PMC8116882_01
Characteristics of IMT. (A) Fascicular proliferation of monotonous spindle cells within myxoid stroma (left), and immunohistochemical staining with anti-ALK antibody (right). (B) Computed tomography (CT) scan before initiating entrectinib (top), and after 9 months of entrectinib (bottom). Arrows indicate lung micronodules. (C) Workflow for CT scan and longitudinal CTCs collection during treatment.
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The tumor cells were interspersed in a myxoid stroma with a heterogeneous inflammatory infiltrate (Figure 1A, left). Positivity for ALK varied, with cells showing more or less diffuse cytoplasm staining (Figure 1A, right). On staging, CT scanning revealed multiple small nodules in both lungs, mainly in the upper left lobe (Figure 1B, top), and 18 fluorodeoxyglucose-PET showed a 29 x 18 x 29 mm osteolytic lesion in the left iliac bone, subsequently confirmed by bone CT scan. A diagnosis of metastatic ALK-positive IMT was made and an anti-ALK therapy (entrectinib 600 mg/day) was adopted (Figure 1C). One year after starting the therapy all lung metastases disappeared (Figure 1B, bottom) and the bone lesion was much reduced in size (23 vs. 29 mm), showing sclerotic margins around the central lytic area.
CTC enrichment and detection. (A) The CTC enrichment approach used to detect and isolate EpCAM-positive (EpCAMhigh/+) and negative (EpCAMlow/-) CTCs from cancer patients' blood samples by the CellSearch system and the ASCD device, respectively. APC-tagged anti-CD45 antibody is used to detect and distinguish peripheral blood leukocytes from CTCs, whereas ferrofluid-tagged and PE-tagged anti-EpCAM and -CK antibodies to isolate EpCAM+/CD45- and/or CK+/CD45- cells, respectively. (B) Representative images of EpCAM+ and EpCAM- CTCs captured by CellSearch and ASCD, and stained for cytokeratins (CK), CD45, ALK or with nuclear dye DAPI.
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Administration) for the In vitro Diagnostic (IVD) detection and collection of circulating tumor cells in cancer patients. According to the system, an event is classified as CTC when its morphological features are consistent with that of a cell expressing the epithelial cell adhesion molecule (EpCAMhigh/+) and exhibiting a CK+, DAPI+ and CD45- phenotype (Figure 2A). However, since EpCAM expression may change during tumor evolution and CTC expressing low levels of EpCAM are discarded by the CellSearch system, cells expressing no or low EpCAM (EpCAMlow/-) were also collected through an antibody-independent Automatic Sample Collection Device (ASCD) coupled to our CellSearch system and subsequently stained for CK, CD45, and DAPI (Figure 2A). ALK-positive IMT cells, a customized assay with a FITC-conjugated anti-ALK antibody was designed and results were expressed as the total number of ALK+CTCs per 7.5 ml of peripheral blood (Figure 2B). As summarized in