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DecisionDx®-UMSeq Summary

Next-Generation Sequencing for Uveal Melanoma

[BOX]Comprehensive genomic profile of an individual uveal melanoma (UM) tumor from a single biopsy

The DecisionDx-UMSeq test is a 7-gene panel that uses next-generation sequencing (NGS) to identify somatic mutations relevant to UM. The test includes hotspot mutations in the genes GNAQGNA11CYSLTR2PLCB4, and SF3B1, mutations in exons 1-2 of EIF1AX, and all coding exons of the BAP1 gene.

 

This information, together with results from the DecisionDx®-UM and DecisionDx®-PRAME gene expression profile (GEP) tests, can help to build a comprehensive genomic profile of an individual UM tumor from a single biopsy. The genomic information can be used now to inform patient care and may become useful in the future as research and therapeutics evolve.

 

DecisionDx-UMSeq can be ordered for patients who are having DecisionDx-UM GEP testing performed. Castle Biosciences has optimized the sequencing so that it can be run using the same fine-needle aspiration biopsy (FNAB) tissue specimen submitted for GEP testing.

 

Mutations in 4 genes affect G-protein-coupled receptor signaling

In UM, mutations in GNAQGNA11CYSLTR2, or PLCB4 result in constitutive activation of G-protein-coupled receptor signaling pathways, such as MAPK, PI3K, PKC, Hippo, etc. (Figure 1) (Van Raamsdonk, 2009; 2011; Johansson, 2016; Moore, 2016).

While some of these pathways may be targeted with inhibitory therapies, these mutations are not currently markers for drug response. These mutations are also not known to be prognostic.

 

The majority of UM tumors will have a mutation in GNAQ or GNA11, while CYSLTR2 and PLCB4 mutations are less common. These mutations are usually mutually exclusive. The presence of one of these mutations may provide some confidence that a melanocytic tumor was sampled. However, other eye lesions can have these mutations, and some UM tumors will not have a mutation in any of these 4 genes. These mutations should not be used exclusively to rule-in or rule-out a UM diagnosis in the absence of other clinical/pathological features.

 

More on GNAQ         More on GNA11         More on CYSLTR2         More on PLCB4

 

Mutations in 3 genes are associated with differential clinical outcomes

Mutations occurring in EIF1AXSF3B1, and BAP1 are usually mutually exclusive:

  • The EIF1AX protein is involved in translation (from mRNA to protein) initiation
  • SF3B1 is a component of the spliceosome, which regulates transcript usage
  • BAP1 is a tumor suppressor gene on chromosome 3 that encodes for a deubiquitinating enzyme that forms protein complexes with BRCA1, BARD1, and ASXL1

 

It is well known that mutations in BAP1 in UM are associated with a higher risk of metastasis and there is considerable overlap with a Class 2 GEP (Harbour, 2010). Some BAP1 mutations are germline and can be inherited (Abdel-Rahman, 2011; Gupta, 2015), but the majority are somatic and thus confined to the tumor.

 

In retrospective studies, having an EIF1AX or SF3B1 mutation has been shown to be associated with a better prognosis than having a BAP1 mutation (Martin, 2013; Ewens, 2014; Decatur, 2016; Furney, 2013; Harbour, 2013; Yavuzyigitoglu, 2016). Tumors with EIF1AX mutations may have the lowest risk of metastasis compared to those with SF3B1 mutations, which may be prone to late metastasis.

 

Importantly, in a recent retrospective study comparing the prognostic value of these 3 mutations when using the DecisionDx-UM GEP test, a Class 2 result was the only significant, independent predictor of metastasis and UM-related mortality in multivariate analysis (Decatur, 2016). Therefore, the GEP test currently provides the best prognostic information about the tumor.

 

More on EIF1AX                             More on SF3B1                             More on BAP1

 

Ordering DecisionDx-UMSeq

DecisionDx-UMSeq is available for patients undergoing DecisionDx-UM GEP testing or those who have already received their GEP results. DecisionDx-UMSeq can be run using the same biopsy taken for the DecisionDx-UM GEP test.

 

A separate DecisionDx-UMSeq requisition form must be completed.

 

Castle Biosciences will  submit the cost for the DecisionDx-UMSeq to your patient’s insurance  and will track the claim on behalf of your patient throughout the process. Castle offers an industry-leading financial assistance program for both insured and uninsured patients. For more information regarding billing or for information on assisting your patients to qualify for financial assistance call 866-788-9007 and select option #3.

 

Understanding the DecisionDx-UMSeq Results

The DecisionDx-UMSeq test results can be expected in 2-4 weeks after receipt of the sample in our laboratory.

