Chronic Myelomonocytic Leukemia Diagnosis By Flow Cytometry: An Italian Center Experience

Abstract

Chronic myelomonocytic leukemia (CMML) is a myeloid neoplasm characterized by dysplasia, abnormal production of monocytes, and an increased risk of leukemic evolution. According to WHO, CMML patients show a persistent (≥3 months) absolute PB monocytosis (≥1×109/L) and relative monocytosis (≥10% of PB leukocytes), with the exclusion of Chronic Myeloid Leukemia (CML), Myeloproliferative Neoplasms, and a blast cell count <20% in PB and/or BM. In addition, morphological and/or histopathological evidence for diagnostic dysplasia ≥1 of the three major BM cell lineages must be present. If dysplasia is absent or not diagnostic (<10%), the presence of cytogenetic or molecular alterations or flow cytometry (FC) abnormalities may be utilized as co-criteria for the diagnosis of CMML. FC is helpful in confirming the monocyte and blast cell counts. In addition, FC is useful for confirming the presence of distinct monocyte populations. Based on the expression of CD14 and CD16, monocytes are classified into classical (MO1) (CD14+/CD16−), intermediate (MO2) (CD14+/CD16+) and non-classical (MO3) (CD14-/CD16+). FC analysis of PB monocytes was proposed as a quick and efficient tool for distinguishing CMML from reactive monocytosis by highlighting an increase in the MO1 fraction above 94% and a decrease in the percentage of MO3. Using FC, we evaluated 25 patients with a confirmed diagnosis of CMML in BM and PB samples, as well as 11 patients with other malignancies (MPN-monocytosis), and 28 patients with reactive monocytosis only in PB samples. Among these patients, 21/25 (84%) of CMML patients displayed MO1 ≥94%, while in patients with reactive monocytosis, MO1≥94% was observed in 11/28 (39%) and 2/11 (18%) in patients with MPN-monocytosis. Then we analyzed the MO3 fraction, which was <1.13% in 18/25 (72%) CMML patients, only one patient with MO1 lower than 94% resulted in an M3 fraction >1.13%. The evaluation of SLAN positivity in this fraction MO3 was always lower than 1.7%. The patient with MO1<94% and MO3 > 1.13% had an active ureteral carcinoma and, at NGS evaluation, had TET2, JAK2, and ZRSR2 mutations. In BM and PB samples from MPN patients, the MO3 fraction was in 2/11 <1.13%, the same patients with MO1> 94%, the diagnosis was a CML in one patient and a PMF triple negative in the other patient. PB samples from reactive monocytosis MO3 fraction were in 11/28 (39%) <1.13%, the same patients with MO1> 94%. The evaluation of SLAN positivity in this fraction MO3 in MPN and reactive monocytosis was always superior at 1.7% (range 4.9-60%). BM samples from CMML and MPN patients reflected the percentage of MO1 and MO3 of PB. Commonly PB and BM monocytes in CMML display aberrant expression of CD56. Furthermore, it was detectable a fraction of non-maturing neutrophils (CD66b+/CD11b-) in nearly half of CMML patients. In conclusion, our real-life experience of routinely FC study on PB and BM CMML samples is similar to the studies described in the literature. It confirms the comparable percentages of monocyte subpopulations in both samples, suggesting that PB samples can be used for diagnostic FC in these patients.