Hematologic Diseases

Detects mutations in key genes recurrently mutated in chronic lymphocytic leukemia (CLL) and related lymphoid neoplasms. DNA sequence of targeted regions of the ASXL1, ATM, BCOR, BIRC3, BRAF, BTK, CCND1, CCND2, CDKN2A, CDKN2B, DDX3X, DNMT3A, FAT1, FBXW7, HIST1H1E, IKZF3, IRAK4, ITPKB, KRAS, MAP2K1, MAP3K14, MAPK1, MED12, MEF2B, MYD88, NFKBIE, NOTCH1, NRAS, PLCG2, PIK3CD, POT1, PTEN, RB1, RIPK1, RPS15, SAMHD1, SETD2, SF3B1, SPEN, SPOP, TET2, TLR2, TP53, TRAF2, TRAF3, UBR5, XPO1 and ZMYM3 genes is determined using next-generation sequencing (NGS) technology.

disease state indication(s)
Chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), Richter's syndrome (RS), mantle cell lymphoma (MCL)

clinical use
The clinical course of chronic lymphocytic leukemia (CLL) is heterogeneous, and ranges from very indolent with a nearly normal life expectancy to rapidly progressive leading to early death. In recent years, unbiased massively parallel sequencing in CLL has yielded fresh insights into the genetic basis of this disease [1,6,10-15].
• The C481S mutation in the BTK gene and the R665W and L845F mutations in the PLCG2 gene have been shown to be associated with acquired resistance to BTK inhibition by ibrutinib [2].
• Genomic alterations in the TP53, BIRC3, NOTCH1, and SF3B1 genes are associated with adverse outcomes, and their presence or absence can improve risk stratification and treatment selection beyond clinical staging and other prognostic biomarkers [3,4,5].
• The National Comprehensive Cancer Network (NCCN) guidelines recommend TP53 sequencing as informative for prognostic and/or therapy determination and other genes such as NOTCH1, SF3B1, and BIRC3 may provide useful prognostic information [4].
• Mutations in NOTCH1, SF3B1, BIRC3, RPS15, NFKBIE, POT1, FAT1, KRAS, and others have also been shown to confer a poorer prognosis and/or resistance to chemo-immunotherapy. Other genes found to be recurrently mutated in CLL, such as XPO1, MAPK1, MAP2K1, BRAF (non-600), KRAS, IKZF3, FBXW7, MYD88, and TLR2, may have therapeutic implications [1,6,10-14].
• An increased number of mutations in untreated CLL patients is associated with a poor overall survival independent of 17p deletion. TP53 mutations occur in approximately 80% of CLL with 17p deletion , resulting in bi-allelic inactivation and worse prognosis compared with 17p deletion alone [7].
• Certain mutations are detected preferentially in a clonal (e.g.: MYD88, HIST1HE) and sub-clonal fashion (e.g. KRAS, NRAS), suggesting an order of alterations corresponding to earlier and later drivers. The presence of a sub-clonal driver mutation by itself was shown to indicate a poor prognosis and more rapid disease progression. Additionally, in a group of patient samples assessed longitudinally, clear patterns of clonal evolution was found to be common after therapy and was associated with worse overall outcomes [1,14,15].
• NOTCH1, TP53, and CDKN2A/B mutations have been correlated to Richter’s transformation and poor survival [8,9].
• Mantle cell lymphoma (MCL), which immunophenotypically can be similar to CLL, are characterized by having mutations affecting many different genes. ATM and CCND1 are among the most frequently mutated genes found in MCL, but other genes including NOTCH1 and NOTCH2 are also mutated, and may be of prognostic and potential therapeutic importance [3].
• Despite chemoimmunotherapy, high-dose cytarabine, and ASCT, younger MCL patients with deletions of CDKN2A and TP53 have shown to have an unfavorable prognosis and may be candidates for alternative therapeutic strategies [16].

1. Gruber M, et al. Evolving understanding of the CLL genome. Semin Hematol 2014;51:177-87.
2. Woyach JA, et al. Resistance Mechanisms for the Bruton's Tyrosine Kinase Inhibitor Ibrutinib. N Engl J Med 2014;37:2286-94.
3. Swerdlow SH, et al. The 2016 revision of the World Health Organization classification of lymphoid neoplasms. Blood 2016;127:2375-90.
4. National Comprehensive Cancer Network (NCCN) Practice Guidelines in Oncology, Non-Hodgkin's Lymphomas.
5. Rossi D, et al. Integrated mutational and cytogenetic analysis identifies new prognostic subgroups in chronic lymphocytic leukemia. Blood 2013;121:1403-1412.
6. Ljungstrom V, et al. Whole-exome sequencing in relapsing chronic lymphocytic leukemia: clinical impact of recurrent RPS15 mutations. Blood 2016;127:1007-16.
7. Yu L, et al. Survival of Del17p CLL Depends on Genomic Complexity and Somatic Mutation. Clin Cancer Res 2017;23:735-745.
8. Villamor N, et al. NOTCH1 mutations identify a genetic subgroup of chronic lymphocytic leukemia patients with high risk of transformation and poor outcome. Leukemia. 2013;27:1100–6.
9. Fabbri G, et al. Genetic lesions associated with chronic lymphocytic leukemia transformation to Richter syndrome. J Exp Med 2013; 210: 2273-2288.
10. Puente XS, et al. Non-coding recurrent mutations in chronic lymphocytic leukemia. Nature 2015;526:519-524.
11. Herling CD, et al. Complex karyotypes and KRAS and POT1 mutations impact outcome in CLL after chlorambucil-based chemotherapy or chemoimmunotherapy. Blood 2016;128;395-404.
12. Mansouri L, et al. Functional loss of NFKBIE leads to NF-kB deregulation in aggressive chronic lymphocytic leukemia. J Exp Med 2015;212;833-843.
13. Messina M, et al. Genetic lesions associated with chronic lymphocytic leukemia chemorefractoriness. Blood 2014;123:2378-88.
14. Landau DA, et al. Mutations driving CLL and their evolution in progression and relapse. Nature 2015;526:525-530.
15. Landau DA, et al. Evolution and impact of subclonal mutations in chronic lymphocytic leukemia. Cell 2013;152:714-26.
16. Delfau-Larue MH, et al. High-dose cytarabine does not overcome the adverse prognostic value of CDKN2A and TP53 deletions in mantle cell lymphoma. Blood 2015;126:604-611.

methodology/product platform
Capture-based, next-generation sequencing (NGS)

specimen type and requirements
Preferred: Peripheral blood: 2-3 mL in EDTA (purple-top) tube.
Bone marrow: 2-3 mL in EDTA (purple-top) tube.
Unacceptable: Specimens received fixed in alternative fixation methods. Decalcified, frozen or fresh tissue.
Note: Use refrigerated cold pack for transport. Make sure cold pack is not in direct contact with specimen.

turnaround time
Global (TC & PC): 10-12 days

cpt code(s)
81450 (x1)

medicare moldx cpt code
Not applicable

regulatory classification
Laboratory developed test (LDT)

ordering option
Global (TC & PC)