There is now evidence that gene fusions activating the MAPK pathway are relatively common in pancreatic acinar cell carcinoma with potentially actionable BRAF or RET fusions being found in ~30%. We ...sought to investigate the incidence of RAF1 fusions in pancreatic malignancies with acinar cell differentiation. FISH testing for RAF1 was undertaken on 30 tumors comprising 25 ‘pure’ acinar cell carcinomas, 2 mixed pancreatic acinar-neuroendocrine carcinomas, 1 mixed acinar cell-low grade neuroendocrine tumor and 2 pancreatoblastomas. RAF1 rearrangements were identified in 5 cases and confirmed by DNA and RNA sequencing to represent oncogenic fusions (GATM-RAF1, GOLGA4-RAF1, PDZRN3-RAF1, HERPUD1-RAF1 and TRIM33-RAF1) and to be mutually exclusive with BRAF and RET fusions, as well as KRAS mutations. Large genome-wide copy number changes were common and included 1q gain and/or 1p loss in all five RAF1 FISH-positive acinar cell carcinomas. RAF1 expression by immunohistochemistry was found in 3 of 5 (60%) of fusion-positive cases and no FISH-negative cases. Phospho-ERK1/2 expression was found in 4 of 5 RAF1-fusion-positive cases. Expression of both RAF1 and phospho-ERK1/2 was heterogeneous and often only detected at the tumor-stroma interface, thus limiting their clinical utility. We conclude that RAF1 gene rearrangements are relatively common in pancreatic acinar cell carcinomas (14.3% to 18.5% of cases) and can be effectively identified by FISH with follow up molecular testing. The combined results of several studies now indicate that BRAF, RET or RAF1 fusions occur in between one third and one-half of these tumors but are extremely rare in other pancreatic malignancies. As these fusions are potentially actionable with currently available therapies, a strong argument can be made to perform FISH or molecular testing on all pancreatic acinar cell carcinomas.
Abstract
Whilst the majority of Endometrial Cancers (ECs) are common Type 1 endometrioid cancers, accounting for 75-80% of cases, with a good prognosis, Type 2 ECs include high-grade, clinically ...aggressive histologies, with poor response rates to hormonal therapies. Serous endometrial carcinoma is the second most common type, accounting for ~10 percent of cases, most with a p53 abnormality and a lesser proportion with HER2 overexpression/amplification/mutation. Clear cell endometrial carcinoma accounts for <5 % of EC, with the most aggressive cases also having p53 mutations. Mixed histologies and undifferentiated ECs are also aggressive. Carcinosarcoma (ECS) is a rare, aggressive, biphasic carcinoma that accounts for <5 percent of ECs, 90% with p53 abnormalities. Endometrial stromal sarcomas (ESS) can be low-grade or high-grade, or undifferentiated or resemble ovarian sex cord tumors. Uterine leiomyosarcoma (uLMS) are epithelioid or myxoid in type. Adenosarcoma involves a benign epithelial component mixed with a malignant stromal element. Whilst there have been considerable improvements in death rates for all cancers combined over the last 20 years, these improvements have not been seen for most rare cancers (RC). In order to ensure that those diagnosed with RC have access to research and novel therapies, we designed the Walter and Eliza Hall Institute of Medical Research (WEHI) Stafford Fox Rare Cancer Program (SFRCP). The WEHI-SFRCP has streamlined ethics, governance, consenting processes, including remote consent at home anywhere in Australia, and data collection protocols to allow the analysis of data and tissue of any type of RC. We have interconnected clinical and laboratory RC Databases within BioGrid Australia using the online REDCap platform. We have extensive laboratory processes in place for processing of tumour and blood samples, including the generation of PBMCs, DNA, RNA, to generate NGS including WGS; patient-derived xenografts (PDX), organoids, cell lines and other derivatives. We have high grade serous endometrial cancer (33 cases, including 4 PDX with 3 pending); uLMS (32 cases, including 2 PDX with 3 pending); endometrial carcinosarcoma (13 cases, including 3 PDX, 1 pending); rare (other, adenosarcoma, STUMP). We perform NGS testing on cases, depending on tumour purity, including Whole Genome Sequencing from fresh tumour samples; characterize PDX according to current chemotherapy and relevant novel therapeutics; study rare endometrial subtypes in specific projects and also provide information back to patients to guide therapy in the clinic. By integrating data sets with endometrial projects of similar depth we will drive forward the study of rare endometrial cancer subtypes.
