save both a child's life and vision. However, anecdotal evidence suggests that many children across the world are diagnosed late. To our knowledge, the clinical presentation of retinoblastoma has never been assessed on a global scale. Objectives: To report the retinoblastoma stage at diagnosis in patients across the world during a single year, to investigate associations between clinical variables and national income level, and to investigate risk factors for advanced disease at diagnosis. Design, Setting, and Participants: A total of 278 retinoblastoma treatment centers were recruited from June 2017 through December 2018 to participate in a cross-sectional analysis of treatment-naive patients with retinoblastoma who were diagnosed in 2017. Main Outcomes and Measures: Age at presentation, proportion of familial history of retinoblastoma, and tumor stage and metastasis. Results: The cohort included 4351 new patients from 153 countries; the median age at diagnosis was 30.5 (interquartile range, 18.3-45.9) months, and 1976 patients (45.4%) were female. Most patients (n = 3685 [84.7%]) were from low- and middle-income countries (LMICs). Globally, the most common indication for referral was leukocoria (n = 2638 [62.8%]), followed by strabismus (n = 429 [10.2%]) and proptosis (n = 309 [7.4%]). Patients from high-income countries (HICs) were diagnosed at a median age of 14.1 months, with 656 of 666 (98.5%) patients having intraocular retinoblastoma and 2 (0.3%) having metastasis. Patients from low-income countries were diagnosed at a median age of 30.5 months, with 256 of 521 (49.1%) having extraocular retinoblastoma and 94 of 498 (18.9%) having metastasis. Lower national income level was associated with older presentation age, higher proportion of locally advanced disease and distant metastasis, and smaller proportion of familial history of retinoblastoma. Advanced disease at diagnosis was more common in LMICs even after adjusting for age (odds ratio for low-income countries vs upper-middle-income countries and HICs, 17.92 [95% CI, 12.94-24.80], and for lower-middle-income countries vs upper-middle-income countries and HICs, 5.74 [95% CI, 4.30-7.68]). Conclusions and Relevance: This study is estimated to have included more than half of all new retinoblastoma cases worldwide in 2017. Children from LMICs, where the main global retinoblastoma burden lies, presented at an older age with more advanced disease and demonstrated a smaller proportion of familial history of retinoblastoma, likely because many do not reach a childbearing age. Given that retinoblastoma is curable, these data are concerning and mandate intervention at national and international levels. Further studies are needed to investigate factors, other than age at presentation, that may be associated with advanced disease in LMICs