Prognostic and treatment predictive significance of SATB1 and SATB2 expression in pancreatic and periampullary adenocarcinoma

Periampullary adenocarcinomas encompass tumours originating in or adjacent to the ampulla of Vater; pancreatic cancer, distal bile duct cancer, ampulla of Vater carcinoma and carcinoma of the periampullary duodenum. Pancreatic cancer is the most common type of periampullary adenocarcinoma, but only a minority can be resected with a curative intent, due to either locally advanced growth or distant metastases at presentation. There are two major morphological types of periampullary adenocarcinomas, which have different prognosis and receive different chemotherapy. Pancreatobiliary type (PB-type) adenocarcinomas include pancreatic cancer, distal bile duct cancer, and some of the ampullary carcinomas. They have a dismal prognosis even after resection and adjuvant gemcitabine-based chemotherapy. Intestinal type (I-type) periampullary adenocarcinomas include duodenal carcinoma and some of the ampullary carcinomas. They have a better prognosis but little is known on risk stratification and optimal chemotherapy [1],[2]. Hence, new biomarkers are needed to better stratify both PB-type and I-type periampullary adenocarcinomas according to risk and expected response to treatment.

Special AT-rich sequence-binding protein 1 (SATB1) is a genome organizing protein which regulates region-specific epigenetic modifications and expression of a large number of genes, and special AT-rich sequence-binding protein 2 (SATB2) is a close homologue with similar functions [3]-[5].

SATB1-expression has been demonstrated to confer a more aggressive tumour phenotype and a shorter patient survival in several cancer forms, e.g. breast cancer [3], prostate cancer [6], laryngeal squamous cell carcinoma [7], nasopharyngeal cancer [8], hepatocellular carcinoma [9], rectal cancer [10], cutaneous malignant melanoma [11], epithelial ovarian cancer [12], glioma [13] and gastric cancer [14].

The SATB2 gene is involved in osteoblast differentiation and craniofacial patterning [15],[16] and has been demonstrated to be abundantly expressed in normal colorectal mucosa and colorectal adenocarcinomas, but more sparsely in other types of carcinomas [17]. Low or absent SATB2-expression has further been shown to be a marker of malignant behaviour and poor prognosis in colorectal cancer [18],[19], whereas high expression correlated to a better response to neoadjuvant chemotherapy in rectal cancer and neoadjuvant/adjuvant chemotherapy in stage III-IV colorectal cancer [20].

The expression and prognostic significance of SATB1 and SATB2 in pancreatic, distal bile duct, ampullary or duodenal adenocarcinomas has not yet been reported. The aim of the present study was therefore to examine the expression, clinicopathological correlates, and prognostic and treatment predictive ability of SATB1 and SATB2 in primary tumours (n = 175) and paired lymph node metastases (n = 105) from a consecutive cohort of patients with periampullary adenocarcinoma, including pancreatic cancer.

The results from this study provide a first demonstration of the expression and prognostic value of SATB1 in pancreatic, distal bile duct, ampullary and duodenal adenocarcinoma. Positive SATB1-expression was observed in 20% of resected PB-type cases, and was associated with a shorter RFS and OS, which is in line with previous publications on the prognostic significance of SATB1 expression in several other major types of cancer [3],[7],[9]-[12],[14]. The findings from the present study thus provide further evidence of SATB1 being a master regulator towards a more aggressive tumour phenotype and a biomarker of poor prognosis in human cancer. In addition, the finding of a potential treatment predictive role of SATB1, its expression being associated with a better response to adjuvant gemcitabine in PB-type tumours, reflected in a prolonged 5-year survival, and an improved response to any adjuvant chemotherapy in I-type tumours, reflected in a prolonged recurrence-free survival, has however not yet been described in any type of cancer. Patients with pancreatic and periampullary adenocarcinomas have a very dismal prognosis even after surgical removal of the tumour. According to contemporary treatment protocols, all patients with pancreatobiliary type adenocarcinoma, including pancreatic cancer are recommended adjuvant treatment, and adjuvant chemotherapy with gemcitabine has recently been shown to increase overall and disease-free survival among patients with radically resected tumours [22]. A challenging task is however to identify which patients will actually benefit from this treatment and not only suffer from the adverse side effects resulting in a reduced quality of life. The here examined retrospective cohort consists of a comparatively large proportion of patients who did not receive any adjuvant chemotherapy, which is in part likely due to the fact that all types of periampullary adenocarcinomas are included. As shown in Table 2, tumour origin and year of surgery differs between the gemcitabine and non-gemcitabine groups of PB-type tumours. During the first part of the included period (2001–2011), the distinction between pancreatobiliary and intestinal tumour morphology was not made, and decision on adjuvant chemotherapy seems to have been based mainly on tumour origin. Many PB-type ampullary tumours did thus not receive adjuvant chemotherapy and tumours of distal bile duct origin were given adjuvant chemotherapy less often than tumours of pancreatic origin. For intestinal type tumours, decision on adjuvant chemotherapy seems to have been based primarily on involved lymph nodes, as this is the only parameter that differs significantly between the group that received and those that did not receive adjuvant chemotherapy. Although treatment predictive effects are best studied in a randomized setting, the nearly equal distribution of patients treated or not treated with adjuvant chemotherapy in this retrospective cohort provides a better setting for discovery of potential treatment predictive markers than studies on cohorts where all patients have received adjuvant chemotherapy.

