Submitting Author: Durham, Benjamin Heath, MD
Institution: University of California, Davis, School of Medicine
Additional authors:Mingyi Chen, MD, PhD, Ralph Green, MD, PhD, Joo Y. Song, M.D., Carol Marshall, MD, Denis M. Dwyre, MD
Session: Erythroleukemia and megakaryoblastic AML and mimics

History of Present Illness:

The patient is a 51-year-old male with a history of hypertension and smoking who presents with increasing shortness of breath that occurred particularly with exertion over the past five-to-six weeks prior to admission. Around three-to-four weeks prior to admission, the patient also experienced weakness and lightheadedness with no associated symptoms of diaphoresis or chest pain. Furthermore, the patient also noted he had been paler than usual. Just one week prior to admission, the patient had epistaxis that lasted about three hours but resolved. Due to this experience, the patient went to his primary care physician and was found to be neutropenic. Therefore, he eventually went to an outside hospital.

Medical History:

Hypertension

Smoking

Medications:

Hydrochlorothiazide = 12.5 mg daily

Family History:

Mother = Peripheral neuropathy

Father = Gout and “low platelets for years that would require monthly CBC, and before he died, he bled out and needed platelet transfusions.” (The patient could not remember the disease name.)

Two sisters = No known medical problems.

Social History:

The patient had a 13 pack year history, and quit smoking recently. The patient drank a six pack of alcohol every other day for 20 years; however, he quit about 10 months prior to his presentation.

Laboratory Values from Outside Hospital (No reference values provided):

CBC:

WBC = 1.4 x 103 cells/mm3

Hemoglobin = 4.8 grams/dL

Hematocrit = 13.8%

Platelets = 22,000 cells/mm3

Manual Differential:

Neutrophils = 32%

Metamyelocytes = 4%

Myelocytes = 4%

Blasts = 8%

Lymphocytes = 52%

Additional Laboratory Values:

AST = 104

ALT = 31

INR = 1.7

Physical Examination:

Vital Signs: Temperature = 98.7ºF (37.1ºC); Blood Pressure: 128/61 mm of Hg; Pulse = 84; SpO2 = 97% on room air

General Appearance: Healthy, alert, no distress, pleasant affect, cooperative, and jaundiced integument.

Eyes: Conjunctivae and corneas clear. PERRL, extraocular muscles intact, and sclerae show a mild yellow discoloration.

HEENT: Moist mucous membranes.

Neck: Supple; no lymphadenopathy; thyroid symmetric and normal in size

Cardiac: Normal rate and regular rhythm, no murmurs, clicks, or gallops.

Pulmonary: Clear to auscultation, except decreased breath sounds in the right lung base with fine crackles in the left lower lung base.

Abdomen: Soft, non-tender, normal bowel sounds, and no hepatomegaly or splenomegaly.

Extremities: No cyanosis, clubbing, or edema.

Skin: Color, texture, and turgor normal. No rashes or lesions.

Neurological: Alert and oriented x 3 with no gross, focal deficits.

Current Laboratory Results of Interest:

CBC:

WBC = 2.2 x 103 cells/mm3 (4.5 - 11.0 K/mm3)

RBC = 2.21 x 106 cells/mm3 (4.5 – 5.9 M/mm3)

Hemoglobin = 7.3 grams/dL (13.5 – 17.5 grams/dL)

Hematocrit = 21.0% (41-53%)

MCV = 95.0 um3 (80 – 100 um3)

MCH = 33.1 pg (27-33 pg)

MCHC = 34.8% (32-36%)

RDW = 19.3 units (0-14.7 units)

MPV = 8.9 um3 (6.8-10.0 um3)

Platelets = 49,000 (130-400 K/mm3)

Manual Differential:

Segs = 7%

Bands = 3%

Lymphs = 80%

Monocytes = 1%

Blasts = 9%

Vitamin B12 and Methylmalonic Acid Level:

Vitamin B12 = 189 pg/mL (211-911 pg/mL)

Methylmalonic acid = 0.18 (0.10-0.40 µmol/L)

Coagulation Studies:

INR = 1.20 (0.87-1.18)

Liver Enzymes and Function Testing:

Aspartate Transaminase (AST) = 99 (15-43 U/L)

Alanine Transferase (ALT) = 28 (6-63 U/L)

Alkaline Phosphatase (ALP) = 87 (35-115 U/L)

Albumin = 3.5 (3.4-4.8 g/dL)

Protein = 6.1 (6.3-8.3 g/dL)

Bilirubin (Total) = 2.0 (0.3-1.3 mg/dL)

Gross Description:

A. Two Wright-Giemsa stained peripheral blood smears received.

B. Two Wright-Giemsa stained intraoperative bone marrow touch preparation smear slides,

three Wright-Giemsa stained bone marrow aspirate smear slides, and one Iron stained

bone marrow aspirate smear slide received.