Citation Format: Clare L. Scott, Ratana Lim, Amandine Carmagnac, Cassandra Vandenberg, Gayanie Ratnayake, Genevieve Dall, Joshua Tram, Justin Bedo, Jocelyn Penington, Joep Vissers, Sean Grimmond, Matthew Wakefield, Anthony Papenfuss, Holly Barker. Building the infrastructure required to research novel therapies for 35% of women with endometrial cancer who have rare subtypes abstract. In: Proceedings of the AACR Virtual Special Conference: Endometrial Cancer: New Biology Driving Research and Treatment; 2020 Nov 9-10. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(3_Suppl):Abstract nr PO026.
Abstract
High-grade serous endometrial carcinoma (HGSEC) accounts for just 10% of endometrial cancer (EC) cases but is responsible for at least 40% of all EC-related deaths. It typically arises in ...post-menopausal women, with 70% of patients presenting with stage III or IV disease, does not respond to hormone therapy unlike the less aggressive forms of EC, and has a lower overall survival rate of just 18-27%, which has not improved over the past two decades. The primary treatment for HGSEC is surgery, followed by a combination of standard chemotherapies (platinum and taxane) with or without localised radiotherapy. However, recurrent HGSEC is less responsive to chemotherapy than are other subtypes of EC and even initial responses to chemotherapy are poor. Therefore, there is a great unmet clinical need to find better treatment options for women with this aggressive cancer. Apart from TP53 (mutated in up to 90% of cases), the other most frequently mutated genes in HGSEC are PPP2R1A (31%), PIK3CA (22%), FBXW7 (28%), CHD4 (17%) and BRCA2 (12%). Focal amplifications of the genes MYC, ERBB2, CCNE1, FGFR3 and SOX17 are also common. The presence of ERBB2 amplification and/or HER2 over-expression in around 30% of HGSEC suggests these patients may respond to HER2-targeting drugs, such as trastuzumab. However, only modest benefit has so far been seen for single-agent HER2-targeted therapies (ie trastuzumab or lapatinib) against HGSEC, suggesting resistance mechanisms are present. Another feature of HGSEC that could be exploited therapeutically is homologous recombination deficiency (HRD), which may be targeted with PARP inhibitors (PARPi). It is not clear what proportion of HGSEC are HRD and neither HER2-targeting drugs or PARPi have been approved for the treatment of HGSEC. Due to its rarity and a lack of pre-clinical models, HGSEC has so far been understudied, resulting in a lack of effective treatment options. We currently have 33 HGSEC patients consented to the WEHI-Stafford Fox Rare Cancer Program and have developed pre-clinical models from fresh patient tumour samples received (4 patient-derived xenograft (PDX) models validated, with 3 pending). Preliminary molecular analysis of whole-genome sequencing (5 samples, one of which gave rise to a PDX model), whole-exome sequencing (4 samples), and cancer panel sequencing (3 samples, 2 of which gave rise to PDX models; one harbouring ERBB2 amplification and one harbouring an AKT mutation) data from our HGSEC cohort has been performed. This has identified potential treatment targets, including ERBB2 amplifications and mutations in HR genes. I am using the PDX models for initial in vivo therapeutic characterization studies and to develop organoid models for use in high-throughput drug assays in vitro. This will guide subsequent novel drug combination testing in our PDX models. By combining specific targeted drugs I hope to overcome de novo resistance mechanisms and prevent acquired resistance. Results from this study will guide future decisions about therapeutic strategies to improve survival of women with HGSEC.
Citation Format: Holly E. Barker, Ratana Lim, Amandine Carmagnac, Cassandra Vandenberg, Gayanie Ratnayake, Genevieve Dall, Briony Milesi, Angela Komiti, Emily O'Grady, Joshua Tram, Kym Pham Stewart, Justin Bedo, Jocelyn Penington, Joep Vissers, Sean Grimmond, Matthew Wakefield, Tony Papenfuss, Clare Scott. Identifying effective combinations of targeted therapies, using novel pre-clinical models, to improve treatment options for high-grade serous endometrial cancer abstract. In: Proceedings of the AACR Virtual Special Conference: Endometrial Cancer: New Biology Driving Research and Treatment; 2020 Nov 9-10. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(3_Suppl):Abstract nr PO037.
