Abstracts of the Scientific Posters, 2013 AACC Annual Meeting ...

Cancer/Tumor Markers
Tuesday, July 30, 9:30 am – 5:00 pm
determined using a Student’s t test and analysis was determined using GraphPad prism
software. In conclusion, this is the first study to show the elevation of serum BCMA in
patients with MM and levels correlated with the change in tumor volume in response
to treatment with cyclophosphamide and bortezomib. We propose that BCMA may be
a new serum biomarker for patients with MM, and is useful to determine prognosis
and monitor the course of their disease.
A-05
PSA Enzymatic Activity: A New Biomarker for Assessing Prostate
Cancer Aggressiveness
D. Georganopoulou 1 , M. Ahrens 1 , P. Bertin 1 , E. Vonesh 2 , T. J. Meade 3 ,
W. J. Catalona 3 . 1 Ohmx Corporation, Evanston, IL, 2 Vonesh Statistical
Consulting, Libertyville, IL, 3 Northwestern University, Chicago, IL
Background and objectives: The recent increase in prostate-specific antigen (psa)
screening rates coupled with improved detection methods have caused a controversial
upsurge in the number of men undergoing prostate biopsy and subsequent treatment.
However, current diagnostic techniques generally suffer from limited ability to
identify which seemingly indolent prostate cancers (pca) are biologically aggressive.
We set out to determine if pca aggressiveness is associated with psa enzymatic activity
in ex vivo prostatic fluid.
Methods: We collected prostatic fluid from 778 post-radical prostatectomy specimens
and randomly selected samples from both the clinically confirmed aggressive (n = 50)
and non-aggressive (n =50) prostate cancer populations for our initial pilot study. In a
blind study, we measured the level of proteolytic enzyme activity of psa (apsa) in each
sample using a fluorogenic peptide probe and used receiver operating characteristic
(roc) analysis to correlate apsa levels with prostate cancer aggressiveness.
Results: We observed that the clinically non-aggressive population had a significantly
higher apsa value (mean = 865μg/ml; median = 654μg/ml) than the clinically
aggressive population (mean = 518μg/ml; median = 449μg/ml), meaning there is a
negative association of apsa with cancer aggressiveness. We performed a roc analysis
appropriate for an unmatched case control study to assess the highest diagnostic effect
for predicting aggressive pca. Among factors considered, apsa and the normalized
ratio of apsa/serum tpsa (rpsa) had the highest discriminatory power for predicting
the presence of aggressive pca. We calculated an area under the curve (auc) of 0.7008
[95% ci: (0.5986, 0.8030)] for apsa and 0.7784 [95% ci: (0.6880, 0.8688)] for rpsa
with the latter being significantly higher (p-value = 0.0300 based on a chi-square test).
Conclusions: Our results show a significant correlation between pca progression and
apsa in prostatic fluid. We found the range of measured apsa for aggressive cases was
94 - 1220 μg/ml while non-aggressive cases ranged from 207 - 2626 μg/ml within
our pilot study. Within the non-aggressive group, there were 11 samples whose apsa
values (1238 - 2626 μg/ml) were greater than the highest apsa value measured within
the aggressive cohort (1220 μg/ml). Using apsa as an aggressiveness biomarker could
result in many (22% in our study population) of the patients diagnosed with nonaggressive
pca being able to avoid or delay radical prostatectomy.
Source of funding: national institutes of health (grant #1r43ca156786-01).
A-06
A Highly Sensitive and Specific Method for Characterization of
Circulating Tumor Cell Subtypes in Breast Cancer Patients
L. Millner, K. Goudy, T. Kampfrath, M. Linder, R. Valdes. University of
Louisville, Louisville, KY,
Introduction: Circulating tumor cells (CTCs) are cells that detach from the primary
tumor, intravasate into the bloodstream, invade distant tissues and produce metastatic
lesions. In breast cancer patients, enumeration of CTCs in blood is used as an adjunct
to assist in predicting overall survival and in clinical management. However, CTCs are
phenotypically heterogeneous and the methods now available for counting these cells
are based on detection of the epithelial marker, Epithelial Cell Adhesion Molecule
(EpCAM). Present methods do not distinguish subtypes and only detect epithelialtype
CTCs. This is significant because CTCs are known to experience epithelial to
mesenchymal transition (EMT), a process that results in increased motility and is
associated with diease progression. Following EMT, a CTC may no longer express
epithelial markers such as EpCAM and evade detection by current methods.
