BrainTumorNext is a next generation sequencing panel that simultaneously analyzes 27 genes associated with increased risk for brain tumors and other cancers/tumors.
BrainTumorNext is a next generation sequencing panel that simultaneously analyzes 27 genes associated with increased risk for brain tumors and other cancers/tumors.
Ambry utilizes next generation sequencing (NGS) to offer a comprehensive panel for hereditary brain tumors. Genes on this panel include: AIP, ALK, APC, CDKN1B, CDKN2A, DICER1, MEN1, MLH1, MSH2, MSH6, NBN, NF1, NF2, PHOX2B, PMS2, POT1, PRKAR1A, PTCH1, PTEN, SMARCA4, SMARCB1, SMARCE1, SUFU, TP53, TSC1, TSC2, and VHL. Full gene sequencing and gross deletion/duplication analysis is performed for all 27 genes. Specific Site Analysis is available for individual gene mutations identified in a family.
Brain tumors are the 16th most common cancer diagnosed in men and women. Fewer than 1% of men and women in the U.S. will be diagnosed with a brain tumor/cancer at some point during their lives.1 The National Cancer Institute (NCI) estimates that approximately 23,700 new cases of brain and other nervous system tumors/cancer will be diagnosed in the U.S. in 2016.2 The median age at diagnosis is age 58, although 43.6% are diagnosed under age 54.2 The majority of brain tumors are sporadic, with some being hereditary and developing due to an inherited genetic mutation. Hereditary brain tumors may be diagnosed at younger ages (especially in childhood).
AIP mutations cause familial isolated pituitary adenomas (FIPA), an autosomal dominant condition. These occur in a familial setting in the absence of multiple endocrine neoplasia 1 (MEN1), Carney complex, or other known inherited conditions. FIPA differs from MEN1 in terms of a lower proportion of prolactinomas and more frequent somatotropinomas in the FIPA cohort.3 Mutations in the AIP gene have been identified in 15-30% of cases of FIPA and have also been reported in individuals with sporadic pituitary adenomas.3,4 Compared to AIP mutation negative individuals with pituitary adenomas, individuals with mutations are more likely to have aggressive disease and present earlier in life.3 The median age of diagnosis of AIP-related FIPA is 23 years and penetrance estimates range from 33-66%.3,5
ALK heterozygous germline mutations in the ALK gene are associated with familial predisposition to neuroblastic tumors including neuroblastoma, ganglioneuroblastoma, and ganglioneuroma.6 Germline mutations in ALK have also been implicated in medulloblastoma risk, but further study is necessary.7 Penetrance for neuroblastoma is variable, and is estimated to be up to 57% across studies.8
APC germline mutations are the primary cause of familial adenomatous polyposis (FAP) and attenuated familial adenomatous polyposis (AFAP). FAP and AFAP are autosomal dominant colon cancer predisposition syndromes characterized by hundreds to thousands of adenomatous polyps in the internal lining of the colon and the rectum. They affect 1 in 8,000 to 1 in 10,000 individuals, and account for about 1% of all colorectal cancers.9 In individuals affected with classic FAP, colonic polyps generally begin developing at an average age of 16 years.10 In these families, colon cancer is inevitable without surgical intervention like colectomy, and the mean age of colon cancer diagnosis in untreated individuals is 35-40 years.11 Individuals with FAP or AFAP may also have increased risks to develop duodenal cancer, pancreatic cancer, papillary thyroid cancer, hepatoblastoma in childhood, and medulloblastoma. Some individuals may also have non-malignant features such as osteomas, congenital hypertrophy of the retinal pigment epithelium (CHRPE), and/or desmoid tumors.9