 

The DecisionDx-UMSeq report will tell you if clinically relevant mutations (variants) were found in any of the 7 gene targets. For each mutation found, the report will describe:

  • Genomic location of the mutation (where in the gene it occurred)
  • Type of mutation (eg, missense, nonsense)
  • Functional change that occurs because of the mutation (ie, an amino acid change in the protein)
  • Frequency that the mutation was detected in the sample (variant allele frequency)
  • Potential consequences of that mutation on gene function, as well as relevant literature references

 

The Tier and Level of Evidence will also be described for each mutation, as recommended by the College of American Pathologists (CAP), the American Society of Clinical Oncology (ASCO), and the Association for Molecular Pathologists (AMP) [Li, 2017].

 

BAP1 germline testing information

If your patient has a detected BAP1 mutation and is interested in germline testing, Castle Biosciences can provide information to assist with finding testing services and genetic counseling. Please call Castle Biosciences at 866-788-9007, option 1 for these resources.

 

References

Abdel-Rahman MH, Pilarski R, Cebulla CM, et al. Germline BAP1 mutation predisposes to uveal melanoma, lung adenocarcinoma, meningioma, and other cancers. J Med Genet. 2011;48:856-859.

Aoude LG, Vajdic CM, Kricker A, et al. Prevalence of germline BAP1 mutation in a population-based sample of uveal melanoma cases. Pigment Cell Melanoma Res. 2013;26:278-279.

Decatur CL, Ong E, Garg N, et al. Driver mutations in uveal melanoma: associations with gene expression profile and patient outcomes. JAMA Ophthalmol. 2016;134:728-733.

Ewens KG, Kanetsky PA, Richards-Yutz J, et al. Chromosome 3 status combined with BAP1 and EIF1AX mutation profiles are associated with metastasis in uveal melanoma. Invest Ophthalmol Vis Sci. 2014;55:5160-5167.

Furney SJ, Pedersen M, Gentien D, et al. SF3B1 mutations are associated with alternative splicing in uveal melanoma. Cancer Discov. 2013;3:1122-1129.

Gupta MP, Lane AM, DeAngelis MM, et al. Clinical characteristics of uveal melanoma in patients with germline BAP1 mutations. JAMA Ophthalmol. 2015;133:881-887.

Harbour JW, Onken MD, Roberson ED, et al. Frequent mutation of BAP1 in metastasizing uveal melanomas. Science. 2010;330:1410-1413.

Harbour JW, Roberson ED, Anbunathan H, et al. Recurrent mutations at codon 625 of the splicing factor SF3B1 in uveal melanoma. Nat Genet. 2013;45:133-135.

Johansson P, Aoude LG, Wadt K, et al. Deep sequencing of uveal melanoma identifies a recurrent mutation in PLCB4. Oncotarget. 2016;7:4624-4631.

Martin M, Masshofer L, Temming P, et al. Exome sequencing identifies recurrent somatic mutations in EIF1AX and SF3B1 in uveal melanoma with disomy 3. Nat Genet. 2013;45:933-6

Moore AR, Ceraudo E, Sher JJ, et al. Recurrent activating mutations of G-protein-coupled receptor CYSLTR2 in uveal melanoma. Nat Genet. 2016;48:675-680.

Njauw CN, Kim I, Piris A, et al. Germline BAP1 inactivation is preferentially associated with metastatic ocular melanoma and cutaneous-ocular melanoma families. PLoS One. 2012;7:e35295.

Onken MD, Worley LA, Long MD, et al. Oncogenic mutations in GNAQ occur early in uveal melanoma. Invest Ophthalmol Vis Sci. 2008;49:5230-5234.

Rai K, Pilarski R, Cebulla CM, et al. Comprehensive review of BAP1 tumor predisposition syndrome with report of two new cases. Clin Genet. 2016;89:285-294.

Shoushtari AN and Carvajal RD. GNAQ and GNA11 mutations in uveal melanoma. Melanoma Res. 2014;24:525-534.

Van Raamsdonk CD, Bezrookove V, Green G, et al. Frequent somatic mutations of GNAQ in uveal melanoma and blue naevi. Nature. 2009;457:599-602.

Van Raamsdonk CD, Griewank KG, Crosby MB, et al. Mutations in GNA11 in uveal melanoma. N Engl J Med. 2010;363:2191-2199.

Yavuzyigitoglu S, Koopmans AE, Verdijk RM, et al. Uveal melanomas with SF3B1 Mutations: a distinct subclass associated with late-onset metastases. Ophthalmology. 2016;123:1118-1128.

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