The diversity between pancreatic head and body/tail cancers: clinical parameters and in vitro models

Hangzhou, China

The diversity between pancreatic head and body/tail cancers: clinical parameters and in vitro models

Qi Ling, Xiao Xu, Shu-Sen Zheng and Holger Kalthoff

Hangzhou, China

BACKGROUND:Pancreatic ductal adenocarcinoma (PDAC) can be divided into head, body and tail cancers according to the anatomy. Distinctions in tissue composition, vascularization and innervations have been clearly identif i ed between the head and body/tail of the pancreas both in embryological development and in histopathology. To understand the postulated genotype difference, we present comprehensive information on two PDAC cell lines as typical representatives originating from pancreatic head and body/tail cancers, respectively.

DATA SOURCE:In the present review, we compare the difference between pancreatic head and body/tail cancers regarding clinical parameters and introducing anin vitromodel.

RESULTS:Increasing evidence has shown that tumors at different locations (head vs body/tail) display different clinical presentation (e.g. incidence, symptom), treatment eff i ciency (e.g. surgery, chemotherapy) and thus patient prognosis. However, the genetic or molecular diversity (e.g. mutations, microRNA) between the two subtypes of PDAC has not been elucidated so far. They present different chemo- and/or radio-resistance, extracellular matrix adhesion and invasiveness, as well as genetic prof i les.

CONCLUSION:Genetic and tumor biological diversity exists in PDAC according to the tumor localization.

(Hepatobiliary Pancreat Dis Int 2013;12:480-487)

pancreatic ductal adenocarcinoma;tumor location; cell lines; diversity

Introduction

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal human cancers. It has a high metastatic potential and the majority of diseases present themselves at a very late stage. PDAC can be divided into head and body/tail cancers according to the anatomy. Pancreatic development begins with the formation of a ventral and a dorsal bud, which becomes the ventral head (lower head and uncinate process) and dorsal pancreas (upper head, body and tail), respectively. This difference in ontogeny leads to signif i cant differences in cell composition, blood supply, lymphatic and venous backf l ow, and innervations between the head and body/tail of the pancreas. For instance, the number of Langerhans Islets is greater in the body and tail. Insulin-positive endocrine cells are highly refractory to malignant transformation under normal conditions, but could serve as a cell-of-origin of PDAC under oncogenic activation in combination with pancreatic injury.[1]In addition, fatty tissue inf i ltration is usually most prominent in the anterior aspect of the pancreas head and may stimulate pancreatic neoplasm.[2,3]Branch duct intraductal papillary mucinous neoplasms are typically arising in the head of the pancreas while mucinous cystic neoplasms are common in the body or tail.[4]In this sense, the head and body/tail of the pancreas may have different malignant potential.

In pancreatic serous cystic neoplasms and intraductal papillary mucinous neoplasms, tumorlocation in the head of the pancreas was independently associated with local invasiveness and recurrence,[5,6]while in pancreatic neuroendocrine tumors, tumors located at the body/tail of the pancreas were more likely to be associated with shorter progression-free survival.[7]In colon cancer, a number of studies have demonstrated that right- and left-sided tumors exhibit different genetic, biological and demographical characteristics and risk factors, suggesting that the carcinogenesis and tumor progression of colon cancer may differ with tumor localization.[8-10]These fi ndings support the hypothesis of different mechanisms in carcinogenesis of tumors at different locations and conf i rm the importance of subsite division. Although increasing evidence has shown differences in clinical presentation between pancreatic head and body/tail cancers,[11-15]the genetic or molecular diversity between the two subtypes of PDAC has not been elucidated so far.

In the present review, we compare the clinical data between pancreatic head and body/tail cancers. In particular, to assess the potential phenotype and genotype difference, we provide comprehensive information on two frequently used PDAC cell lines that originate from pancreatic head and body/tail cancers, respectively.

