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case_images: Morphology and immunophenotype of neoplastic cells in peripheral blood and bone marrow. (A) Peripheral smears revealed neoplastic lymphocytes of 9-10 microm, with irregular, often angulated nuclei and prominent nucleoli (1000x magnification). (B) Flow cytometry at diagnosis showed 54% lymphocytes, the majority of which demonstrated the following phenotypes: CD2+/CD3+/CD4+/CD5+/CD7+/CD8-/CD52+/CD56-. SSC indicates side scatter. Green gate: lymphocytes; orange gate: monocytes; blue gate: granulocytes. The numbers on the other scatter plots represent percentages of the quadrants. (C) Histology showed normocellular bone marrow with approximately 15% interstitial and vaguely nodular lymphoid infiltrate in the bone marrow (200x magnification, Naphthol AS-D chloracetate esterase staining. (D) Immunohistochemistry using TCL1 antibody (Abcam, Cambridge, UK, 1:100 dilution), 200x magnification.
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case_images: Peripheral blood smears showed 46% of atypical lymphocytes with slightly angulated, 8-10 microm large nuclei and prominent nucleoli (Figure 1A). Peripheral blood and bone marrow showed 44% and 33% of CD2+, CD3+, CD4+, CD5+, CD7+, CD8-, CD52+, CD56- T-cells, respectively (Figure 1B). Bone marrow biopsy showed 10% of focal, interstitial lymphoid infiltrate showing CD3, CD4 and TCL1 (T-cell leukemia/lymphoma protein 1, protein product of TCL1A gene) positivity, as well as TDT (terminal deoxynucleotidyltransferase), CD10, and CD34 negativity with normal hematopoiesis in the background (Figure 1C,D).
case_images: Cytogenetic findings. (A) 47,XX, del(6)(q22q23), i(8)(q10), +i(8)(q10), t(9;22;18)(q34;q11;q21), inv(14)(q11;q32), der(15)t(8;15)(q22;p13), add(16)(p13) karyotype was seen in 5 out of 30 metaphases obtained from peripheral blood (an arrow marks the Philadelphia chromosome). Four metaphases showed the same abnormalities with the addition of add(17)(q25), while one metaphase showed the addition of +der(22). (B) Interphase cells showed one fusion, two red, and two green signals with BCR::ABL1 dual fusion probe (Abbott, Chicago, United States of America). Approximately half of the cells showed an additional fusion signal representing +der(22) (630x).
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case_images: All metaphases with clonal alterations (10/30 in peripheral blood and 11/18 in bone marrow) showed del(6q), i(8q), inv(14), as well as t(9;22;18) and two other structural aberrations (Figure 2A). Fusion of BCR and ABL1 genes (Figure 2B), as well as the disruption of the TCL1A gene, was demonstrated.
case_images: Results of Sanger sequencing. Based on alignment with reference RNA sequences, the transcript was identified as in-frame with the breakpoint showing fusion of BCR exon 6 to ABL1 exon 2.
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case_images: Following reverse-transcriptase polymerase chain reaction (PCR) using primers for BCR exon 1 and ABL1 exon 2, Sanger sequencing of the transcript revealed an e6a2 BCR::ABL1 transcript indicating an atypical BCR breakpoint (Figure 3).
case_images: Summary of therapies and clinical data. ALEM: alemtuzumab; CYCLO: cyclophosphamide; IMA: imatinib; CHOP: cyclophosphamide-, doxorubicin-, vincristine-, and prednisone-containing regimen; NILO: nilotinib.
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case_images: Figure 4 summarizes the sequence of therapies (Figure 4).