Objective: To establish a model using heterogeneous breast cancer cell lines and a
method for capturing and characterizing distinct CTC subsets. This method should
have high separation efficiency of subtypes, high recovery in spiked blood samples,
and be capable of distinguishing heterogeneous subtypes independent of EMT status.
Materials: We have established a breast cancer cell line panel including all 4 of the
breast cancer molecular subtypes including luminal, HER2, basal-like, and claudinlow.
This model of 4 breast cancer cell lines was used to represent the heterogeneity
in CTCs from a breast cancer patient and the plasticity in the EMT process. We have
chosen to include 1 breast cancer cell line that does not express EpCAM (MDA-
MB-231). The cell lines and their molecular subtypes include MCF-7 (luminal A),
SK-Br-3 (HER2), MDA-MB-231 (claudin-low) and HCC1954 (basal-like) and
each represents an identifiable subtype. 25,000 or 2,500 cells of each cell line were
combined and then identified using a combination of antibodies including HER2,
EpCAM, and CD44. Experiments to determine separation efficiency and percent
recovery were performed on the BD Accuri C6 flow cytometer. Results: The
specificity (separation efficiency) for each subtype was determined to be 67.3% ± 7.1
(±SE) (HCC1954), 91.7% ± 9.7 (MCF-7), 57.3% ± 8.7 (MDA-MB-231), and 100%
± 19.0 (MCF-7). The overall separation efficiency was determined to be 79.4 ± 5.9%
(± SE). Spiking experiments of a single mesenchymal cell line that does not express
EpCAM were conducted in whole human blood, and a sensitivity (percent recovery)
of 84.9 ± 14.6% (±SE) was achieved.
Conclusion: High percent recovery of a spiked mesenchymal breast cancer cell
line into whole blood was achieved. This method isn’t limited to cells that express
EpCAM so CTCs that have undergone EMT are able to be detected. The combination
of antibodies has high separation efficiency with 2 of the 4 cell lines. Enrichment
processes and antibody selection are being optimized to improve specificity of all 4
subtypes. This data indicates that phenotypically diverse CTCs are capable of being
subtyped and characterized. Subtype characterization will allow therapies to be
individually tailored to address each patient’s own CTCs.
Support: P30ES014443 and T32ES011564
A-09
SAP155-mediated c-myc suppressor FBP-interacting repressor splicing
variants as colon cancer screening biomarkers
K. Matsushita, S. Itoga, F. Nomura. Chiba University Graduate School of
Medicine, Chiba, Japan
Background: The c-myc transcriptional suppressor, FUSE-binding protein (FBP)-
interacting repressor (FIR), is alternatively spliced in colorectal cancer tissue.
Recently, the knockdown of SAP155 pre-mRNA-splicing factor, a subunit of SF3b,
was reported to disturb FIR pre-mRNA splicing and yielded FIRΔexon2, an exon2-
spliced variant of FIR, which lacks c-myc repression activity.
Methods: The expression levels of FIR variant mRNAs were examined in the
peripheral blood of colorectal cancer patients and healthy volunteers to assess its
potency for tumor detection. As expected, circulating FIR variant mRNAs in the
PB of cancer patients were significantly overexpressed compared to that in healthy
volunteers.
Results: In this study, novel splicing variants of FIRs, Δ3 and Δ4, were also
generated by SAP155 siRNA and those variants were also found to be activated in
human colorectal cancer tissue. In particular, the area under the receiving operating
characteristic curve of FIRs FIRΔexon2 or FIRΔexon2/FIR was greater than those of
conventional carcinoembryonic antigen (CEA) or carbohydrate antigen 19-9 (CA19-
9). In addition, FIRΔexon2 or FIR mRNA expression in the peripheral blood was
significantly reduced after operative removal of colorectal tumors.
Conclusion: Circulating FIR and FIRΔexon2 mRNAs are potential novel screening
markers for colorectal cancer testing with conventional CEA and CA19-9. Our results
indicate that overexpression of FIR and its splicing variants in colorectal cancer
directs feed-forward or addicted circuit c-myc transcriptional activation. Clinical
implications for colorectal cancers of novel FIR splicing variants are also discussed.
CLINICAL CHEMISTRY, Vol. 59, No. 10, Supplement, 2013
A3