autologous blood and marrow transplantation - Blog Science ...

Detection of Cytokeratin +
Cells in Marrow as a
Predictor for Tumor Cell Contamination and
Measurement of Tumor Cell Loss in Apheresis Products
Deborah L Hood, Karel A. Dicke, Peggy J. Donnell, Lorene K.
Fulbright, George R. Blumenschein
Arlington Cancer Center, Arlington, TX
ABSTRACT
Metastasis of solid tumors, such as breast cancer, to bone marrow can be
detected using an immunofluorescent assay for cytokeratin with a sensitivity of 1
tumor cell in 10 6
cells. Initial comparisons to pathology in 70 consecutive aspirates
revealed that our methodology not only confirmed positive pathology but was 3
times more likely to identify micrometastases; 35 of 70 vs. 12 of 70. Between
January 1996 and April 1998, 79 stage III and IV breast cancer patients were
leukapheresed after cytoreductive therapy. Marrow samples from each patient were
assayed for the presence of cytokeratin +
cells before apheresis.
Aspirate results Mean tumor frequency - apheresis + apheresis
Negative 70/79(89) 0 64/70(91%) 6/70(9%)
Positive 9/79(11%) 1/3 X10 5
1/9(11%) 8/9(89%)
Of the patients with a negative aspirate, 91% had negative collections.
Similarly, 89% of the patients with a positive aspirate had at least one positive
apheresis procedure, suggesting that marrow status is a strong predictor of tumor
involvement in apheresis products.
Flow cytometric analysis of 10 6
stained cells identified 12 positive apheresis
from eight patients, containing a me*an of 2.3 X10 5
cytokeratin +
cells. Apheresed
cells were not immediately frozen but were centrifuged, resuspended in an
isotonic electrolyte solution with 10% autologous plasma, and held overnight at
4°C. Repeat staining and analysis of 24-hour-old specimens revealed that tumor
cells had fallen below the detection level of 1/10 6
cells in 10 of 12 apheresis
products. The other two specimens still contained positive cells but showed
decreases in total tumor cell numbers: 1.6X10 5
to 8.8X10 3
and 1.1 X10 5
to
2.1 X10 3
. These overnight changes emphasize the necessity of assaying fresh
apheresis samples.
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