Prognostic significance of the IGVH mutation status and immunohistochemical analysis of ZAP70 and CD38 in bone marrow biopsies in chronic lymphocytic leukemia

Chronic lymhocytic leukemia (CLL) is a common malignancy among adults in Western countries. It may have a various processes in a range of between subtle and aggressive nature. It is important to predict whether the patients would have the agressive disease. Rai and Binnet Classification systems are not enough to make this prediction. Thus, a number of complementary factors have been introduced to predict the prognosis of CLL. Recently, efforts focus on the immunoglobulin heavy chain variable domain (IgVH); it is thought that the abcence of mutation is associated with aggressive disease and poor survival outcomes. Identification of IgVH mutations are expensive and difficult; moreover, there is no consensus on any standarts about how to perform or evaluate, so indirect methods indicating the mutation status become more important. It is reported that Zeta chain-associated protein 70 and CD38 are expressed in the absence of IgVH mutations in CLL. It is said these indicators may be used instead of IgVH mutation identification.This study aims to evaluate the relationships between IgVH, ZAP70 and CD38 proteins, which has been shown to be associated with poor prognosis, and to investigate whether their expression is related with the bone marrow infiltration patterns.MATERİAL&METHOD: Paraffin blocks of bone marrow biopsy specimen of 84 patients with CLL was collected within 5 years, then ZAP70 and CD38 antibodies were performed to the slices from the parafin blocks via avidin-biotin method. Staining of tumor cells with these two antibodies were compared, each antibody was assessed with respect to marrow infiltration patterns. On the other hand, DNA samples of 20 patients were isolated from the paraffin blocks and PCR was performed using Biomed-2 primers and IgVH mutation status with sequence analysis was evaluated; then results were compared to immunohistochemistry.RESULTS: Well known poor prognostic factors, ZAP70 and CD38 had positive correlation between their staining (p=0,000); but that correlation was not found between this two indicators and IgVH (p=1.000 and p=0.931, respectively). It is also observed that ZAP70 staining cases required earlier treatment following diagnosis (p=0,046).Bone marrow infiltration pattern and ZAP70 and CD38 expression were analyzed and with immunohistochemistry, both markers were found to have increased expression in diffuse infiltration (for ZAP70; p=0,000, for CD38; p=0,001). The patients with diffuse infiltration needed earlier treatment (p=0,019) and they had advanced disease according to Binnet classification (Stage B,C) (p=0,000).CONCLUSION: we conclude that ZAP70 and CD38 expressions may be used in routine practice as a negatif prognostic factor in CLL. Larger study groups are required to demonstrate the risk of IgVH mutation status on the nature and the progression of the disease. It is also revealed that we should propose the diffuse infiltration as a poor prognostic factor. Of note, patients with positive ZAP70 staining may have more agressive disease and earlier treatment should be considered and so they should have closer follow up.Keywords: CLL, bone marrow biopsy, IgVH mutation, ZAP70, CD38, immunohistochemistry, PCR