Comparison of clinical and diagnostic parameters between pancreatic head and body/tail cancers

Incidence

Data from Surveillance, Epidemiology, and End Results (SEER) registries of the United States (1973-2002) have shown that about 77.5% (34 072/43 946) of PDACs originate at the head of the pancreas, on which consequently most discussion on pancreatic cancer has been focused.[11]The overall annual incidence of pancreatic head cancer is 5.6 per 100 000, compared with 1.6 of pancreatic body/tail cancer.[11]Data from National Pancreatic Cancer Registry of Japan (1981-2002,n=9290)[13]and Eindhoven Cancer Registry of Netherland (1995-2000,n=1128)[14]also demonstrated much higher incidences of pancreatic head cancer (62.3% and 56.5%, respectively) than body/tail cancer (17.5% and 12.7%, respectively). For resectable tumors (Stage I: T1N0M0 and T2N0M0), about 69.8% (6676/9559) are located in the head of the pancreas as shown by the United States National Cancer Data Base, 1995-2004.[12]

Early diagnosis

Symptoms often do not appear until the disease is in an advanced stage, thus making early detection diff i cult. Notably, a patient's symptoms will vary depending on the location of the tumor within the pancreas. Both pancreatic head and body/tail cancers can cause non-specif i c symptoms, such as abdominal pain, nausea, loss of appetite and weight loss. However, only tumor blocking the bile ducts, which pass through the head of the pancreas, can cause jaundice. A study from China investigated the clinicopathological characteristics between pancreatic head cancer (n=541) and body/tail cancer (n=106) from 1980 to 2003. It was found that patients primarily diagnosed with pancreatic body/tail cancer were associated with much less jaundice but more pain, higher serum albumin level, higher carcinoembryonic antigen (CEA) but lower carbohydrate antigen 19-9 (CA19-9) positivity, and higher metastasis rate.[15]Another study from Japan reviewed 209 PDAC patients and showed that patients with pancreatic body/tail cancer were signif i cantly more likely to have abdominal pain but less likely to have jaundice.[16]Other studies conf i rmed that patients with pancreatic body/tail cancer were more likely to be diagnosed at an advanced stage.[11,17]SEER registries database (1973-2002) reported that patients with pancreatic body/tail cancer have a higher proportion of the distant stage diseases (72.7%) compared to patients with pancreatic head cancer (39.2%).[11]

Diabetes is a risk factor of pancreatic cancer. A meta-analysis demonstrated that individuals in whom diabetes had only recently been diagnosed (<4 years) exhibited a 50% higher risk of the malignancy compared with individuals who had diabetes for ≥5 years.[18]On the other hand, pancreatic cancer can induce a diabetic status.[18]Theoretically, patients with pancreatic body/ tail cancer are more prone to have islet dysfunction and subsequent diabetes, since islet concentration is much higher in the tail than in the head of the pancreas. But the data comparing the onset of pancreatic cancer-induced diabetes between the head, body, and tail of the pancreas are limited. Of note, postoperative diabetes may be higher in patients receiving distal pancreatectomy than those treated with pancreaticoduodenectomy.[19]

Perfusion computed tomography (CT) and dynamic contrast-enhanced magnetic resonance imaging (DCEMRI) can provide important information in the diagnosis of pancreatic cancer. Perfusion CT allows measurement of tumor vascular physiology including tumor blood fl ow (BF), blood volume (BV), mean transit time, and vascular permeability surface area product (PS).[20]It has been reported that no signif i cant difference in perfusion values (BF, BV and PS) was found between the head, body, and tail of the pancreas.[20,21]However, there is lack of studies comparing the imaging modalities between the head, body, and tail of the pancreas.

Survival

It is not surprising that pancreatic head cancer has a better overall patient survival than pancreatic body/tail cancer, because more patients with pancreatic body/tail cancer are diagnosed at a relatively advanced stage. Data from SEER database (1988-2004) including 33 752 patients with pancreatic cancer presented a signif i cantly lower rate of cancer-related surgery (16% vs 30%) and also a signif i cant lower median survival (4 months vs 6 months) in patients with body/tail cancer compared with those with head cancer.[22]The SEER registries database (1973-2002) including 43 706 cases of pancreatic cancer showed that pancreatic head cancer had a 4% lower overall mortality risk compared with pancreatic body/ tail cancer in multivariate COX analysis.[11]However, not consistent with the results from Western countries, data from the National Pancreatic Cancer Registry of Japan showed a signif i cantly lower 5-year survival rate (10.7% vs 13.8%) for patients with pancreatic head cancer (n=5788) than those with pancreatic body/tail cancer (n=1629).[13]