CDKN1B Heterozygous germline alterations are associated with multiple endocrine neoplasia type 4 (MEN4), characterized by parathyroid, anterior pituitary, and neuroendocrine tumors.12 Nearly 100% individuals with CDKN1B mutations develop hyperparathyroidism, however, penetrance estimates for other features of this syndrome are not currently available.13 MEN4 and multiple endocrine neoplasia type 1 share similar phenotypes, and mutations in CDKN1B account for approximately 1-3% of individuals with clinically diagnosed MEN1 lacking a germline mutation in the MEN1 gene.12,14
CDKN2A encodes two distinct proteins, p16 and p14ARF, which are both involved in cell cycle regulation. Germline p16/CDKN2A mutations are associated with familial atypical multiple mole melanoma (FAMMM) syndrome. FAMMM is an autosomal dominant disorder characterized by an increased risk for atypical mole malignant melanoma, often associated with dysplastic or atypical nevi.CDKN2A mutation carriers have an approximate 28-67% lifetime risk of developing melanoma, with penetrance estimates varying widely based on study design and geographic region.15-17 Individuals carrying CDKN2A mutations also have an approximate 17-25% lifetime risk for pancreatic cancer; however, recent reports suggest this risk may be as high as 58% and elevated further in smokers.18-20 Rare mutations that affect the p14ARF mutations have also been reported to predispose to melanoma and possibly pancreatic cancer.19,21,22
DICER1 mutations have been shown to cause a tumor predisposition syndrome associated with an increased risk for various benign and malignant tumors. Studies have demonstrated an increased risk for tumors including pleuropulmonary blastoma, cystic nephroma, ovarian sex cord stromal tumors (primarily Sertoli-Leydig cell tumors), multinodular goiter and thyroid cancer, embryonal rhabdomyosarcomas, ciliary body medulloepithelioma, nasal chondromesenchymal hamartomas, and pituitary blastoma, as well as various other tumor types. At this time, lifetime risks for each tumor type have not been well described.23-25
MEN1 mutations cause multiple endocrine neoplasia type 1 (MEN1) and familial isolated hyperparathyroidism (FIHP). FIHP is defined by primary hyperparathyroidism as the sole endocrinopathy in a family.26 In contrast, MEN1 is characterized by primary hyperparathyroidism due to parathyroid adenomas (present in over 90% of affected individuals), gastro-entero-pancreatic neuroendocrine tumors (in 30-70%), pituitary adenomas (in 30-60%), adrenocortical tumors (15-50%), bronchial and thymic carcinoids (up to 10%), facial angiofibromas, collagenomas, and lipomas.27-32 The majority of patients (94%) carrying a mutation in the MEN1 gene exhibit clinical or biochemical symptoms by age 50.33
MLH1, MSH2, MSH6, and PMS2 germline mutations are associated with Lynch syndrome (previously called hereditary non-polyposis colorectal cancer, HNPCC). Lynch syndrome is an autosomal dominant condition estimated to cause 2-5% of all colorectal cancer. It is associated with a significantly increased risk for colorectal cancer (up to 82% lifetime risk), uterine/endometrial cancer (25-60% lifetime risk in women), stomach cancer (6-13% lifetime risk), ovarian cancer (4-12% lifetime risk in women) and prostate cancer (up to 2 fold). Risk for cancer of the small bowel, hepatobiliary tract, upper urinary tract (including transitional cell carcinoma of the renal pelvis), brain, and sebaceous glands may also be elevated.34-38
NBN gene is involved in the Fanconi anemia (FA)-BRCA pathway, critical for DNA repair by homologous recombination, and interact in vivo with BRCA1 and/or BRCA2.39-41 Mutations in these genes are associated with an increased risk for female breast and ovarian cancer.41,43 NBN has more recently been associated an increased risk of prostate cancer and medulloblastoma.44-46 NBN is also associated with a rare autosomal recessive disorder that affects multiple body systems.
