CD147 is a multifunctional glycoprotein that acts to induce the expression of MMPs,
among other roles 4]–6], 25]. CD147 also forms heterodimers with proton-coupled monocarboxylate transporters (MCT)
MCT1, MCT3, and MCT4 13], 25], helping to maintain lactate and pH homeostasis in epithelial cells, while N-glycosylation
of CD147 leads to self-aggregation and MMP induction 26]. Overexpression of CD147 of MMPs and CD147 has been noted in many malignancies, including
breast and colorectal cancer 7], 8], as the mechanistic role of highly glycosylated CD147 is suspected to assist in tumor
invasion and metastatic spread 26], 27]. However, the prognostic role of CD147 is still controversial, particularly in PCa.
Our results show that CD147 is decreased in malignant prostate samples compared to
tumor-adjacent normal tissue, and further decreases in expression are associated with
advanced pathologic stage and Gleason score, indicating that CD147 may be important
in the progression to advanced stages of PCa. Using a xenograft-derived cell line
model of prostate cancer progression, we confirmed that CD147 expression was decreased
in aggressive PCa. Investigation of CD147-encoding BSG mRNA expression in publicly available microarray data shows that CD147 protein changes
may be regulated transcriptionally. In this study, we also found that CD147 predicts
biochemical recurrence after prostatectomy.
Our results are in disagreement with earlier studies on the expression and prognostic
role of CD147 in PCa 9]–13]. However, one recent study of 11,152 patients found a decrease in CD147 expression
between PCa and BPT and with increasing stage and Gleason score 14], and these results are in agreement with one earlier study on CD147 expression in
PCa 13]. However, these studies concluded that CD147 does not play a significant prognostic
role in determining post-surgical PSA recurrence 14] or that PCa patients with higher CD147 expression performed significantly worse 13]. This is the first study, to our knowledge, to demonstrate that low expression of
CD147 is indicative of poor prognosis after prostatectomy independent of clinico-pathological
features.
One limitation of previous studies is the semi-quantitative approach of evaluating
immunohistochemical staining. While these methods are generally effective in quantitating
“on/off†proteins, this approach is less effective when analyzing proteins with a
heterogeneous staining pattern, particularly when proteins are localized to the membrane
or cytoplasm, as small differences are not readily detected using manual methods of
quantification. In this study, we show that CD147 is primarily localized to the cellular
membrane. Quantitation of E-cadherin in the membrane portion of the epithelium resulted
in a significant decrease in expression in all PCa samples compared to benign prostatic
tissue, while HGPIN and BPH samples showed no significant differences in expression.
Furthermore, membrane-associated E-cadherin expression predicted biochemical recurrence
after prostatectomy in our patient cohort. This data is in agreement with previous
studies on E-cadherin in PCa 16], 28], and thus validates the epithelial cell membrane segmentation for investigation of
CD147.
Antibodies to N-terminal synthetic peptides or recombinant fragments of CD147 have
shown predominant localization to the basal and lateral plasma membrane, and studies
using these antibodies have shown similar decreases in CD147 expression along PCa
progression 13], 14]. As has been argued previously 14], the use of different antibodies may account for some of the contrasting results
found in current literature on CD147. Some previous studies have shown cytoplasmic
and nuclear staining of CD147 in PCa specimens 11], rather than expected membrane-associated localization. Like previous studies with
similar localization and expression results 13], 14], CD147 protein expression in our cohort of patient samples was largely present in
the plasma membrane. Additionally, our immunoblot results showed bands at molecular
weights consistent with both glycosylated and nonglycosylated forms of CD147. When
considering the known roles and expected localization of CD147 5], 27], our results suggest high antibody specificity for known forms of CD147.
Our results differ from previous studies on CD147 and other malignancies 7], 8]. This may be attributable to the diverse roles and unusually high amount of post-translational
modifications to CD147. The mechanistic role of CD147 of inducing MMP expression and
promoting extracellular matrix degradation and reconstruction has implicated CD147
in tumor invasion. Caveolin-1 is a putative tumor suppressor in other malignancies
26], 29], 30] that binds CD147 and suppresses N-glycosylation and CD147-induced fibroblast MMP
activation. Caveolin-1 serves a unique role in PCa, as secretion and overexpression
of caveolin-1 is associated with advanced disease 31]. Though the interaction of caveolin-1 and CD147 was not investigated in this study,
the overexpression of caveolin-1 in PCa progression may serve to bind CD147 and preventing
N-glycosylation, self-aggregation, and MMP induction, thus accounting for the decrease
in CD147 expression in advanced PCa that we observed.
Furthermore, the N-terminal domain of CD147 is essential for MMP induction 27], as N-glycosylation of CD147 is associated with self-aggregation and MMP expression
26]. In HT1080 fibrosarcoma and A431 epidermoid carcinoma cells, purification of a 22-kDa
CD147 fragment from culture medium demonstrated cleavage and shedding of CD147 by
membrane-type MMPs in the linking region between the two Ig-like domains 27]. This soluble 22-kDa fragment was adequate for augmentation of MMP-2 expression in
human fibroblasts, while the cleavage of membrane CD147 is expected to downregulate
the membrane-specific MMP-activating function of CD147 27]. N-terminal cleavage and shedding of CD147 has not yet been studied in PCa, but may
provide a mechanistic explanation for the decrease in CD147 with PCa progression that
was observed in this and previous studies 13], 14], as antibodies to the N-terminal Ig1 domain may not recognize cleaved forms of CD147.
In this study, we also found that BSG mRNA expression was not significantly associated with recurrence, indicating that
post-translational modifications of CD147 may be important in the metastatic spread
of PCa. Further studies are needed on post-translational modified forms of CD147 in
PCa to reconcile the opposing results found throughout the literature.
Limitations of this study include both sample size and the use of tumor-adjacent normal
as a baseline for comparison of expression. Compared to previous studies 14] our sample size is relatively low, but the use of a quantitative multispectral platform
increases the sensitivity of our assay and helps to make our sample size sufficient
to draw conclusions. The use of tumor-adjacent normal tissues is common practice due
to the difficult of obtaining truly normal tissues. However, future studies should
incorporate autopsy normal prostate tissues to avoid the well-documented field effects
of tumors.