As long as operation is possible, pancreatic head cancer is surgically treated by pancreaticoduodenectomy, whereas pancreatic body/tail cancer is treated by a distal pancreatectomy. As mentioned above, more tumors are diagnosed at an early stage and the resectability is higher in pancreatic head cancer. The SEER database (1988-2004) including 5118 and 663 patients with resected pancreatic head and body/tail cancers, respectively, revealed that tumor location at the body/tail of the pancreas was an independent negative predictor of survival in patients after surgery.[22]Of note, most of the patients undergoing resection were not at an early stage: 53% showed with regional lymph node metastasis and 74% were at T-stage 3/4.[22]Within the same local-stage, pancreatic head cancer had a much lower 3-year survival rate than pancreatic body/tail cancer (9% vs 20%).[11]Moreover, a Japanese study enrolling 80 consecutive patients with resectable pancreatic cancers presented similar overall survival and recurrence rates after a curative resection between patients with pancreatic head cancer (n=56) and those with pancreatic body/tail cancer (n=24).[23]

Although surgical resection remains to be the only potential cure for PDAC, only a small proportion of patients newly diagnosed with PDAC are considered for surgical resection. Chemo- and/or radiotherapy has emerged as a key factor to improve patient outcome. A retrospective study from Japan including 66 patients with metastatic PDAC who were treated with gemcitabine revealed tumor location at the head of the pancreas as an independent risk factor of poor prognosis after chemotherapy.[24]Similarly, a study from France using a cohort of 42 patients with metastatic PDAC treated by fl uorouracil/leucovorin combined with irinotecan and oxaliplatin (FOLFIRINOX) demonstrated that only tumor location in the head of the pancreas was associated with poor outcome.[25]However, some other reports showed that tumor site (head vs body/tail) did not relate to progression freesurvival or overall survival in patients with advanced or metastatic pancreatic cancer treated with chemo- and/ or radiotherapy.[26-28]So far, no large-sample and welldesigned study has yet been conducted for comparing the different response to chemo- and/or radiotherapy such as toxicity and tumor progression between different tumor sites. The association between tumor localization and response to chemo- and/or radiotherapy needs to be further evaluated.

Diagnostic characterization of genetic markers

Microarray-based approaches have allowed genomewide, high throughput screening for novel candidate genes or microRNAs (miRNAs) involved in the pathogenesis of PDAC. Most recently, as a consequence of technical advances, whole sequencing of the cancer exome has been performed and leads to a greater insight into the mutational spectrum of human cancers. In 2008, a comprehensive genetic analysis containing 24 PDACs determined an average of 63 genetic alterations in the sequences of 23 219 transcripts, representing 20 661 protein-coding genes. These alterations def i ned a core set of 12 cellular signaling pathways and processes implicated in the deregulation that gives rise to the major features of pancreatic tumorigenesis.[29]When the data from 24 PDAC patients (head/tail: 19/5) were correlated with several clinical parameters (e.g. age, gender, tumor location), no signif i cant difference was found in the numbers of mutations (63.1±32.7 vs 72.6 ±24.0 per tumor), deletions (8.4±4.8 vs 7.8±3.1 per tumor), or amplif i cations (5.2±6.0 vs 9.0 ±10.9 per tumor) between pancreatic head and tail cancers.[30]However, details on point mutations in relation to topology were not given and the study cohort was not appropriately matched between patients with pancreatic head and tail cancers.

