Berg, Holly

Abstract The diagnosis of acute promyelocytic leukemia (APL) relies on the identification of PML::RARA fusion. While the majority of APL cases harbor a typical t(15;17)(q24;q21), atypical genetic mechanisms leading to the oncogenic PML::RARA fusion have been reported yet their frequency and scope remain poorly characterized. We assessed the genetic findings of 831 cases with APL investigated with concurrent chromosome banding analysis and dual‐color dual‐fusion fluorescence in situ hybridization (D‐FISH) analysis at our institution over an 18.5‐year timeframe. Seven hundred twenty‐three (87%) cases had a typical balanced t(15;17) with both testing modalities. Atypical karyotypic results including complex translocations, unbalanced rearrangements and insertional events occurred in 50 (6%) cases, while 6 (0.7%) cases were cryptic by conventional chromosome studies despite PML::RARA fusion by D‐FISH evaluation. Atypical FISH patterns were observed in 48 (6%) cases despite apparently balanced t(15;17) on chromosome banding analysis. Two hundred fifty (30%) cases displayed additional chromosome abnormalities of which trisomy/tetrasomy 8 (37%), del(7q)/add(7q) (12%), and del(9q) (7%) were most frequent. Complex and very complex karyotypes were observed in 81 (10%) and 34 (4%) cases, respectively. In addition, 4 (0.5%) cases presented as an apparently doubled, near‐tetraploid stemline clone. This report provides the largest appraisal of cytogenetic findings in APL with conventional chromosome and PML::RARA D‐FISH analysis. By characterizing the frequency and breadth of typical and atypical results through the lens of these cytogenetic testing modalities, this study serves as a pragmatic source of information for those involved in the investigation of APL in both the clinical and research laboratory settings.

Abstract KMT2A gene rearrangements are a major oncogenic driver in multiple hematologic neoplasms. Apart from t(9;11)(p21;q23) (KMT2A/MLLT3) in acute myeloid leukemia (AML), KMT2A gene rearrangements are considered to convey high risk and poor overall survival. Herein, we report a case of a 7 year old boy with newly diagnosed AML and a cryptic KMT2A/AFDN gene fusion resulting from a 5′KMT2A insertional event. The results of conventional chromosome studies revealed trisomy 8 in all 20 metaphases, with normal-appearing chromosomes 6 and 11. A KMT2A break-apart FISH probe identified 2 intact copies of the KMT2A gene region and an extra 5′KMT2A signal in 85% of interphase nuclei. Subsequent FISH studies using a KMT2A/AFDN dual-color dual-fusion FISH probe revealed positive results for a single fusion in 82% of interphase nuclei, indicating a KMT2A/AFDN gene fusion. Subsequently, metaphase FISH confirmed the location of the KMT2A/AFDN fusion at 6q27. To our knowledge, this represents only the second time in the literature that a cryptic KMT2A/AFDN gene fusion resulting from a 5′KMT2A insertional event was reported.