NF1 mutations cause neurofibromatosis type 1 (NF1), an autosomal dominant disorder affecting multiple body systems. It is characterized by multiple café-au-lait spots, axillary and inguinal freckling, multiple cutaneous neurofibromas, and Lisch nodules. The most common neoplasms observed in individuals with NF1 include peripheral nerve sheath tumors, gastrointestinal stromal tumors (GIST), central nervous system gliomas, leukemias, paragangliomas (PGLs) and pheochromocytomas (PCCs), and breast cancer. Multiple population-based studies have demonstrated a 3 to 5-fold increase in lifetime breast cancer risk for women with NF1, with the highest risks for those less than 50 years of age.47 In addition, individuals with NF1 have an estimated lifetime risk for PGLs and PCCs of up to 7%.100
NF2 heterozygous germline pathogenic mutations cause neurofibromatosis type 2 (NF2), characterized by development of multiple nerve sheath tumors, most notably bilateral vestibular schwannomas (present in over 90% of individuals), as well schwannomas of other cranial nerves (in 24-51%), intracranial meningiomas (in 45-58%), and spinal tumors including meningiomas, schwannomas, gliomas, ependymomas, and rarely astrocytomas (combined 63-90%).48 Approximately 50% of all NF2 mutations are inherited while the remaining 50% are de novo events. Of the NF2 mutations that are de novo events, up to 30% are mosaic.49
PHOX2B functions as a transcription factor involved in the development of several major noradrenergic neuron populations and the determination of neurotransmitter phenotype. Expansion of a 20 amino acid polyalanine tract in this protein by 5-13 aa has been associated with congenital central hypoventilation syndrome (CCHS), characterized by apparent hypoventilation due to autonomic nervous system dysregulation (ANSD) as well as abnormalities of neural crest-derived structures, such as Hirschsprung disease (HSCR), and tumors of neural crest origin.50 Heterozygous germline mutations outside the polyalanine repeat region predispose to neuroblastic tumors such as neuroblastoma, ganglioneuroblastoma, and ganglioneuroma, with or without other neurocristopathies of CCHS and HSCR.51 Genotype-phenotype correlations are under study to help better define disease mechanisms and the extent of overlap of the various phenotypes associated with PHOX2B mutations.52
POT1 heterozygous germline alterations have been identified in familial and unselected cases of glial tumors such as glioma, astrocytoma, and oligodendroglioma, and appear to demonstrate incomplete penetrance.53,54 Lifetime cancer risk estimates for POT1 mutation carriers are not currently available.
PRKAR1A heterozygous germline pathogenic mutations cause Carney Complex, which is characterized by primary pigmented nodular adrenocortical disease (PPNAD)(present in 26-60% of individuals), pituitary adenoma (in 10-12%), cardiac myxoma (in 32-53%), skin and breast myxomas (in 20-33%), thyroid nodules (in 25%) and/or carcinoma (in 2-5%), large-cell calicifying sertoli cell tumor (in 33-41%), and psammomatous melanotic schwannomas (in 8-10%). 55-58
PTCH1 mutations cause nevoid basal cell carcinoma syndrome (NBCCS), also referred to as Gorlin syndrome. NBCCS/Gorlin syndrome is an autosomal dominant condition with a de novo mutation rate of approximately 20-30%, and up to 90% of affected individuals develop basal cell carcinoma. In addition, NBCCS can cause odontogenic keratocysts, congenital skeletal anomalies, cerebral calcifications, macrocephaly, polydactyly, intellectual disability, palmar epidermal pits, and cardiac and ovarian fibromas.59-63 Up to 5% of children with NBCCS develop medulloblastoma, also called primitive neuroectodermal tumor (PNET), most often the desmoplastic subtype. The diagnosis of NBCCS has historically been made based on clinical features.
