Phospho-IkappaB Is Abnormally Expressed in Bone Marrow of CMML Patients.

Introduction: Chronic myelomonocytic leukemia (CMML) is a heterogeneous group of bone marrow disorders currently grouped into the MDS/MPD overlap by WHO classification. Its clinical and laboratory features includes the presence of up to 20% blast in the bone marrow and the peripheral blood, a PB monocyte count >1000/mL, splenomegaly, variable reticulin fibrosis in the bone marrow and cytopenias. The exact pathogenesis remains in question and there are no effective therapies. Recent studies in certain myeloid disorders suggest that the nuclear transcription factor NFkB regulates cell survival, proliferation, and differentiation. It has been found to be highly expressed in AML and may serve as an important therapeutic target. Little is known about NFkB in other hematologic disorders, including CMML. Examination of NFkB activation in situ has been technically difficult due to lack of quality antibody reagents suitable for fixed tissues. Recently, phosphospecific antibodies have become available, which reflect the functional status of proteins. IkB regulates NFkB subunits by sequestering them in the cytoplasm. Phosphorylation of IkB by IKK results in release of NFkB and translocation to the nucleus. Thus, phospho-IkB (pIkB) is an indicator of NFkB activation. We studied the pattern of pIkB expression in CMML as a surrogate of NFkB activation.

Methods: We identified a cohort of 24 CMML (CMML1=17; CMML2=7) patients and 9 healthy controls. Cases were characterized clinically and pathologically. Immunohistochemistry (IHC) for pIkB was performed in trephine biopsies using a phosphor-specific antibody (Cell Signaling). The staining pattern was compared to normal bone marrow. We utilized JMP 5.1.2 statistical software to compare a variety of clinical and laboratory parameters.

Results: The mean age of patients at diagnosis was 61 (range:38–72). Median WBC, Hgb, PLT, absolute monocytes were 18.6K/ul, 10.2g/dL, 93K/ul, 4K/ul respectively. As expected, the overall survival (OS) was short (mean OS = 11.1 months). Compared to normal bone marrow, pIkB was found to be abnormally activated in maturing granulocytic cells and was found to be present in cytoplasmic as well as nuclear locations. The mean % neutrophils that expressed pIkB was 36.6 compared to 14.9 for normal bone marrow (P<0.00001). No clear association between OS or PFS was detected in this relatively small series based on neutrophil pIkB expression. However, neutrophil pIkB expression was correlated with higher WBC (R = 0.9, P=0.03) and absolute monocyte count (R =0.19, P=.04).

Conclusions: An abnormal in situ pIkB expression pattern is present in CMML compared to normal bone marrow, suggesting abnormal activation of NFkB. Interestingly, maturing myeloid cell expression of pIKB was associated with higher WBC and absolute monocyte count. Further studies are warranted in examining the role of NFkB activation in CMML and potential therapeutic intervention in this pathway, such as with proteasome inhibitors.