Submitting Author: Jamali, Mina, MD MSc FRCPC
Institution: Sunnybrook Health Sciences Centre and the University of Toronto
Additional authors:Hubert Tsui Marciano D. Reis
Session: B Lymphoblastic Leukemia/Lymphoma

The patient is a 36-year-old, previously healthy man presenting with a month long history of shortness of breath, night sweats and fatigue. Routine bloodwork revealed severe anemia and neutropenia. As no precipitating causes were found on preliminary investigation, a bone marrow study (BMS) was considered.

Laboratory investigations reveal:

WBC 1.8k/uL, Hb 59g/L, Platelets 191k/uL

A 200 cell count differential reveals the following absolute counts: neutrophils 0.1k/uL, lymphocytes 0.3k/uL, myelocytes 0.1k/uL, metamyelocytes, blasts 1.2k/uL

HCT 0.160L/L, RBC 1.66x1012/L, MCV 95.9fL, RDW 14.5%, MCH 35.2pg, MCHC 367g/L, Retic 3x109/L

Slightly elevated liver function tests and normal creatinine

Peripheral blood smear:

The peripheral blood smear shows leukopenia due to severe absolute neutropenia. Granulopoiesis is left shifted with occasional myelocyte and metamyelocyte showing toxic granulation. Blasts account for the majority of leukocytes accounting for ~69% of total WBCs. They show a spectrum of size and morphology from small to large and round to irregular nuclear contours. They all show fine nuclear chromatin with occasional prominent nucleolus and scant to modest deep blue cytoplasm. Cytoplasmic granulation is rare and Auer rods are not seen. Eosinophils, basophils and monocytes are not apparent. There is marked anemia with poikilocytosis. Platelets show significantly poor granulation with numerous large and giant forms.

Bone marrow biopsy:

Process: Left posterior iliac crest, B.Plus fixation, RDO Rapid decalcifier (EDTA-HCl based) decalcification

The bone marrow core subcortical biopsy consists of a single core measuring 1.1cm which is excellent for evaluation. It is close to 100% cellular. The marrow is replaced by sheets of blasts showing a spectrum of size and morphology as was seen in the blood. Granulopoiesis and erythropoiesis are markedly reduced and non-progressive. Megakaryocytes are slightly increased in number. There is focal loose clustering and occasional hyper and hypolobated forms.

Bone marrow aspirate:

The bone marrow aspirate smears show no spicules for evaluation and are hemodilute. Blasts comprise the majority of the cells present, again showing variation in size and morphology. Rare granulocyte, erythroid precursor and small lymphocyte is noted. Eosinophils and basophils are rare. Megakaryocytes are not seen on these preparations.

Immunohistochemical stains:

CD34 shows positive mononuclear cell expression in ~10% of total cells. CD117 is negative. Almost half the blasts are positive for TdT. CD20 is strongly positive in the blasts. IgM is negative. MPO reveals reduced but residual granulopoiesis with negative expression in the blasts.

Flow cytometric analysis:

Flow cytometry was performed on a portion of the bone marrow aspirate and shows a tight clustered population spanning the negative-dim CD45 vs low side scatter region accounting for 53% of total events. These cells express partial/dim CD45, partial CD34, HLA-DR, dim CD33, CD19, dim CD20, cytoplasmic CD79a, bright CD10 and show dim surface kappa immunoglobulin light chain restriction. They are negative for cytoplasmic MPO and all other B/T/NK/myelomonocytic antigens.

Cytogenetic analysis was performed on a portion of the aspirate.

46,XY,t(9;22)(q34;q11.2)[2]

/45,idem,-3,-7, der(8)t(3;8)(q12;p11.2~21),add(9)(p11),+mar[5]

/46,XY[13]

Results by G-banded karyotype: Abnormal clonal proliferation with a complex karyotype including t(9;22).

Seven out of twenty metaphases analyzed showed the classic t(9;22)(q34;q11.2). Five of these cells in addition showed a complex karyotype including an unbalanced 9p rearrangement resulting in deletion of the short arm of chromosome 9, monosomy 7, an unbalanced rearrangement between chromosomes 3 and 8 with breakpoints estimated at 3q12 and 8p11.2~21, resulting in deletion of the majority of the short arm of chromosome 3, and an unidentified small marker chromosome. The remaining thirteen metaphases showed a normal male karyotype.

BCR-ABL1 fusion transcripts (p210 breakpoint) were detected by real-time quantitative PCR. This is consistent with the presence of Philadelphia chromosome positive cells in this specimen.

Course:

CSF was negative. The patient was admitted to hospital and initiated on the Dana-Farber chemotherapy regimen as well as tyrosine kinase inhibitor (TKI), imatinib mesylate. He is also being considered for allogeneic stem cell transplantation.

The application of cytogenetics to the clinical study of acute lymphoblastic leukemia (ALL) has lagged behind its use in acute myeloid leukemia (AML), largely because of the rarity of ALL and each specific cytogenetic subgroup. We describe here a case of Ph-positive B-ALL also associated with a complex karyotype, CD20 positivity and surface light chain restriction. The increased risk associated with t(9;22) may change with the use of newer TKIs; however, the multiple karyotypic aberrations seen in this case portend very poor prognosis, especially given what appears to be a ‘monosomal karyotype’ equivalent in ALL. There is insufficient evidence to support chronic myelogenous leukemia (CML) in blast phase or mixed phenotypic acute leukemia (MPAL).

Our case illustrates the rare occurrence of surface light chain immunoglobulin restriction (sIg+) in ALL. The B-lymphoblasts express nuclear TdT and HLA-DR as well as CD10 which are seen in at least the intermediate stage. IgM negativity would suggest the blasts are not late stage. There is contention as to whether sIg+ represents a late precursor stage of B-cell differentiation. It may be that sIg+ ALL arises when a subset of leukemic cells progress towards greater maturational status ultimately becoming the predominant clone. Along with CD20 expression, this may provide an immunophenotypic signature for post-therapy assessment of minimal residual disease (MRD) or use of lymphoma-like approaches including rituximab. It remains to be seen whether these findings represent biologically distinct entities or have important prognostic implications.

Precursor B-cell lymphoblastic leukemia with surface light chain immunoglobulin restriction and complex karyotype including t(9;22)

B-acute lymphoblastic leukemia with t(9;22)(q34;q11.2); BCR-ABL1 and complex karyotype (surface light chain immunoglobulin restriction)

Case 335