Hematological Malignancies MYD88 and CXCR4 Mutation Panel

MYD88 L265P mutations are found in >90% of lymphoplasmacytic lymphoma (LPL) and approximately 50% if IgM MGUS patients. This mutation is also quite common in diffuse large B-cell lymphoma, especially DLBCL’s at immune privileged sites such as the skin, testis, and CNS (including ocular). It can also be detected in rare cases of low-grade B-cell lymphoma/leukemia such as marginal zone lymphoma (4%), and chronic lymphocytic leukemia/small lymphocytic lymphoma (2%). The MYD88 gene encodes a cytoplasmic adaptor protein that plays a central role in innate and adaptive immunity and functions as a key linker protein in the Toll Like Receptor (TLRs) and IL-1 receptor signaling pathway. The L265P mutation favors cell survival through IRAK1/IRAK4 and NF-kB signaling. 

CXCR4 encodes a G-protein coupled receptor that activates the AKT1 and mitogen-activated protein kinase signaling pathways. CXCR4 mutations occur in about 40% of LPL and are associated with primary ibrutinib resistance. In a phase II study, responses to ibrutinib in Waldenstroms macroglobulinemia varied depending on mutations status of MYD88 and CXCR4;  highest responses were seen in MYD88Mut/CXCR4WT, intermediate responses in MYD88Mut/CXCR4Mut and lowest responses in MYD88WT/CXCR4WT cases. Germline CXCR4 mutations are also described in in warts, hypogammagloblulinemia, immunodeficiency, and myelokathexis (WHIM) syndrome.


Clinical Utility: 
Although not specific, identification of a MYD88 L265P mutation may aid in the diagnosis of LPL in the appropriate clinicopathologic context. CXCR4 mutations are associated with primary resistance to BTK inhibitors. Recent consensus guidelines strongly encourage molecular studies for MYD88 and CXCR4 in the workup for LPL

This test is performed by PCR-based Next Generation Sequencing of DNA extracted from formalin fixed paraffin embedded (FFPE) tissue or fresh tissue including peripheral blood and bone marrow.  The entire coding regions of MYD88 and CXCR4 are sequenced using massively parallel sequencing (next-generation sequencing) with a combination of multiplexed PCR (customized QIAseq Targeted DNA panel) and sequencing on an Illumina platform.  An in-house bioinformatics analysis pipeline has been used that employs multiple established variant calling tools (FreeBayes, MuTect2 and Scalpel) and variant annotation tools.  The genomic variants have been interpreted in accordance with the 2017 guideline recommendations by AMP/ASCO/CAP (PMID: 27993330).  The assay is validated in accordance with the AMP guidelines (PMID: 28341590).

Clinical Sensitivity:
MYD88 L265P mutations are found in >90% of lymphoplasmacytic lymphoma (LPL). CXCR4 mutations occur in about 40% of LPL. 

Clinical Specificity:
MYD88 and CXCR4 mutations may be identified across a spectrum of lymphomas. Interpretation of this test requires clinical, morphologic, and immunophenotypic correlation.

• 5-10 mL of blood or bone marrow — yellow (ACD) or purple (EDTA) tube
• Formalin-fixed paraffin-embedded (FFPE) tissue blocks
• 10 unstained slides (4-5 microns)
• Fresh Tissue:
  • Stabilize in Allprotect Tissue Reagent (Qiagen) and ship at room temperature – OR
    • Suspend in sterile culture media (RPMI or DMEM) or non-bacteriostatic normal saline in a sterile container and shipped at room temperature – OR
    • Snap Frozen and shipped on dry ice
• If sending DNA: please send 200ng at a minimum of 10ng/µL (DNA must be extracted in a CLIA-certified laboratory or a laboratory meeting equivalent requirements as determined by the CAP and/or CMS)
• Pathology report MUST accompany sample for interpretation of results.
  
  A REQUISITION FORM MUST ACCOMPANY ALL SAMPLES.  Please include detailed clinical information.

Once per week

14-18 days

• Package and ship specimen to remain cold, but not frozen. 
• Ship via overnight express, using the FedEx priority overnight label provided. 
• Contact Client Services for shipping kits and instructions at (855) 535-1522.

1. Campo E et al. The International Consensus Classification of Mature Lymphoid Neoplasms: A Report from the Clinical Advisory Committee. Blood. 2022 Jun 2:blood.2022015851. doi: 10.1182/blood.2022015851. Epub ahead of print. PMID: 35653592.
2. Varettoni et al.Prevalence and clinical significance of the MYD88 (L265P) somatic mutation in Waldenstrom's macroglobulinemia and related lymphoid neoplasms.Blood. 2013. 121(13):2522-2528.
3. Landgren et al. MYD88 and beyond: novel opportunities for diagnosis, prognosis and treatment in Waldenström's Macroglobulinemia.Leukemia. 2014. 28(9):1799-803.
4. Kaiser LM et. Al CXCR4 in Waldenström's Macroglobulinema: chances and challenges. Leukemia. 2021 Feb;35(2):333-345. doi: 10.1038/s41375-020-01102-3. Epub 2020 Dec 3. PMID: 33273682; PMCID: PMC7862063.