CHROMOPROBE MULTIPROBE ALL , 5 DEVICE

cMYC (MYC) Breakapart

Translocations involving the MYC (cMYC) oncogene (Avian Myelocytomatosis Viral Oncogene Homologue) are a molecular feature of Burkitt lymphoma and occur in almost every case. These rearrangements can also be found in 5 to 10% of diffuse large B-cell lymphoma (DLBCL)¹.

The majority of rearrangements result in the juxtaposition of MYC to IGH, IGL or IGK in the t(8;14), t(8;22) or t(2;8) respectively. In each case, this results in dysregulation of MYC (as a result of being close to the constitutively active immunoglobulin locus), increased transcription and neoplastic growth². The breakpoints involved are widely scattered throughout the gene but the t(8;14) translocation always spares the protein coding exons which become attached to the derived 14. In the t(2;8) and t(8;22), MYC remains on chromosome 8 and the immunoglobulin locus joins it, resulting in close positioning of the MYC and immunoglobulin loci. Rare cases of translocations not involving immunoglobulin partners have also been reported³.

Patients with MYC rearrangements, in isolation of a confirmed diagnosis of Burkitt lymphoma, were originally thought to have poor prognoses but they do respond well to intensive chemotherapy affording them an increased survival rate. This shows that cytogenetic confirmation of the rearrangement is necessary to manage the patient effectively⁴.

References

1. Savage et al., Blood. 2009 Oct 22;114(17):3533-7

2. Robertson. Nature. 1983 Apr 7;302(5908):474-5

3. Seo et al., Ann Lab Med. 2012 Jul;32(4):289-93

4. Hoelzer et al., Blood. 1996 Jan 15;87(2):495-508

P16 (CDKN2A) Deletion

Deletions of chromosome 9p21 are implicated in a wide variety of tumours including approximately 10% of paediatric ALL patients¹, though the incidence is higher in T-ALL².

The region has been the subject of much study but deletion of the potential tumour suppressor gene P16 (Cyclin-Dependent Kinase Inhibitor 2A (CDKN2A)) was found to take place in 90% of newly diagnosed cases of paediatric ALL showing cytogenetic deletions of 9p21 by FISH³. This study showed that deletion of P16 only (rather than both P16 (CDKN2A) and P15 (CDKN2B)) was the critical step as one case was found to be deleted for P16 but P15 was present. The deletion is usually homozygous (81% compared to 9% hemizygous) in cases of T-ALL whilst homozygous and hemizygous deletions are roughly equal in B-ALL (23% vs. 20% for homozygous and hemizygous respectively).

The gene product inhibits the Cyclin Dependent Kinases CDK4 and CDK6, which are important in controlling cell cycle progression from G1 to S phase⁴. Disruptions of this process are therefore likely to result in the proliferation of mutated cells.

References

1. Heerema et al., Blood 1999; 94:1537-1544

2. Secker-Walker et al., Br J Haematol 1997;96(3):601-10

3. Okuda et al., Blood 1995;85(9):2321-30

4. Fry et al., Mol Cancer Ther. 2004 Nov;3(11):1427-38

E2A (TCF3) Breakapart

Translocations involving the E2A (TCF3: Transcription Factor 3) gene are some of the most common rearrangements in childhood B-ALL, specifically B-cell precursor ALL¹.

The two main partner genes are PBX1 and HLF (on chromosomes 1 and 17 respectively) which become fused to E2A as a result of the t(1;19) and t(17;19) translocations, forming the E2A/PBX1 and E2A/HLF fusion proteins. Other translocation partners have also been detected.

The t(1;19) is the most common E2A rearrangement, being present in around 5% of B-ALL and 20% of pre B-ALL², whilst the t(17;19) is present in around 1% of all ALL cases³. Both are historically associated with a poor outcome, though treatment therapies have overcome this in the case of the t(1;19)⁴. Detection of the t(1;19) is best carried out using molecular methods, such as FISH, as the fusion has been shown to be missed in 20 to 25% of patients by standard cytogenetics⁵. Similarly, other E2A translocations have been shown to be cytogenetically cryptic⁶.

References

1. Barber et al., Genes Chromosomes Cancer. 2007 May;46(5):478-86

2. Alonso CN. t(1;19)(q23;p13) TCF3/PBX1. Atlas Genet Cytogenet Oncol Haematol. July 2012

3. Viguié F . t(17;19)(q22;p13). Atlas Genet Cytogenet Oncol Haematol. May 1999

4. Felice et al., Leuk Lymphoma. 2011 Jul;52(7):1215-21

5. Izraeli et al., Leukemia 1993;7(5):671-8 6. Huret JL. TCF3 (transcription factor 3 (E2A immunoglobulin enhancer binding factors E12/E47)). Atlas GenetCytogenet Oncol Haematol. January 2012

Hyperdiploidy

Approximately 30% of childhood ALL cases have cells exhibiting a chromosome number between 51 and 67. Within this group are prognostically significant subsets, with individuals possessing 51 to 55 chromosomes having a poorer prognosis than those with 56 to 67. The chromosomes that are more readily hyperdiploid are chromosomes 4, 6, 10, 14, 17, 18, 21 and X, with gains of chromosomes 4, 10 and 17 leading to a relatively better prognosis.

TEL/AML1 (ETV6/RUNX1) Translocation, Dual Fusion

The TEL (or ETV6 – Erythroblastosis Variant Gene 6 translocation, ETS) / AML1 (or RUNX1 – Runt-Related Transcription Factor 1) fusion is brought about by the cytogenetically invisible t(12;21) translocation.