Abstract
Uterine sarcomas make up 1-4% of uterine malignancies. Of these 60% are classified as leiomyosarcoma (uLMS). The 5-year survival rate of uLMS is 35-65.2% for tumours that have not spread ...beyond the uterus. However, women are often diagnosed at a late stage due to a lack of screening options by which time the tumour has often spread to adjacent and distant tissues. Current standard of care for uLMS patients is surgical de-bulking followed by adjuvant chemotherapy, but significant improvement in progression free survival and overall survival is not consistently observed. The lack of advances for the treatment and screening of uLMS is due in part to the scarcity of appropriate research resources for this rare disease. Genetic analyses have been performed but on relatively small samples and there are just 14 reported patient-derived xenograft (PDX) models of uLMS in the literature to date. Through the WEHI Stafford Fox Rare Cancer Program, as well as collaborations (facilitated by ANZGOG) throughout the country and internationally, we have access to a large biobank of uLMS tissue. We have received 8 fresh uLMS samples in the laboratory, 2 of which have established PDX lines that were validated as uLMS by our anatomical pathologist. All fresh samples received into the laboratory are snap frozen for whole-genome sequencing as well as viably frozen to enable regeneration of the tissue for future applications. One application is organoid culturing, which allows the tissue to retain its 3D growth properties and is significantly cheaper than growing tissue as a PDX. Organoid culturing also allows for higher throughput of samples in drug screening assays, enabling us to fast-track the selection of drugs for validation in our PDX models. In addition to these fresh samples we also have 23 archival uLMS samples (formalin fixed, paraffin embedded) that can be used for lower coverage genetic analysis, and protein expression by immunohistochemistry. This unique biobank of uLMS tissue is the first of its kind in Australia and with it we will endeavour to gain a comprehensive understanding of this disease. Through our PDX modelling we also have the opportunity to predict resistance to therapy and test emerging therapies in a clinically relevant context. We believe this biobank will provide a critical resource which will ultimately lead to better outcomes for uLMS patients.
Citation Format: Genevieve Dall, Cassandra Vandenberg, Amandine Carmagnac, Ratana Lim, Briony Milesi, Angela Komiti, Emily O'Grady, Joshua Tram, Gayanie Ratnayake, Kym Pham Stewart, Justin Bedo, Jocelyn Penington, Joep Vissers, Inger Olesen, Sean Grimmond, Holly Barker, Tony Papenfuss, Clare Scott. Developing pre-clinical models of uterine leiomyosarcoma abstract. In: Proceedings of the AACR Virtual Special Conference: Endometrial Cancer: New Biology Driving Research and Treatment; 2020 Nov 9-10. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(3_Suppl):Abstract nr PO021.
In this study, exome sequencing yielded a genetic diagnosis in 16% of patients who had previously been evaluated to rule out known causes of intellectual disability.
Severe intellectual disability, ...which is also referred to as cognitive impairment or mental retardation, affects approximately 0.5% of the population in Western countries
1
,
2
and represents an important health burden. A clinical diagnosis of severe intellectual disability is generally based on an IQ of less than 50 and substantial limitations in activities of daily living. In early childhood, the diagnosis is based on substantial developmental delays, including motor, cognitive, and speech delays. Children with different nonsyndromic forms of intellectual disability are clinically indistinguishable.
Intellectual disability can be caused by nongenetic factors, such as infections and perinatal asphyxia. In developed countries, . . .
Family-based next generation sequencing (NGS) has recently pointed to an important role for de novo germline point mutations in both rare and common genetic disorders associated with reduced fitness. ...In this review we highlight the impact of the mutational target size on the frequency of diseases caused by these de novo point mutations. In addition, we will discuss the human per-generation mutation rate, its relation to advanced paternal age and how these factors affect the frequency of genetic disease caused by de novo events.
In a predominantly biomedical healthcare model focused on cure, providing optimal, person-centred palliative care is challenging. The general public, patients, and healthcare professionals are often ...unaware of palliative care's benefits. Poor interdisciplinary teamwork and limited communication combined with a lack of early identification of patients with palliative care needs contribute to sub-optimal palliative care provision. We aimed to develop a national quality framework to improve availability and access to high-quality palliative care in a mixed generalist-specialist palliative care model. We hypothesised that a whole-sector approach and a modified Delphi technique would be suitable to reach this aim. Analogous to the international AGREE guideline criteria and employing a whole-sector approach, an expert panel comprising mandated representatives for patients and their families, various healthcare associations, and health insurers answered the main question: 'What are the elements defining high-quality palliative care in the Netherlands?'. For constructing the quality framework, a bottleneck analysis of palliative care provision and a literature review were conducted. Six core documents were used in a modified Delphi technique to build the framework with the expert panel, while stakeholder organisations were involved and informed in round-table discussions. In the entire process, preparing and building relationships took one year and surveying, convening, discussing content, consulting peers, and obtaining final consent from all stakeholders took 18 months. A quality framework, including a glossary of terms, endorsed by organisations representing patients and their families, general practitioners, elderly care physicians, medical specialists, nurses, social workers, psychologists, spiritual caregivers, and health insurers was developed and annexed with a summary for patients and families. We successfully developed a national consensus-based patient-centred quality framework for high-quality palliative care in a mixed generalist-specialist palliative care model. A whole-sector approach and a modified Delphi technique are feasible structures to achieve this aim. The process we reported may guide other countries in their initiatives to enhance palliative care.
