Cancer Cytogenetics Laboratory

Common pediatric malignancies

B-Cell Acute Lymphoblastic Leukemia (ALL)

Precursor B-cell acute lymphoblastic leukemia (ALL) accounts for the majority (85%) of leukemia diagnoses in children. Most patients with ALL have an abnormal clone by conventional cytogenetic studies. The most common karyotype among children with ALL is hyperdiploidy (50-55 chromosomes per cell), which is associated with a good prognosis in the absence of any structural anomalies, and when associated with trisomies of chromosomes 4, 10, and 17. The most common chromosome translocations in pediatric ALL include t(12;21)(p13;q22), t(1;19)(q23;p13), t(8;14)(q24;q32), t(11;var)(q23;var) and t(9;22)(q34;q11.2). The t(12;21)(p13;q22) is associated with a TEL/AML1 fusion and is not possible to detect by conventional cytogenetic studies. This translocation, along with other common translocations, can be successfully detected by FISH. Deletions within the short arm of chromosome 9 (9p21) are also common in ALL, although the clinical significance of this finding is still uncertain.

A representative profile of a B cell leukemia is shown below. (Karyotype, FISH and array results)

The karyotypes demonstrates a dicentric 9;20 and loss of one normal copy of chromosome 20; a deletion of the long arm of a chromosome 16; trisomy 21; and an isochromosome of the short arm of an X.

Metaphase FISH demonstrating two normal copies of the X, and an isochromosome Xp.

Array profile for chromosome 9, demonstrating loss of the short arm of chromosome 9, as well as a homozygous deletion in 9p21.3.

Array profile for chromosome 20, showing the deletion of most of the long arm. The results are consistent with an unbalanced t(9;20).

T-Cell Leukemia/Lymphoma

T-ALL is a neoplastic disorder of lymphoblasts committed to the T-cell lineage. These malignancies comprise 15% to 20% of acute leukemias. While one half of T-ALL patients have normal chromosome studies, molecular cytogenetic or SNP array analysis can identify abnormalities in almost all cases. These include:

• Cryptic deletions of p16/CDKN2A in 9p21.3
• Rearrangements involving 11q23 (MLL)

Myelodysplastic syndrome (MDS)

Myelodysplastic syndromes (MDS) may present as a primary finding or after treatment with chemotheraputic agents. Clonal cytogenetic abnormalities are more frequently observed in cases of secondary MDS (80% of patients) than in primary MDS (40%-60% of patients). The common chromosomal abnormalities associated with MDS include: inv(3), -5/deletion 5q, -7/deletion 7q, trisomy 8, deletion 13, and deletion 20q. MLL rearrangements have also been observed.

Lymphoma

Lymphomas fall into two major categories, Hodgkin lymphoma and all other lymphomas (non-Hodgkin lymphomas or NHLs). The various types and sub-types of non-Hodgkin lymphoma are classified by both microscopic appearance of the tumor cells and well-defined genetic rearrangements. FISH with probes to the IGH locus in 14q32 are typically employed to rule out a B cell lymphoma. Large cell anaplastic lymphoma often results in a translocation of the ALK locus in 2p23. SNP array analysis may also be performed to rule out a variety of unbalanced rearrangements.

Rhabdomyosarcoma

This group of sarcomas includes the embryonal, alveolar and undifferentiated subtypes of rhabdomyosarcoma. The alveolar subtype (ARMS) is clinically much more aggressive, and accounts for 20-30% of rhabdomyosarcoma. Most ARMS cases contain one of two recurrent chromosomal translocations: t(2;13)(q35;q14) or t(1;13)(p36;q14). DNA probes for the PAX7 gene on chromosome 1, for the PAX3 gene on chromosome 2 and for the FKHR gene on chromosome 13 are used in combination to detect these rearrangements and any subsequent amplifications. Embryonal rhabdomyosarcomas are often characterized by copy number changes of chromosomes 2, 8, 9, 11, and 13, or loss of heterozygosity for chromosome 11p15, which are best detected by SNP array analysis.

