Background Measurable residual disease (MRD) is a strong independent poor prognostic factor for acute leukemia. Multiparameter flow cytometry (FCM) is a commonly used MRD detection method. However, FCM MRD detection is not well standardized, and the interpretation is subjective. There are normal/reactive minor cell populations in bone marrow (BM) and peripheral blood (PB), which could be confused with MRD. Methods The FCM data of 231 BM and 44 PB pediatric samples performed in a recent 15‐month period were retrospectively reviewed. These samples were from 56 B‐lymphoblastic leukemia (B‐ALL) patients, 11 T‐lymphoblastic leukemia (T‐ALL) patients, 28 acute myeloid leukemia (AML)/myelodysplastic syndrome (MDS) patients, 44 cytopenia/leukocytosis patients, and five patients with mycosis fungoides. Results There were over 10 normal or reactive minor cell populations identified with certain phenotypes mimicking MRD of acute leukemia. These mimickers included CD19+ NK cells, CD22+ basophils, CD22+ dendritic cells (DCs), and plasma cells for B‐ALL MRD; CD4/8 double‐negative T cells, CD4/8 double‐positive T cells, cytoplasmic CD3+ NK cells, CD2− T cells, CD7− T cells, CD5− gamma delta T cells, CD56+ NKT cells for T‐ALL MRD; CD33+ NK cells, CD117+ NK cells, basophils, plasmacytoid DCs, non‐classical monocytes, CD56+ and/or CD61+ monocytes for AML MRD. Conclusions These data confirm the presence of a variety of normal/reactive minor cell populations that could mimic MRD of acute leukemia by FCM. Recognizing these MRD mimickers is important for correct FCM MRD interpretation.

Background/objectives: Rhabdomyosarcoma (RMS) is characterized by the expression of the myogenic regulatory protein MYOD1. Histologic types include alveolar, embryonal (ERMS), and spindle cell sclerosing RMS (SRMS). SRMS harbors MYOD1 mutations in a subset of adult cases in association with poor prognosis.Design/methods: To study the level of MYOD1 protein expression and its clinical significance, we have analyzed variable numbers of pediatric (<18 years of age) and adult (age range ≥18 to 35 years) ERMS and SRMS cases for presence or absence of MYOD1 immunoreactivity in correlation with clinical outcome and MYOD1 L122R mutations.Results: Lack of MYOD1 immunoreactivity, identified in 23.8% of nonalveolar RMS (non-ARMS) cases, was more prevalent in SRMS (44%) than ERMS (17.2%) and was significantly associated with low overall survival and unfavorable tumor sites (p < .05).Lack of MYOD1 immunoreactivity was not associated with MYOD1 L122R mutations, which were identified in 3/37 (8%) cases including only two of 31 (6.5%) pediatric cases, one of 11 or 9% pediatric SRMS, and one case of infant ERMS. Conclusion:These studies highlight the prognostic role of MYOD1 in non-ARMS. Lack of MYOD1 immunoreactivity is associated with poor prognosis in ERMS and SRMS.MYOD1 gene mutations are generally infrequent in pediatric RMS. Although mutations are predominant in SRMS, they may exceptionally occur in infantile ERMS.

The perinucleolar compartment (PNC) is a small nuclear body that plays important role in tumorigenesis. PNC prevalence correlates with poor prognosis and cancer metastasis. Its expression in pediatric Ewing sarcoma (EWS) has not previously been documented. In this study, we analyzed 40 EWS tumor cases from Caucasian and Hispanic patients for PNC prevalence by immunohistochemical detection of polypyrimidine tract binding protein and correlated the prevalence with dysregulated microRNA profiles. EWS cases showed staining ranging from 0 to 100%, which were categorized as diffuse (≥77%, n = 9, high PNC) or not diffuse (<77%, n = 31) for low PNC. High PNC prevalence was significantly higher in Hispanic patients from the US (n = 6, p = 0.017) and in patients who relapsed with metastatic disease (n = 4; p = 0.011). High PNC was associated with significantly shorter disease-free survival and early recurrence compared to those with low PNC. Using NanoString digital profiling, high PNC tumors revealed upregulation of eight and downregulation of 18 microRNAs. Of these, miR-320d and miR-29c-3p had the most significant differential expression in tumors with high PNC. In conclusion, this is the first study that demonstrates the presence of PNC in EWS, reflecting its utility as a predictive biomarker associated with tumor metastasis, specific microRNA profile, Hispanic ethnic origin, and poor prognosis.

Rhabdomyosarcoma (RMS) is a common malignancy of soft tissue, subclassified as alveolar (ARMS), pleomorphic (PRMS), spindle cell/sclerosing (SRMS), and embryonal (ERMS) types. The Yes-associated protein (YAP) is a member of the Hippo pathway and a transcriptional regulator that controls cell proliferation. We have studied the immunohistochemical expression of YAP in different RMSs, arranged in tissue microarray (TMA) and whole slide formats. Pertinent clinical data including patient age, gender, tumor location, and clinical stage were collected. Out of 96 TMA cases, 30 cases (31%) were pleomorphic, 27 (28%) were embryonal, 24 (25%) alveolar, and 15 (16%) spindle cell. Positive nuclear YAP staining was seen in the PRMS (17/30, 56.7%), SRMS (7/15, 46.7%), ERMS (19/27 or 70%), and less in ARMS (37.5%). YAP nuclear staining was significantly more prevalent in ERMS than ARMS ( p=0.02). Of the 41 whole slide cases, nuclear staining was detected in all ARMS but was restricted in distribution to <30% of the cells, in contrast to ERMS and SRMS, which had diffuse or >30% staining. These results highlight the role of YAP in RMS tumorigenesis, a fact that can be useful in engineering targeted therapy. Restricted nuclear YAP staining (<30% of cells) may be of value in the diagnosis of ARMS.