The FDA has recently approved a PCA3 test assay which is
owned by a Canadian company, Gen-Probe. This opens up a whole new avenue for
examining PCa amongst men. I examine some of the issue here at a fairly high
level.
There has been a great deal of discussion regarding PSA and
its lack of sufficient specificity and sensitivity to PCa and there is some
evidence that PCA3 will improve the situation. This is yet to be determined in
extensive clinical trials. One of the problems with PSA is that it is
reflective of total prostate volume and it also naturally increases with age.
Thus a male of say 70 years of age and with a 70 cc prostate may easily have a
PSA of 2.5 just based upon the size and age factors.
Likewise if the male were
40 and had a 35 cc prostate then this may be indicative of PCa. In a recent
paper by McGarty, we detailed the issue
of PSA sampling and the percent change, ie velocity, as a means to assess the
nature of the underlying cause. Namely the more prostate basal cells and
luminal cells the higher the PSA. As we shall see there is better correlation
with PCA3 but the underlying molecular and cellular dynamics do not appear as
well defined at this time, namely we have a marker with no clear underlying
genomics cause.
The PCA3 measurement is define as follows:
PCA3 Score = 1000 [mRNA PCA3]/[mRNA PSA]
where [mRNA PCA3] is the concentration of mRNA of PCA3 and
the same for the denominator. The range is such that a PCA3 score of less than
5 gives a very low likelihood of PCa and >35 gives a very high probability.
The issue here often is repeat biopsy. The suggestion then is that one use PCA3
as a test for repeat biopsy indication (see Gen-Probe PCA3 documentation).
Details on ROC for PCA3 are not broadly available and repeatable at this time.
PCA3 was first discussed in 1999 in a paper by Bussemakers
et al, at which time it was called DD3. In their abstract the authors stated at
the time:
The DD3 gene was mapped to chromosome
9q21–22, and no homology of DD3 to any gene present in the computer databases
was found. Our data indicate that DD3 is
one of the most prostate cancer-specific genes yet described, and this makes DD3
a promising marker for the early diagnosis of prostate cancer and provides a
powerful tool for the development of new treatment strategies for prostate
cancer patients.
It further turns out that PCA3 is a noncoding mRNA and thus
there is no protein resultant. This was speculated by Bussemakers et al when
they published their work in 1999. The key question seems to be why does PCA3
increase when there is a PCa and what is the details of the mechanism.
Furthermore where does PCA3 fit within the context of the many pathways we know
exist in PCa development.
As Cao an Yao report:
The DD3PCA3 encoding
gene is located on chromosome 9 (9q2122). The gene includes four exons and
three introns. In PCa, the most frequent mutation is the selective splicing of
the second exon. At present, there is a vast body of ongoing studies on PCA3.
Hopefully they can further confirm the role of PCA3 in the occurrence and the
development of PCa and provide new treatment targets for patients with PCa.
Hessels
suggested that using quantitative reverse transcriptase polymerase chain
reaction (RTPCR) for the detection of urine DD3PCA3 was a valuable molecular
detection method in patients with PCa and could help reduce unnecessary
biopsies.
In a multicenter study designed to examine the diagnostic capacity of
urine PCA3 detection, the AUC of urine PCA3 detection was 0.66, while the AUC
of serum PCA3 detection was merely 0.57. The sensitivity and specificity of
PCA3 detection were 65% and 66% , respectively .
Recently, researchers have suggested that serum PSA level plus PCA3 detection
was the most promising diagnostic method for PCa . All these studies show that
PCA3 is probably an important urine marker for PCa. It also provides a new clue
for developing noninvasive detection methods for PCa. Hence, PCA3 may have
considerable significance in multiple tumor marker screening of patients for
PCa in the future.
Thus one of the questions is what is PCA3 and why does it
reflect PCa presence. We know that we are measuring mRNA concentrations, and we
know that in measuring them we have experimental evidence that PSA reflects
total cell concentration. But what of PCA3, what does that reflect.
In a recent paper by Clarke et al the authors attempt to
clarify what the role of PCA3 is.
In order to understand
further the importance of the PCA3 gene
in PCa we undertook a more detailed investigation of this gene and its
chromosomal locus. This investigation points to a considerably more complex
transcriptional unit for PCA3 than
originally reported including additional novel exons. We describe a number of
novel PCA3 splice variants with more specific expression
in PCa tissues and metastases. We also demonstrate that PCA3 is embedded in the intron of a
second gene, BMCC1, a gene implicated
in controlling oncogenic transformation and that both genes showed increased
expression in PCa and metastases. The absence of a TATA box element within a
human gene promoter has been associated with promiscuous transcriptional initiation.