PTEN is a gene associated with Cowden syndrome (CS), PTEN hamartoma tumor syndrome (PHTS), Bannayan-Riley-Ruvalcaba syndrome, Proteus syndrome, and autism spectrum disorder. CS is a multiple hamartoma syndrome with a high risk of developing tumors of the thyroid, breast, and endometrium. Mucocutaneous lesions, thyroid abnormalities, fibrocystic disease, multiple uterine leiomyomata, and macrocephaly can also be seen. Affected individuals have a lifetime risk of up to 50% for breast cancer, 10% for thyroid cancer, and 5-10% for endometrial cancer. Over 90% of individuals with CS will express some clinical manifestations by their twenties.64,65 Recent studies noted increased risks for renal cell cancer, colorectal cancer, and other cancers.66,67 One study quotes up to a 31-fold increase in RCC risk for PTEN mutation carriers as compared to the general population.68
SMARCA4 truncating mutations cause rhabdoid tumor predisposition syndrome type 2. SMARCA4-associated tumors are highly aggressive and include atypical teratoid/rhabdoid tumors (AT/RT) of the central nervous system, malignant rhabdoid tumors of the kidney, and small cell carcinoma of the ovary, hypercalcemic type (SCCOHT). The lifetime cancer risks for SMARCA4 mutation carriers has yet to be defined; however, age of onset and penetrance are extremely variable, with some carriers presenting prenatally while others remain unaffected through adulthood.69-72
SMARCB1 heterozygous germline pathogenic mutations cause rhabdoid tumor predisposition syndrome type 1 (RTPS1) and schwannomatosis. Individuals with RTPS1 are at risk to develop atypical teratoid/rhabdoid tumors of the CNS and malignant rhabdoid tumors of the kidney.73 Age of onset and penetrance are extremely variable, with some carriers presenting at birth while others remain unaffected through adulthood.74 Approximately 5% of individuals with schwannomatosis due to SMARCB1 mutation also develop meningiomas.73 SMARCB1 mutations are thought to account for 35% of AT/RT, 48% of familial schwannomatosis, and 10% of sporadic schwannomatosis.73,74 It is believed that individuals with non-truncating mutations in SMARCB1 have a low risk for AT/RT, but genotype-phenotype correlations are still under study to help better define disease mechanisms and the extent of phenotypic overlap of the three syndromes associated with SMARCB1 mutations.73
SMARCE1 heterozygous loss of function alterations such as truncations, frameshifts, and gross deletions cause predisposition for multiple spinal and cranial clear cell meningiomas.75,76 The lifetime risk for clear cell meningiomas in SMARCE1 mutation carriers has yet to be defined; however, age of onset and penetrance are extremely variable, with some carriers presenting in early childhood while others remain unaffected through adulthood. 76
SUFU heterozygous germline pathogenic mutations cause nevoid basal cell carcinoma syndrome (NBCCS), also referred to as Gorlin syndrome. NBCCS/Gorlin syndrome is a genetically and clinically heterogeneous condition characterized by multiple basal cell carcinomas, jaw keratocysts, skeletal anomalies, palmar and plantar pits, calicification of the falx cerebri, coarse facial features, macrocephaly, increased risk for childhood-onset desmoplastic medulloblastoma, ovarian and cardiac fibromas, ocular anomalies, and cleft lip and palate.77 Approximately 5% of individuals with NBCCS will develop medulloblastoma, however current data suggests the risk is higher (~30%) in individuals with mutations in SUFU than in other genes that cause NBCCS.78,79 Some individuals with mutations in SUFU develop isolated medulloblastoma without other symptoms of NBCCS.79 SUFU has also been implicated in familial meningioma.80
TP53 is a tumor suppressor gene, and germline mutations within it are associated with Li-Fraumeni syndrome (LFS). An individual carrying a TP53 mutation has a 21-49% lifetime risk of developing cancer by age 30 and a lifetime cancer risk of 68-93%.81 The most common tumor types observed in LFS families include soft tissue and osteosarcomas, breast cancer, brain tumors (including astrocytomas, glioblastomas, medulloblastomas and choroid plexus carcinomas), and adrenocortical carcinoma (ACC); other cancers, including colorectal, gastric, ovarian, pancreatic, and renal, have also been reported.82,83 Studies have shown that a small percentage of women with early onset breast cancer that do not carry BRCA1 and BRCA2 mutations are identified to have mutations in TP53.84-86
