In this hypothesis generating study ALCAM and s-ALCAM expression levels in tumor tissue and sera of cervical carcinoma patients were evaluated for the first time. Here we were able to show that ALCAM expression in tumor cells might be a predictive marker for response to chemoradiation in this cancer entity.
By evaluating ALCAM reactivity in tumor cells we were able to show that ALCAM expression is present in a substantial portion of tumors but does not correlate with prognostic markers like clinical stage, grading, age or histological type, or with patients' survival. Although few other groups distinguish between cytoplasmic and membranous ALCAM expression in neoplastic tissue following the idea that a stronger cytoplasmic expression pattern might be abnormal and associated with tumor progression [19
], we refrained from separating ALCAM expression in membraneous and cytoplasmic staining for two reasons: first we found that cytoplasmic and membranous ALCAM staining correlated positively with each other, second, in cases of intense ALCAM expression it is often difficult to assess the staining intensity of each compartment properly (Additional file 1
: Figure S1 and unpublished data).
Prior studies in other cancer entities have reported that ALCAM is upregulated in some and downregulated in others [8
]. Contrary to our results high ALCAM expression in the primary tumor was found to be associated with reduced survival or unfavourable prognostic markers in some tumor types, i.e. colorectal [4
], oral [10
], esophageal [11
], pancreatic [22
] and gastric carcinomas [23
] as well as neuroblastoma [24
]. However, similar to previous studies in breast cancer, our results indicate that ALCAM might represent a predictive marker in cervical cancer: In patients who did not receive any further therapy after tumor excision, high ALCAM expression levels were associated with shorter CSS and DFS, whereas in patients receiving adjuvant chemoradiation high ALCAM expression was indicative of a better prognosis. In a prior study in breast cancer patients we analysed ALCAM expression in tumor tissues on protein and mRNA level using c-DNA microarray data and western blot analysis [12
]. Similar to the present data obtained in cervical carcinomas, ALCAM expression was not relevant for prognosis in the total breast cancer cohort. However, in the subgroup of chemotherapy-treated patients, high ALCAM expression levels were associated with significantly longer DFS and CSS. These results could be at least partly explained by our experimental study where ALCAM expression was increased or silenced by stable transfection in breast cancer cell lines: In those cells, the consequences of high ALCAM expression were complex and included enhanced invasive potential (which might result in a more aggressive tumor growth) and increased apoptosis (which might lead to higher chemosensitivity and a better prognosis in chemotherapy-treated patients) [25
]. In pancreatic cancer cells, a reduced ALCAM expression has also been shown to be associated with chemoresistance, in that case to gemcitabine and actinomycin D [13
]. The results of our present study suggest that the consequences of high ALCAM expression might be similar in cervical cancer cells.
Comparison of the subcohorts treated with radiotherapy alone and with chemoradiation suggests that ALCAM expression is more predictive for successful chemotherapy compared to radiation. The mechanisms of resistance to radiotherapy and resistance to chemotherapy share similar features (role of apoptosis) as well as different characteristics (i.e. drug activation and accumulation in chemoresistance, and DNA repair mechanisms in radioresistance). Our results indicate that both mechanisms might be influenced by ALCAM in different, as yet unknown ways.
Since ALCAM is shedded into the blood by proteases, we also analysed s-ALCAM levels in a subcohort of 55 patients. The resulting s-ALCAM levels in our patients (mean 28.9 ng/ml) are below those reported for breast cancer patients (74 ng/ml; [15
]) and ovarian cancer patients (mean 44 ng/ml; [14
]), but slightly above those found in patients with esophageal carcinoma (mean 23.9 ng/ml; [26
]). Healthy control groups showed mean s-ALCAM levels of 60 ng/ml [15
], 20.6 ng/ml [26
] and 29 ng/ml [14
], respectively. During our experiments, there was no control group available which was comparable to the patients with respect of sex, age and sample storage. Yet, the low s-ALCAM levels in our cervical cancer cohort and the lack of association of s-ALCAM values with ALCAM immunostaining in cancer tissue or patients' outcome suggests that s-ALCAM is not an acceptable serum biomarker in this tumor type.
For squamous cell carcinomas of the cervix or other sites there are only a few studies available, which provide predictive markers for response to radiotherapy or chemotherapy, mainly by analysing cDNA expression arrays [27
]. Adhesion molecules have been identified to be involved in chemoresistance in some prior studies on different cancer types: In neuroblastoma cell lines, acquired resistance to vincristine and doxorubicine was associated with downregulation of the neural cell adhesion molecule (NCAM) [30
]. Similarly, paclitaxel-resistant ovarian carcinoma cell lines showed a decreased expression of the epithelial adhesion molecule E-cadherin [31
]. ALCAM silencing has been shown to induce chemoresistance in pancreatic cancer cells in vitro [13
]. In breast cancer patients [12
] and in this study analysing cervical cancer patients, ALCAM expression in cancer tissue correlates with increased sensitivity to chemoradiation or chemotherapy which might at least partly result from its influence on apoptosis [25
Limitations of this study are its retrospective design and the heterogeneous group of patients with relatively small numbers in the treatment groups. However, the heterogeneity in this first, hypothesis-generating study on ALCAM expression in cervical cancer led to interesting data which suggest that the impact of ALCAM overexpression might be treatment-dependent. To further investigate the potential predictive value of ALCAM expression for chemoradiation in cervical cancer, it will be necessary to investigate ALCAM expression levels in a prospective manner, using i.e. tissue biopsies of patients with advanced disease (FIGO III/IV) to analyse outcome after chemoradiation.