The PCA3 gene does not contain an upstream
TATA sequence and it was therefore of interest to determine whether any additional
transcription initiation sites existed for PCA3
Perhaps this relationship to BMCC1 may lead to some insight.
They continue:
BMCC1
is upregulated in PCa and androgen inducible Since PCA3 is upregulated in PCa
and since we showed here that this gene is embedded in a second gene BMCC1,
implicated in cellular proliferation, we determined whether BMCC1 was also differentially
regulated in PCa. We used a set of RT-PCR primers that span that region of the BMCC1
gene (exons 6 and 7), specific for the full-length BMCC1-1 transcript.
Expression of BMCC1-1 was evident in normal prostate and BPH specimens and was upregulated
in PCa and metastases. This was confirmed using
primers corresponding to the BCH C-terminal region of BMCC1 and for BMCC1-2.
Indeed amplification of this isoform gave better discrimination between PCa and
BPH. Extending these experiments to PCa and other cell
lines revealed that both genes were highly expressed, specifically in the PCa
cell line LNCaP. In addition BMCC1-1 was detected in a second PCa cell line
DU145 but at lower levels. PCA3 is also expressed in DU145 but required further
rounds of amplification for detection. The shorter BMCC1 isoforms (BMCC1-3 and/or
BMCC1-4) were also detected (using primers specific for the BCH region) in an
EBV-transformed lymphoblastoid cell line (JHP), but the longer BMCC1-1 isoform was
not detected. Previous data have shown that the level of PCA3 can be induced in
LNCaP cells after treatment with dihydrotestosterone, which mimics the effects
of binding of the androgen receptor (DHT). We determined whether BMCC1- 1 was
also responsive to hormonal induction. The results …. demonstrate that both PCA3
and BMCC1 are maximally induced in the LNCaP cell line at a concentration of
0.5 mM DHT.
Thus there seems to be some means of related induction but
again no definitive relationship to well defined pathways.
The following is the PCA3 and PSA ROC for comparison. Note
the following (see de la
Taille):
The area under the
curve of the receiver operating characteristics (AUC ROC) of the PCA3 Score was
compared with that of serum total PSA, PSAD and % free PSA. The diagnostic
accuracy of the PCA3 Score was statistically significantly better than that of
serum total PSA, PSAD and % free PSA. The greatest diagnostic accuracy of the
PCA3 Score was obtained at a cut-off of 35: specificity 76% and sensitivity 64%
. At a sensitivity of 80%, the PCA3 Score specificity of 58% was higher than the
44% for PSAD and 27% for serum total PSA and % free PSA.
The following from de la Taille is the comparative ROC. It appears that from the
limited data available that the ROC curve is substantially better with PCA3
than PSA alone.
The key questions remaining in my mind are:
1. What pathway elements does PCA3 reflect. What genetically
is happening and what is the underlying system model. This is always an issue.
As with primary medicine we have underlying physiology, here we must have some
underlying genomics.
2. What are the cellular mechanism which control PCA3. Again
this is a pathways issue.
3. How sustainable is PCA3 ROC for this assay. Many tests
have been done and FDA approval is merely acceptance of some limited tests.
4. How does one relate PSA and PCA3. Note that the PCA3
measure does reflect PSA concentration, so we have not abandoned PSA.
5. Why do we normalize PCA3 on PSA? If PSA has such a
variability are we normalizing on something which is inherently unpredictable?
References
1.
Bussemakers, M., et al, DD#: A New Prostate specific Gene, Am
Assn Cancer Res, 1999.
2.
Clarke, R.,
New Genomic Structure for Prostate Cancer Specific Gene PCA3 within BMCC1, Plus
One March 2008.
3.
de la Taille, A. et al, The PCA3 Assay Improves the Prediction of Initial Biopsy Outcome, 1
CHU Henri Mondor, Paris, France; 2 CHU.
4.
DeMarzo, A.,
et al, Molecular Alterations in Prostate Cancer as Diagnostic, Prognostic, and
Theraputic Targets, Int Soc Uro Path 2008.
5. McGarty, T., PSA Evaluation Methodologies, MIT/RLE Draft Paper 2010, http://www.telmarc.com/Documents/Papers/2010%20PSA%20Evaluation%20Methodologies%20Short.pdf
6.
Rattue, P.,
Repeat Prostate Biopsies, Medical News Today, February 2012.
7.
Schmidt, U.,
et al, Quantitative Multi Gene Expression Profiling of Primary Prostate Cancer,
The Prostate V 66 pp 1521-1534, 2006.
8.
Wang, R. et al, Rational Approach to
Implementation of Prostate Cancer Antigen 3 into Clinical Care, Cancer, Nov
2009.
9.
Wright, J., P. Lange, Newer Potential Biomarkers for Prostate
Cancer, Rev Uro V 9 2008 pp 207-213.
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