TSC1 and TSC2 are genes associated with tuberous sclerosis complex (TSC), a multi-system neurocutaneous disorder characterized by presence of benign hamartomas in multiple tissues and organs, seizures, and intellectual disability.87 Benign hamartomas can be found in the heart (rhabdomyomas), brain (astrocytomas), kidneys (angiomyolipomas), skin, eyes, lungs (pulmonary lymphangioleimyomatosis), skeleton, and endocrine glands.87-91 The lifetime risk of renal cancer development in TSC is 2-5%.88
VHL mutations are associated with von Hippel-Lindau disease (VHL). VHL is an autosomal dominant cancer predisposition syndrome with about a 20% de novo mutation(139) rate and an estimated incidence of 1 in 36,000.92 VHL is characterized by renal tumors, adrenal pheochromocytoma (PCC), retinal angiomas, central nervous system hemangioblastomas, pancreatic cysts, and neuroendocrine tumors. The associated lifetime risk of RCC in those with VHL is estimated at 25-70%, depending on disease subtype. VHL-associated renal tumors tend to be earlier-onset (average age of diagnoses is 39 years) and multifocal.93 Published literature supports that patients carrying a partial germline VHL gene deletion have a higher RCC risk than those carrying full-gene deletions.94,95
Indications for Testing
BrainTumorNext may be appropriate in the following situations, combined with common red flags for hereditary cancer:
* On the same side of the family
Common Red Flags for Hereditary Cancer
The American Society of Clinical Oncology (ASCO) recommends that genetic testing be offered to individuals with suspected inherited (genetic) cancer risk in situations where test results can be interpreted, and when they affect medical management of the patient. It is sufficient for cancer risk assessment to evaluate genes of established clinical utility that are suggested by the patient’s personal and/or family history.96-99
Benefits of Testing
Identifying patients with an inherited susceptibility for certain cancers can help with medical management and risk assessment. For example, this information can:
BrainTumorNext analyzes 27 genes (listed above). All genes are evaluated by next generation sequencing (NGS) or Sanger sequencing of all coding domains, and well into the flanking 5’ and 3’ ends of all the introns and untranslated regions. In addition, sequencing of the promoter region is performed for the following genes: PTEN (c.-1300 to c.-745), MLH1 (c.-337 to c.-194), and MSH2 (c.-318 to c.-65). The inversion of coding exons 1-7 of the MSH2 gene is detected by NGS and confirmed by PCR and agarose gel electrophoresis. For ALK, only variants located within the kinase domain (c.3286-c.4149) are reported. For PHOX2B, the polyalanine repeat region is excluded from analysis. Clinically significant intronic findings beyond 5 base pairs are always reported. Intronic variants of unknown or unlikely clinical significance are not reported beyond 5 base pairs from the splice junction. Additional Sanger sequencing is performed for any regions missing or with insufficient read depth coverage for reliable heterozygous variant detection. Reportable small insertions and deletions, potentially homozygous variants, variants in regions complicated by pseudogene interference, and single nucleotide variant calls not satisfying 100x depth of coverage and 40% het ratio thresholds are verified by Sanger sequencing.101 Gross deletion/duplication analysis is performed for the covered exons and untranslated regions of all 27 genes using read-depth from NGS data with confirmatory multiplex ligation-dependent probe amplification (MLPA) and/or targeted chromosomal microarray. For APC, all promoter 1B gross deletions as well as single nucleotide substitutions within the promoter 1B YY1 binding motif are analyzed and reported. If a deletion is detected in exons 13, 14, or 15 of PMS2, double stranded sequencing of the appropriate exon(s) of the pseudogene, PMS2CL, will be performed to determine if the deletion is located in the PMS2 gene or pseudogene.
BrainTumorNext can detect >99.9% of described mutations in the included genes listed above, when present (analytic sensitivity).
|TEST CODE||TEST NAME||TURNAROUND TIME (days)|
|8847||BrainTumorNext||14 - 21|