Abstract Introduction/Objective Matrix-assisted laser desorption ionization/ time-of-flight mass spectrometry (MALDI-TOF MS) direct from positive blood culture bottles has facilitated drastic drops in turn-around times for microorganism identification but has been poorly studied for anaerobes. We investigated the ability of MALDI-TOF to provide early anaerobe identification at 4 hours and 18-24 hours of growth on agar from anaerobic blood culture bottles. Additionally, we reviewed medical records of such patients to ascertain impact of early identification on antimicrobial treatment. Methods Over 9 months, we ran MALDI-TOF on early growth from blood cultures positive for growth in BACTEC™ Lytic/ 10 Anaerobic/F bottles. Broth from each bottle was subbed to sheep blood agar (4 hours, 5% CO2) and 2 CDC (BD-BBL™) anaerobic blood agar plates (examined 18-24 hours and 48 hours). Bruker Biotyper® RUO v7854 and Mayo Clinic Custom MALDI-TOF MS libraries were used for identification. Results 144/184 (78%) bottles resulted in growth of aerobic bacteria. Of the remaining bottles with growth of anaerobes, 38 were assessed by early MALDI-TOF. Early MALDI-TOF at 4 hours identified 3 Clostridium perfringens (8%) and an additional 26/38 (68%) isolates at 18-24 hours (both Gram-positive and –negative). Routine 48 hour identification was required for 9 (24%) isolates. In 7 cases, early MALDI-TOF resulted in a change to more appropriate antimicrobial therapy, most often for Bacteroides. Conclusion 29/38 (76%) of anaerobes from blood culture bottles were identified by early MALDI-TOF and reported to the clinician at least 24 hours before routine review of anaerobic sub plates for growth. All C. perfringens and Bacteroides were identified by 4 and 24 hours, respectively. Although early MALDI-TOF resulted in antimicrobial therapy adjustments in a minority of cases, it may allow for more targeted and earlier antimicrobial therapy. Early MALDI-TOF from anaerobic blood culture bottles should be considered for improved patient care and antimicrobial stewardship.

AbstractAcute undifferentiated leukemia (AUL) is a very rare hematologic neoplasm that expresses no markers specific for either myeloid or lymphoid lineages. While commonly observed in several acute leukemias, KMT2A rearrangements in AUL have been rarely reported in the literature. We report the third case to our knowledge of AUL harboring a KMT2A rearrangement. Furthermore, the KMT2A/GIMAP8 gene fusion identified in this case represents a novel KMT2A rearrangement.

Abstract Zinc-finger protein 384 (ZNF384) gene fusions with EP300 have recently been described as a recurrent fusion in B-cell acute lymphoblastic leukemia (B-ALL) with a good response to conventional chemotherapy, suggesting a favorable prognosis. Herein, we report on a female patient aged 12 years with uninformative conventional chromosome and B-ALL panel fluorescence in situ hybridization studies with chromosomal microarray showing multiple copy number gains, including relative gains in the ZNF384 (12p13.31) and EP300 (22q13.2) gene regions, suggesting a cryptic EP300/ZNF384 fusion. Ultimately, a next-generation sequencing assay, mate pair sequencing, was utilized to confirm EP300/ZNF384 fusion in this B-ALL clone, which may confer a favorable overall prognosis and potential targeted therapy.

Abstract Background Flow cytometric detection of T‐cell clonality is challenging. The current available methodology for T‐cell receptor (TCR) Vβ repertoire evaluation is a complex assay and has limited sensitivity especially for detecting low levels of disease. Therefore, there is an unmet need for a reliable, simple, and rapid assay to identify T‐cell clonality. The rearrangement of the TCRB gene involves the random and mutually exclusive expression of one of two constant β chain genes (TRBC1 and TRBC2), analogous to the kappa and lambda gene utilization by B cells. Methods Here, we used a single TRBC1 antibody, in conjunction with other T‐cell associated markers, to detect T‐cell clonality in tissue biopsies and body fluids. A total of 143 tissue/body fluid specimens from 46 patients with a definitive diagnosis of a T‐cell neoplasm and 97 patients with no T‐cell malignancy were analyzed with a cocktail of monoclonal antibodies including CD2/CD3/CD4/CD5/CD7/CD8/CD45/TCRγδ/TRBC1. Results We examined TRBC1 expression on neoplastic T‐cell populations identified based on their immunophenotypic aberrancies, and monotypic TRBC1 expression was identified in all 46 known T‐cell lymphoma cases. We applied a similar gating strategy to the 97 cases without T‐cell neoplasms, and arbitrarily dissected T‐cell populations into immunophenotypically distinct subsets; in this group, we found that all cases revealed an expected polytypic TRBC1 expression in all subsets. Conclusions Single TRBC1 antibody detection of T‐cell clonality by flow cytometry is a simple, rapid, and robust assay that could be routinely utilized in flow cytometric laboratories.