C. Received in formalin, decalcified, designated "left posterior iliac crest," is a

1.8 x 0.3 cm brown-red cylindrical segment of roughened tissue. The specimen is

submitted intact in cassette C1.

D. No clot section.

Sent 13.0 mLs of bone marrow aspirate to a reference laboratory for CEBPA, KIT, NPM1, FLT3, and AML FISH.

Microscopic Description:

PERIPHERAL BLOOD: The erythrocytes are normocytic-to-near macrocytic in size with anisocytosis and poikilocytosis. There are schistocytes, elliptocytes, and dacrocytes noted. Also, there are numerous nucleated red blood cells, and a portion of the nucleated red blood cells show irregular nuclear contours and nuclear budding. Polychromasia is prominently seen. The leukocytes show a leukopenia with neutropenia and monocytopenia. There are occasional dysplastic myeloid cells noted that have a pelgeroid morphology. Occasional circulating blasts are seen. The platelets are decreased in number with scattered large and giant platelet forms noted.

BONE MARROW ASPIRATE SMEARS/TOUCH IMPRINTS: The bone marrow aspirate shows cellular spicules and is adequate for interpretation. There is a left-shifted myeloid lineage that is markedly decreased with dysmyelopoiesis noted. A population of blasts with a high nuclear-to-cytoplasmic ratio; open chromatin; and scant, granular cytoplasm is seen. The erythroid lineage is markedly increased with a left-shifted maturation where the majority of the erythroid precursors are proerythroblasts. There is megaloblastic maturation with dyserythropoiesis noted. The megakaryocytes are nearly aplastic with a few scattered dysplastic forms seen.

A 500 cell differential count performed on the bone marrow aspirate shows the following:

Bands/Segs 1.0%

Myelocytes/Metamyelocytes 3.0%

Promyelocytes 0.0%

Blasts 10.0%

Erythroid Precursors 82.0%

Lymphocytes 3.0%

Monocytes 0.0%

Plasma Cells 0.0%

Eosinophils 1.0%

Basophils 0.0%

Blasts = 10% of all nucleated cells

Blasts = 56% of non-erythroid cells

BONE MARROW CORE BIOPSY: The decalcified bone marrow core biopsy is adequate for evaluation. The bone is normal in morphology. The bone marrow is markedly hypercellular (90-95%). The myeloid lineage is nearly aplastic with the absent of mature forms. The erythroid lineage shows near replacement of the marrow by erythroid precursors, of which many appear to be immature. The megakaryocytes are markedly decreased in number, and most of the scattered megakaryocytes show dysmegakaryopoiesis. There are also many blasts/immature cells that are difficult to assess as to whether or not they are of the myeloid or erythroid lineage. Thus, special staining and immunohistochemical staining are applied to the specimen.

Special Stains:

Bone Marrow Aspirate:

Iron: Histiocytic and sideroblastic iron present (3+/6); occasional ringed sideroblast noted.

NSE: Negative in the immature cells.

MPO: Negative in the immature cells, positive in more mature

myeloid cells.

PAS: Positive in some erythroid cells with some of these showing

a globular cytoplasmic staining pattern.

Bone Marrow Core Biopsy:

Reticulin: Focal 1-2+/4+ reticulin fibrosis.

PAS: Positive in some immature cells. Lymphoid aggregate

identified.

Immunohistochemistry:

CD117: Positive in approximately 5-10% of cells.

CD138: Clusters of plasma cells positive.

E-cadherin: Focally positive.

CD34: Negative.

MPO: Negative with approximately 5% cells positive.

CD3: Scattered lymphocytes positive with identification of lymphoid

aggregate.

Ki-67: Positive in approximately 30-40% of cells in marrow.

CD20: Positive in scattered lymphocytes and some lymphocytes in aggregate.

Flow Immunophenotypical Analysis:

MORPHOLOGY: One cytospin preparation and one direct aspirate smear from bone marrow are examined. The slides show a cellular marrow with a predominance of erythroid precursors. The erythroid cells shows megaloblastic maturation, and dysplasia is noted. A population of blasts is seen. Myeloid precursors and megakaryocytes are decreased in number, and dysplasia is noted in these populations.

FLOW DATA: Area A, the lymphocyte gate, comprises 2% of the cells analyzed. The cells in

this area are a mix of T-cells and B-cells without immunophenotypic abnormality.

Area D, an immature cell / blast gate, comprises 25% of the cells analyzed. The cells in

this area express dim CD33 and CD7. No significant expression of CD34, HLA-DR, T-cell

antigens, B-cell antigens, or TDT is seen.

