Background
Soft tissue sarcomas are a heterogeneous group of rare malignancies often having poor outcome [
1]. Soft tissue sarcomas constitute less than 1 % of all cancers [
1] while there are more than 50 histological subtypes with sometimes overlapping histological features [
2]. Distinction is essential as subtypes differ in biological behaviour and sensitivity to chemotherapy, and as such an adequate histological diagnosis, is crucial for clinical decision making [
3]. Fifty-six percent of soft tissue sarcomas present as localized disease at the time of diagnosis, and surgery is the mainstay of treatment, sometimes combined with radiotherapy or chemotherapy [
4].
From the molecular point of view, soft tissue sarcomas can be distinguished into two categories. The first class includes sarcomas with a simple genome, in which recurrent translocations, amplifications or specific mutations can be found. The second class includes sarcomas with a complex genome, characterized by a multitude of chromosomal alterations and genomic instability, often reflected by pleomorphic histological features [
3]. This group includes high grade leiomyosarcoma, myxofibrosarcoma, undifferentiated pleomorphic sarcoma, undifferentiated spindle cell sarcoma, pleomorphic liposarcoma, and pleomorphic rhabdomyosarcoma.
Leiomyosarcomas constitute 5–10 % of all soft tissue sarcomas, displaying smooth-muscle differentiation [
1]. Studies showed for leiomyosarcoma that the metastasis-free 5-year survival rate is about 60 % [
5]. Histological grade is the most important prognostic factor for most soft tissue sarcomas. By using FNCLCC grading system, which is the most widely used 3-grade system, soft tissue sarcomas are divided into low, intermediate and high grade based on the sum score of three histologic parameters including tumor differentiation, mitotic count and tumor necrosis. About 65 % of leiomyosarcomas are reported to have high-grade areas [
6]. High grade leiomyosarcomas often have poor patient outcome [
4]. Until now, the genetics and pathology of leiomyosarcomas are not completely understood and as they have a complex genome, no molecular diagnostic tests or specific therapeutic targets are available. Hence, there is a strong need for new molecular markers that can aid in the stratification of leiomyosarcomas patients with respect to their disease outcome.
In a previous study, we used imaging mass spectrometry to compare these soft tissue sarcomas with a complex genome. A panel of protein signatures that could distinguish between different subtypes, or were associated to patient survival were discovered [
7]. Among them, proteasome activator complex subunit 1 (PSME1) was found indicative of poor survival in soft tissue sarcomas. PSME1 (also known as REGalpha and PA28A), is a multicatalytic proteinase complex, implicated in immunoproteasome assembly and required for efficient antigen processing [
8]. Intriguingly, PSME1 was also found to associate with diagnosis or prognosis in other tumor types, e.g. prostate cancer [
9], breast cancer [
10] and ovarian cancer [
11,
12].
In this study, we used tissue microarrays of soft tissue sarcomas with complex genomes, to evaluate whether PSME1 expression can predict clinical outcome in soft tissue sarcomas, especially leiomyosarcomas.
Discussion
Using imaging mass spectrometry we previously identified PSME1 as a prognostic biomarker indicating poor survival in soft tissue sarcoma patients [
7]. Imaging mass spectrometry is a sensitive discovery tool (zepto-molar sensitivity [
23]) enabling the detection of hundreds of molecules directly from tissue [
24,
25]. To further explore the prognostic value of PSME1 we analysed PSME1 expression in a larger, independent set of soft tissue sarcomas using immunohistochemistry on tissue microarrays. PSME1 (or PA28A) encodes a subunit of the proteasome system, which is a major source for generation of tumor antigens presented by MHC class I molecules [
26,
27]. Escape of immune response is one of the hallmarks of cancer [
28]. In addition, elevated proteasome activity in tumor cells has been described to influence transcription factors involved in cell survival or apoptosis [
29,
30]. Novel strategies using the proteasome have been proposed for cancer treatment for example by alternating the NAD
+/NADH ratio to change kinetics of proteasomal degradation [
30] or inhibiting proteasome to induce apoptosis [
31‐
34].
PSME1 is expressed in many different cell types, especially antigen presenting cells, and its expression can be controlled by interferon gamma. Both chemotherapy and TNF-alpha may induce a local inflammatory reaction within the tumor microenvironment and therefore may influence expression of PSME1. It is of interest that all sarcoma subtypes included in our study expressed PSME1 to a variable extent, while neoadjuvant chemotherapy or treatment with interferon gamma is not standard practice in our hospital. As far as clinical data were available, only four patients received preoperative chemotherapy or TNF-alpha, and expression levels were not significantly different. In the control group, consisting of uterine leiomyomas, expression was low to absent, both in the nucleus as well as in the cytoplasm.
High PSME1 expression was also described in other tumors. For example, increased PSME1 expression was also found in primary and metastatic human prostate cancer and was suggested as a potential target for therapeutic intervention [
9]. PSME1 was previously also detected using imaging mass spectrometry in other tumors: Dekker et al. detected PSME1 as a marker of stromal activation in breast cancer [
10]. Previous studies also showed that PSME1 could be a molecular signature to discriminate between benign and malignant ovarian tumors [
11,
35], and an early diagnosis and tumor-relapse biomarker [
12]. Zhang et al. detected PSME1 as a tumor marker in human oesophageal squamous cell carcinoma [
36]. The proteasome can be present in the cytoplasm as well as in the nucleus of all eukaryotic cells, although their distribution and function can be variable [
17]. We here show that in soft tissue sarcomas with a complex genome, PSME1 is expressed both in the cytoplasm and in the nucleus. Proteasome-dependent protein degradation is important in the cytoplasm for MHC class 1 antigen presentation [
8]. In the nucleus, PSME1 plays an important role in maintaining the nuclear function including gene expression and cell proliferation [
19,
37].
To further evaluate its clinical relevance, we analysed the largest subgroup, comprising 34 leiomyosarcomas of different histological grade, in more detail. Both nuclear as well as cytoplasmic expression of PSME1 significantly increased with increasing histological grade. Moreover, high nuclear expression of PSME1 was significantly associated to poor outcome (overall survival, metastasis-free survival and event-free survival) in leiomyosarcoma patients, although the patient cohort is rather small (n = 34). In multivariate analysis only the association with decreased metastasis-free survival was independent of histological grade, while an independent association to poor overall survival and decreased event-free survival was at the border of significance. Although PMSE1 expression is a promising biomarker, our results need to be validated in an independent cohort of leiomyosarcomas.
In summary, we found elevated expression of the proteasome subunit PSME1 in leiomyosarcomas compared to control tissues, and an association of the expression with increasing histological grade in leiomyosarcoma. Moreover, high nuclear PSME1 expression was found to be an independent predictor of metastasis-free survival in leiomyosarcoma patients. Our results suggest that the expression of proteasome subunits such as PSME1 could be taken into account for leiomyosarcoma patients when considering immunotherapeutic strategies in these tumors [
38].
Authors’ contributions
SL: data analysis and interpretation, writing of manuscript. AHGC: immunohistochemistry scoring, data interpretation. BB: statistical analysis. MG: tissue microarrays’ construction. MK: data collection and analysis. IBB: experimental work. LAM: design and supervision of the study. JVMGB: immunohistochemistry scoring, design and supervision of the study, writing of manuscript. All authors read and approved the final manuscript.