Apart from considerations in the adjuvant situation, SATB1 could also prove to be a useful biomarker for identification of patients with borderline resectable tumours who will respond well to neoadjuvant chemotherapy, thus increasing the number of resectable tumours. Therefore, the indication of a treatment predictive value of SATB1 expression in periampullary adenocarcinoma is of high potential clinical relevance and merits further validation in additional patient cohorts. The mechanistic basis for SATB1-related increased sensitivity to various combinations of chemotherapy should also be pursued in future studies.

Given the high homology of SATB1 and SATB2, it is important to use well-validated antibodies to ensure target specificity. The antibodies used in the present study have been validated previously [23] and cross-reactivity should therefore not be an issue.

SATB1 was often heterogeneously expressed and the number of positive cells was often low, which justifies assessment not only of the primary tumour, but also metastases in order to improve the detection of positive cases.

Immunohistochemistry has several advantages compared to e.g. analyses of mRNA levels in that it allows for assessment of candidate protein biomarkers in a morphological and subcellular context. The results from this study are quite in line with several previous studies demonstrating that even a small fraction of SATB1 positive cells by immunohistochemistry is sufficient to confer a poor prognosis [3],[12]. Moreover, results from studies on the prognostic value of mRNA levels of SATB1 have shown discrepant results in relation to its protein expression in e.g. breast cancer [24],[25]. A likely explanation for this is the more or less abundant expression of SATB1 in activated lymphocytes, also serving as a positive internal control in immunohistochemical studies. Therefore, immunohistochemistry should be the method of choice for assessment of the utility of SATB1 as a prognostic and treatment predictive biomarker in human cancer.

Some methodological aspects on the TMA technique need consideration. Although heterogeneity issues cannot be fully circumvented, it is reasonable to assume that analysis of whole tissue sections will lead to an improved detection rate of positive primary tumours and/or metastases. However, while the use of whole sections is feasible in the clinical setting and in prospective studies, the TMA technique has become a well-established platform for high-throughput tissue biomarker studies in the retrospective setting, and has been demonstrated to provide similar or even better prognostic information for heterogeneously expressed markers than whole section-based analyses [26]. Moreover, a comparative strength of the here used TMA is that tissue cores had, whenever possible, been obtained from different donor blocks of the primary tumours, and from different lymph node metastases in cases with more than one metastasis.

In a previous study related to the potential utility of SATB2 as a diagnostic marker for colorectal cancer, screening of its expression in a multitude of normal and cancerous tissues revealed that none out of 25 pancreatic adenocarcinomas and only one out of 15 bile duct adenocarcinomas were positive for SATB2 [17], which is in line with the finding in the present study of SATB2 being positive in only three out of 107 pancreatobiliary type adenocarcinomas. Although a significant association was found between SATB2 expression and poor prognosis, the small number of positive cases makes it hazardous to draw any conclusions on the potential prognostic value of SATB2 expression in PB-type tumours. In the group of I-type tumours, where SATB2 expression was more frequent (8/61), no prognostic effect was seen. Moreover, while there were no significant treatment predictive effects of SATB2 expression in I-type adenocarcinoma, it is noteworthy that SATB2-positive I-type cases receiving adjuvant chemotherapy had no recurrences or fatalities during the follow up period. This observation may suggest a similar treatment predictive function for SATB2 in I-type tumours as observed for SATB1 and would also be in line with the previously described treatment predictive function of SATB2 in colorectal adenocarcinoma [20]. Along this line, while some studies have suggested antagonistic effects of SATB1 and SATB2 [5],[18],[23], it cannot be ruled out that SATB1 and SATB2 both increase chemotherapy sensitivity in the here examined types of cancer.

Expression of SATB1 is associated with poor prognosis in pancreatobiliary type adenocarcinomas, and predicts response to adjuvant treatment in both intestinal type and pancreatobiliary type periampullary adenocarcinomas, including pancreatic cancer. These findings are of potential clinical relevance and merit further validation in additional patient cohorts as well as in a mechanistic context.