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ASCO 2009 POSTER: Circulating Tumor Cells Monitored
Over Time in Lung Cancer Patients
Madelyn S. Luttgen1, Dena Marrinucci1, Daniel Lazar1,
Michael Malchiodi1, Peter Clark1, Edward Huynh2, Kelly Bethel2, Lyudmila
Bazhenova3, Jorge Nieva4, Peter Kuhn11The Scripps Research Institute, 10550
N. Torrey Pines Rd, 92037. 2Scripps Clinic, 10666 N. Torrey Pines Rd, La Jolla ,
CA 92037. 3Moores UCSD Cancer Center, 3855 Health Sciences Dr, La Jolla, CA
92093. 4Billings Clinic, 2825 Eighth Ave North, Billings, MT 59107.
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BACKGROUND & METHODS
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Fig1: Example CTCs from a single NSCLC patient. Red: cytokeratin,
green: CD45, blue: DAPI.
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Circulating tumor cell (CTC) detection and enumeration is
a valuable tool for monitoring cancer patient status and outcome. While
many current techniques employ immunomagnetic-enrichment based protocols focused
on the importance of a particular CTC number as the indicator of patient status
or outcome, we employ a cytometric, enrichment free approach using an
immunofluorescent protocol to monitor CTC counts in patients with non-small cell
lung cancer (NSCLC) over the course of treatment.
METHODS
8cc of anti-coagulated blood is collected from NSCLC patients at the time of
progression, 3 weeks, 3 months, 6 months, 9 months, and one year after the
initial draw. Red blood cells are lysed and CTCs are identified via an
immunofluorescent staining protocol that uses a pan anti-cytokeratin antibody
cocktail directed against 9 cytokeratins, a DAPI nuclear counterstain, and a
white blood cell stain, CD45. FAST detects the positions of fluorescing cells so
relocation and retrospective CTC analysis is possible. CTCs are morphologically
reviewed and identified as cytokeratin and DAPI positive, and CD45 negative.
(Figure 1A-1D) To date, we have assayed 44 blood samples obtained from 22 NSCLC
patients. A schematic of the blood processing protocol is displayed in Figure 2.
Patient response to therapy was determined by RECIST1.1 criteria every 3 months
from the time of enrollment.
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Figure 2. Schematic of blood processing protocol.
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Figure 3.
Marrinucci, D, Bethel K, Nieva, J, et al: Circulating tumor cells from
well-differentiated lung adenocarcinoma retain cytomorphologic features of
primary tumor type: A case study. The Archives of Pathology, in press.
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RESULTS
PATIENT CHARACTERISTICS
There are 65 patients currently enrolled in the study. Forty-six patients have
follow-up data which was included in the analysis. Twenty-five of those 46
patients are deceased. The histological subtypes in the 46 cases for which the
data was available included adenocarcinoma (29/46), squamous cell carcinoma
(6/46), large cell carcinoma (2/46), mixed adeno-squamos carcinoma (1/46), and
non-small cell lung carcinoma not further described, poorly differentiated, or
with a mixed pattern (7/46).
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Figure 6. Case Study 1: Correlation between Change in CTC Count and
Clinical Response Based on RECIST1.1 Criteria
Figure 7. Case Study 2: Correlation between Change in CTC Count and
Clinical Response Based on RECIST1.1 Criteria
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RESULTS 1 of 2
126 of 136 (93%) blood samples from the 46 NSCLC patient blood samples have
CTCs. Seventeen of 46 patients have CTC data from time 0 and 3 weeks. However,
only 4 of 17 (24%) show a correlation between CTC count change at 3 weeks and
clinical response at 3months. Twenty of 46 patients have CTC data for time 0 and
time 3 months. Eight of 13 (62%) patients with stable disease or partial
response at 3 months show a correlation with a decrease or stable CTC count at 3
months. Four of the 5 (80%) patients that do not correlate at 3 months show a
correlation between change in CTC count and response at subsequent time points
(Figure 5). Six of 7 (86%) patients with progressive disease at the 3 month time
point show a correlation with an increase in CTC count at 3 months. The one
patient with progressive disease but a low and stable CTC count at 3 months
subsequently had a partial response at the next time point (Figure 4).
RESULTS 2 of 2
Patients that do not show a correlation between change in CTC count and clinical
response at 3 months show a correlation with subsequent time points. Six of 6
(100%) patients with stable disease at 3 months and progressive disease at 6
months have an increase in CTC counts at either 3 months or 6 months. Six of 7
(86%) patients with stable disease or partial response at both 3 months and 6
months have a decreased or stable CTC count over this time period. The one
patients with stable disease or partial response but an increase in CTC count at
6 months had progressive disease at 9 months.
Nineteen of 20 (95%) patients with blood samples at time 0, 3
months, 6 months, and 9 months either show a direct correlation between change
in CTC count and clinical response at that time point or a predictive response
to therapy at a later time point.
CONCLUSIONS
CTCs can be effectively enumerated in metastatic NSCLC patients, with the
majority demonstrating CTCs in the setting of progressive disease. The
change in CTC count at 3 mos, but not at 3 wks, correlates with response to
chemotherapy. Further follow-up will determine the predictive value of CTC
enumeration on survival.
FUTURE DIRECTIONS
The Kuhn Laboratory is committed to learning more about the physical properties
of cancer. Please refer to
www.physicsoncology.org for more information.
Special thanks to Scripps Clinic, Billings Clinic, and the Palo Alto Research
Center for their help and support. Also, thank you to NIH for support in funding
this research through grant NIH NCI RO1CA125653.
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