Neuroblastoma

Neuroblastoma is the most common extracranial solid tumor in childhood. The tumors are derived from developing neuronal cells of the sympathetic nervous system and are found mostly (but not exclusively) in infants and young children. Neuroblastomas have distinct genetic profiles that predict the clinical phenotype. MYCN amplification and/or deletions involving 1p36 are frequently associated with poor outcome. Interphase FISH to rule out a 1p36 deletion or MYCN amplification, in combination with a high resolution SNP array are the most appropriate tests to perform for characterization of these malignancies.

Synovial Cell Sarcoma

Synovial cell sarcoma is a rare soft tissue tumor that accounts for 5-8% of soft tissue sarcomas. A t(X;18)(q11.2;p11.2) is found in 80% of synovial sarcomas in successful standard cytogenetic studies. This translocation results in two gene fusions in which the SYT gene at 18q11.2 is joined with one of two closely related genes at Xp11.2, designated SSX1or SSX2. FISH may be used to detect the disruption of the SYT gene on chromosome 18q11.2 by employing a dual color break apart (BAP) DNA probe set.

Ewings Sarcoma

Ewing tumors form a histologically heterogeneous family belonging to the group of small round-cell tumors derived from the neural crest. Almost 90% of Ewing’s tumors show a t(11;22)(q24;q12), which fuses the EWS gene on chromosome 22 with the transcription factor gene FLI1 on chromosome 11, leading to a hybrid transcript and an oncogenic chimeric protein. In about 5% of cases, the EWS gene is involved in variant translocations, such as a t(21;22)(q22;q12) or t(7;22)(p22;q12). The disruption of the EWS gene on chromosome 22q12 can be visualized using a dual color break apart (BAP) DNA probe set. Additional abnormalities include trisomy 8 and an unbalanced t(1;16)(q12;q11.2).

Atypical Teratoid Rhabdoid Tumor/Renal /Extra-renal Rhabdoid Tumor

Rhabdoid tumor is a highly aggressive tumor that may present in the brain (AT/RT), kidney or other soft tissue site. Mutations, deletions and loss of heterozygosity of the INI1/SMARCB1 gene in chromosome band 22q11.2 characterize the majority of rhabdoid tumors in all anatomic locations. Please see below for a further description of the mutation and deletion assays of INI1/SMARCB1 employed for patients with rhabdoid tumors.

Medulloblastoma and PNET

Medulloblastoma is the most malignant brain tumor in the pediatric population. Subgroups of medulloblastoma have deletions of chromosome 6, alterations of the PTCH pathway, or contain a deletion of 17p or isochromosome 17q. MYC amplification is characteristic of the anaplastic subtype, which is associated with the worst prognosis. A SNP array and specific FISH analyses are the most commonly utilized tests for patients with malignant brain tumors.

Gliomas

Gliomas are tumors of glial cell origin and account for >70% of all primary brain tumors. They are classified according to the World Health Organization (WHO) criteria, which provides grading from I – IV.

Juvenile Pilocytic Astrocytoma (JPA)

JPA, WHO grade I, is the most common glioma in the pediatric population. Fusions of the 5’ region of KIAA1549 with the distal region of BRAF have been identified in approximately 80% of pilocytic astrocytomas. Reverse-transcription polymerase chain reaction (RT-PCR) is performed to detect these fusions.

Other Low Grade (Grades I and II) Gliomas

Ganglioglioma (GG) is a mixed glial and neuronal tumor which represents about 4% of all pediatric brain tumors. A specific mutation in exon 15 of BRAF has been detected in 50% of gangliogliomas, as well as a variety of other grade II gliomas. PCR followed by DNA sequencing of exons 11 and 15 of BRAF is performed to detected this mutation.

High Grade Gliomas

Anaplastic astrocytoma (AA), WHO grade III, and glioblastoma multiforme (GBM), WHO grade IV, account for approximately 10% of pediatric CNS tumors. These tumors are characterized by a variety of chromosomal gains ad loss best detected by a whole genome approach. DNA isolated from primary or recurrent tumors is analyzed using a high resolution SNP array. Constitutional or germline alterations in tumor predisposition genes are evaluated using DNA isolated from a peripheral blood sample.