This is the most common rearrangement in childhood B-ALL and has been detected using FISH in around 17% of cases¹, compared to a pick-up rate of 0.05% by conventional cytogenetics². The translocation is associated with a favourable outcome and has been associated with late relapse³. TEL has also been shown to be deleted in some children with ALL with loss of heterozygosity (LOH) of chromosome 12p12-13 and this deletion is often associated with a TEL/AML1 translocation⁴. Both the TEL and AML1 genes encode transcription factors and TEL has been shown to be required for proper transcription during haematopoiesis within the bone marrow⁵.

References

1. Jamil et al., Cancer Genet Cytogenet 2000;122(2):73-8

2. Borkhardt et al., Blood. 1997 Jul 15;90(2):571-7

3. Mosad et al., Journal of Hematology & Oncology 2008;1:17

4. Raynaud et al., Blood 1996;87(7):2891-9

5. Wang et al., Genes Dev. 1998 Aug 1;12(15):2392-402

MLL (KMT2A) Breakapart

Rearrangement of the MLL (KMT2A: lysine (K)-specific methyltransferase 2A) gene on chromosome 11q23.3 can be detected in the leukaemic cells of approximately 85% of infants with B-ALL^1,2,3. They can also be found in 3% of de novo and 10% of therapy related AML cases⁴. Translocations involving the MLL gene are generally associated with increased risk of treatment failure⁵.

In infant ALL, the most frequent of these translocations is the t(4;11)(q21;q23.3) translocation involving MLL and the AFF1 (AF4) gene on chromosome 4^6,7. A poor outcome for infants with ALL is strongly associated with the presence of this rearrangement in particular⁶. The discovery that a single YAC spanned breakpoints in four of the more common translocations led to the naming of the candidate gene MLL (Myeloid/Lymphoid or Mixed Lineage Leukaemia). The gene has homology with a drosophila gene (‘trithorax’), which is highly conserved in humans and gives rise to a protein that can be folded to give six zinc finger domains and is a developmental regulator. The zinc finger domains are translocated to the AFF1, MLLT3 and MLLT1 genes on the partner chromosomes in the aforementioned t(4;11), t(9;11)(p22;q23.3) and t(11;19)(q23.3;p13.3) translocations, respectively. Each of the genes involved in these translocations have been shown to have high sequence homology. There have been over 85 recurrent translocations involving the MLL gene reported, with 66 partner genes so far identified⁸.

References

1. Rubnitz et al., Blood 1994;84(2):570-3

2. Secker-Walker et al., Leukaemia 1998;12(5):840-4

3. Rowley, Annu Rev Genet 1998;32:495-519

4. Grossman et al., Leukemia 28 March 2013; doi10.1038/leu.2013.90

5. Pui and Evans, New Engl J Med 1998;339(9):605-15

6. Felix and Lange, Oncologist 1999;4(3):225-40

7. Heerema et al., Leukemia 1999;13(5):679-86

8. Huret JL. KMT2A (myeloid/lymphoid or mixed lineage leukemia). Atlas Genet Cytogenet Oncol Haematol. October 2005

BCR/ABL(ABL1) Translocation, Dual Fusion

The stereotypical Philadelphia chromosome forming translocation, t(9;22), represents a significant abnormality in ALL and is associated with an extremely poor outcome. In a small number of cases, the translocation does not result in a cytogenetically visible Philadelphia chromosome. In these cases, FISH is essential for highlighting the fusion gene.

IGH Breakpart

In Burkitt’s Lymphoma, IGH is most notably involved in rearrangements involving the MYC oncogene as a result of the t(8;14)(q24.21;q32.33) translocation¹. However, other rearrangements of the IGH gene are also seen in a number of different malignancies, including T-ALL, Chronic Lymphocytic Leukaemia (CLL) and Acute Lymphpblastic Leukaemia (ALL).There are a number of stereotypical translocations involved in each of the diseases and more are being described regularly.

In T-ALL for example, IGH is observed in the t(14;14)(q11;q32) translocation (or inv(14)(q11q32) rearrangement)² that is found in T-cell leukaemia associated with ataxia-telangiectasia (AT). However, rare reports have indicated that this abnormality also occurs in B-ALL. The recurrent t(14;19)(q32;q13) translocation associated with chronic B-cell lymphoproliferative disorders, such as atypical CLL, has also been shown to occur in B-ALL and results in the juxtaposition of the IGH and BCL3 genes and subsequent over expression of BCL3³. More recently, a report suggested the involvement of IGH in a novel cryptic translocation in paediatric T-ALL, which also involved TLX3 (HOX11L2) or NKX2-5 (CSX) on 5q35 brought about by a t(5;14)(q35;q32) translocation⁴. IGH is involved in a large number of different rearrangements with fusion partners on almost every other chromosome. Many of these rearrangements have been reported in only one or a few cases but some are more common, such as IGH/BCL2, caused by the t(14;18) translocation⁵, and IGH/CCND1, a result of the t(11;14) translocation⁶.All these rearrangements do, however, have breakpoints within the IGH gene. We have designed a split probe set for IGH, which allows the detection of rearrangements, regardless of the partner gene involved.

References

1. Hoffman, Ronald (2009). Hematology : basic principles and practice (5th ed. ed.). Philadelphia, PA: Churchill Livingstone/Elsevier. pp. 1304-1305

2. Liu et al., Cancer Genet Cytogenet 2004;152:141-5

3. Robinson et al., Genes Chromosomes Cancer 2004;39(1):88-92

4. van Zutven et al., Haematologica 2004;89(6):671-8

5. Huret JL . t(14;18)(q32;q21) (IgH/BCL2); t(2;18)(p11;q21); t(18;22)(q21;q11). Atlas Genet Cytogenet Oncol Haematol. 
May 1998

6. Huret JL . t(11;14)(q13;q32). Atlas Genet Cytogenet Oncol Haematol. May 1998