Area F, the CD45 negative gate, comprises 67% of the cells analyzed. The cells in this

area are comprised of erythroid precursors with CD235a and CD71 expression. No CD34

expression is seen in these cells.

IMPRESSION: Abnormal study showing erythroid hyperplasia and increased blasts with

CD33+/CD7+ expression. Study suspicious for AML/erythroid leukemia vs. a high grade MDS with increased blasts.

COMMENTS: The patient is a 51 year old male presenting with pancytopenia with circulating blasts. The bone marrow flow shows erythroid hyperplasia with increased blasts. This is suggestive of AML subtype erythroid leukemia (erythroid cells >50% of marrow cells) vs. high grade MDS with increased blasts. Lab testing on the patient also has shown B12 deficiency. This finding may also contribute to some of the findings on the flow.

Correlation with the bone marrow study is required.

STAINING TECHNIQUES: GTG/FISH

BANDING LEVEL: 425

# CELLS COUNTED: 20

# CELLS ANALYZED: 20

# CELLS KARYOTYPED: 2

RESULTS:

46,XY[5]

nuc ish (p53,ATM,D13S319,Cep 12,13q34)x2[200]

Insufficient metaphases were available to evaluate inv(16)

FISH.

MALE KARYOTYPE

INTERPRETATION

Using an oligonucleotide culture method, there were no clonal abnormalities. Only 5 metaphases were available for analysis, which does not constitute a complete study. Using fluorescence in situ hybridization for chromosome regions 11q, 12, 13q, 17p; all interphase nuclei showed a normal result. No deletions or duplications were detected. These results are consistent with a favorable clinical outcome.

AML Molecular FISH studies:

NPM1+

This bone marrow case demonstrates several interesting features and leads to a challenging differential diagnosis. The marrow shows a prominent erythroid hyperplasia with a hypoplasia of the other two cell lineages. Trilineage dysplasia is prominent. The blast population is aberrantly expressing dim CD33 and CD7. Also, the cytogenetic results show a normal male karyotype; however, this case demonstrates NPM1 positivity. In addition, B12 deficiency was also noted on lab testing. It is unclear how much the B12 deficiency is contributing to the marrow findings because flow immunophenotypic, clonal cytogenetic, and morphological abnormalities have been described in B12 deficiency, and these features can cause B12 deficiency to mimic myelodysplastic syndromes. [1]

Thus, the challenging differential diagnosis includes erythroleukemia (erythroid/myeloid), pure erythroid leukemia, megaloblastic anemia secondary to vitamin B12 deficiency, refractory anemia with excess blasts, acute myeloid leukemia with myelodysplasia-related changes, or acute myeloid leukemia with maturation and an increase in erythroid precursors due to erythropoietin therapy. [2]

Thus, the presence of normal cytogenetics and NPM1 positivity; the effect of B12 deficiency on the marrow; and whether or not the patient has erythroleukemia, pure erythroid leukemia, acute myeloid leukemia with myelodysplasia-related changes, or a high-grade myelodysplastic syndrome are interesting points that make this case challenging.

Therefore, clinical follow up was conducted with this case and is reported. Forty-eight hours following the initiation of vitamin B12 replacement therapy, a follow up bone marrow was performed. It showed similar immunophenotypical and morphological features to the diagnostic marrow. Furthermore, despite the presence of a low serum level of vitamin B12, the methylmalonic acid level was normal. Therefore, there is an absence of a functional B12 deficiency, which was not known at the time of the presented case.

Reference:

1. Parmentier S, Meinel J, Oelschlaegel U, et al. Severe pernicious anemia with distinct cytogenetic and flow cytometric aberrations mimicking myelodysplastic syndrome. Ann Hematol 2012; 91: 1979-1981.

2. Arber DA, Brunning RD, Orazi A, Porwit A, et al. Acute myeloid leukemia, not otherwise specified. WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues, 4th Edition. Eds. Swerdlow SH, Campo E, Harris NL, et al. Lyon: IARC, 2008. 134-136.

Acute erythroid leukemia, erythroid/myeloid

Flow cytometry showed that the blasts were also CD71 and CD235a (heterogeneously) positive. The blasts did not express CD56, CD13, CD117, or megakaryocytic markers. The patient was treated with vitamin B12 for 48 hours and then was re-marrowed. The subsequent bone marrow showed the same features. Next, the patient was started on Induction chemotherapy with the 7 + 3 (cytarabine and idarubicin) regimen. After induction chemothearpy, the patient underwent 2 cycles of consolidation chemotherapy with high dose Cytarabine. The patient is in complete remission at present and waiting for allogeneic bone marrow.

Acute myeloid leukemia with NPM1 mutation and prominent erythroid component

Case 311