A de novo paradigm for mental retardation Brunner, Han G; Veltman, Joris A; Vissers, Lisenka E L M ...
Nature genetics,
12/2010, Letnik:
42, Številka:
12
Journal Article
Recenzirano
The per-generation mutation rate in humans is high. De novo mutations may compensate for allele loss due to severely reduced fecundity in common neurodevelopmental and psychiatric diseases, ...explaining a major paradox in evolutionary genetic theory. Here we used a family based exome sequencing approach to test this de novo mutation hypothesis in ten individuals with unexplained mental retardation. We identified and validated unique non-synonymous de novo mutations in nine genes. Six of these, identified in six different individuals, are likely to be pathogenic based on gene function, evolutionary conservation and mutation impact. Our findings provide strong experimental support for a de novo paradigm for mental retardation. Together with de novo copy number variation, de novo point mutations of large effect could explain the majority of all mental retardation cases in the population.
Invertebrate model systems are powerful tools for studying human disease owing to their genetic tractability and ease of screening. We conducted a mosaic genetic screen of lethal mutations on the ...Drosophila X chromosome to identify genes required for the development, function, and maintenance of the nervous system. We identified 165 genes, most of whose function has not been studied in vivo. In parallel, we investigated rare variant alleles in 1,929 human exomes from families with unsolved Mendelian disease. Genes that are essential in flies and have multiple human homologs were found to be likely to be associated with human diseases. Merging the human data sets with the fly genes allowed us to identify disease-associated mutations in six families and to provide insights into microcephaly associated with brain dysgenesis. This bidirectional synergism between fly genetics and human genomics facilitates the functional annotation of evolutionarily conserved genes involved in human health.
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•A Drosophila resource of mutants affecting adult neural development and maintenance•Essential fly genes with multiple human homologs are often associated with disease•Variants in human homologs of these genes are associated with Mendelian disease•ANKLE2 is associated with small brain size and microcephaly in fly and man
Combining datasets from a Drosophila resource of mutants with neurodevelopmental or -degenerative phenotypes with human exomes of patients with rare Mendelian disorders provides new insights into human neurodevelopmental diseases.
ABSTRACT
Copy number variation (CNV) is a common source of genetic variation that has been implicated in many genomic disorders. This has resulted in the widespread application of genomic microarrays ...as a first‐tier diagnostic tool for CNV detection. More recently, whole‐exome sequencing (WES) has been proven successful for the detection of clinically relevant point mutations and small insertion–deletions exome wide. We evaluate the utility of short‐read WES (SOLiD 5500xl) to detect clinically relevant CNVs in DNA from 10 patients with intellectual disability and compare these results to data from two independent high‐resolution microarrays. Eleven of the 12 clinically relevant CNVs were detected via read‐depth analysis of WES data; a heterozygous single‐exon deletion remained undetected by all algorithms evaluated. Although the detection power of WES for small CNVs currently does not match that of high‐resolution microarray platforms, we show that the majority (88%) of rare coding CNVs containing three or more exons are successfully identified by WES. These results show that the CNV detection resolution of WES is comparable to that of medium‐resolution genomic microarrays commonly used as clinical assays. The combined detection of point mutations, indels, and CNVs makes WES a very attractive first‐tier diagnostic test for genetically heterogeneous disorders.
Copy number variation (CNV) is a common source of genetic variation implicated in many genomic disorders. We evaluate the utility of WES to detect clinically relevant CNVs in DNA from 10 patients with intellectual disability and compare these results to data from two independent high‐resolution microarrays. Of the 12 clinically relevant CNVs 11 were detected via read‐depth analysis of WES‐data. The combined detection of small mutations and CNVs makes WES an attractive diagnostic test for genetically heterogeneous disorders.
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