The incidence of K-ras point mutations was not different signif i cantly between pancreatic head cancer (79%, 22/28) and body/tail cancer (72%, 13/18) as shown by combining the data from a Japanese and a Chinese study.[31,32]A recent report from USA described 2 patients presented with synchronous PDACs arising from intraductal papillary mucinous neoplasm (each with one tumor in the head and the other inthe tail of the pancreas). They found both tumors from each patient shared identical K-ras mutations but different clonal genetic instability as ref l ected by loss of heterozygosity analysis, indicating that molecular diversity existed between tumors at different localizations even if the synchronous tumors are likely initiated from the same precursor lesions.[33]

MicroRNAs play key roles in diverse biological processes, and accordingly, unique expression prof i les of miRNAs have been found in PDAC. Aberrant expression of a number of miRNAs was closely associated with tumor development, inf i ltration, metastasis and poor prognosis.[34,35]However, no study has taken tumor location into consideration as part of the miRNA analysis to date. We could only obtain the data on circulating miR-210 expression from a study containing 16 and 6 patients with locally advanced unresectable stage T4 PDACs in the head and body of the pancreas, respectively. Here, miR-210 was found to be higher in patients with pancreatic body cancer than those with pancreatic head cancer.[36]Given the lack of detailed information on potential genetic differences between pancreatic head and body/tail cancers, we propose anin vitrosystem based on the comparison of two PDAC cell lines in the following section.

Comparison of pancreatic cancer cell lines originating from pancreatic head and body/ tail cancers

Cancer cell lines ref l ect the genomic events leading to tumor changes seen in clinical tissues and are valuable tools in studies of tumor cell biology.[37]Different PDAC cell lines arising from primary tumors, liver metastasis, ascites, or lymph node metastasis exhibit a great deal of diversity in structure and function. To compare the cell lines derived from pancreatic head and body/tail cancers, we reviewed current information characterizing frequently used PDAC cell lines originating from the primary tumors (BxPC-3, Capan-2, MIA PaCa-2 and Panc-1). Other well known cell lines such as PSN-1 and Panc Tu-1, which were derived from primary tumor but the exact site of origin was not def i ned, were not included.

Panc-1 and MIA PaCa-2, the source of which are matched by donor age (±10 years), tumor stage, histological differentiation and ultrastructural features,[37,38]were selected as representing pancreatic head and body/tail cancers, respectively.

Cell migration and invasion

Tumor cell motility is the hallmark of invasion and is an initial step in metastasis, whereas extracellular matrix (ECM) adhesion is a key mediator in cell motility. Therefore, comparing the ability of cell adhesion, migration and invasion between Panc-1 and MIA PaCa-2 might help in discovering the metastatic potential of pancreatic head and body/tail cancers. Compared to MIA PaCa-2, Panc-1 showed a higher binding aff i nity to collagens (type I and IV), the most abundant protein in ECM, and also exhibited higher adhesion ability to endothelial cells.[38,39]In addition, Panc-1 expressed greater levels of a series of cell adhesion molecules such as E-selectin, intercellular adhesion molecule-1 (ICAM-1), sialyl Lewis a (sLea), sialyl Lewis x (sLex), lymphocyte function-associated antigen-1 (LFA-1) and very late activation antigen-4 (VLA-4).[39]Moreover, Panc-1 exhibited much higher expression of invasion-associated molecules than MIA PaCa-2, such as EphA2.[40]

Actually, invasion is a consequence of the cross talk that occurs between cancer cells and stroma cells. For better mimicking of the tumor environment, a study using a co-culture system with PDAC cells and tumorderived fi broblasts demonstrated that hepatocyte growth factor produced by fi broblasts could initiate an apparent invasion-stimulating response in strongly c-met expressing Panc-1 cells but not in weakly expressing MIA PaCa-2 cells.[41]In this sense, pancreatic head cancer (Panc-1) showed higher metastatic potential than pancreatic body/tail cancer (MIA PaCa-2).

Pro-angiogenic potential

Angiogenesis is critical for tumor growth and metastasis. Vascular endothelial growth factor (VEGF), which principally mediates angiogenesis, can determine the angiogenic switch in cancer. Compared with Panc-1, MIA PaCa-2 secreted lower levels of VEGF.[42]Both Panc-1 and MIA PaCa-2 do not express cyclooxygenase-2 (COX-2) protein, an important mediator of angiogenesis and tumor growth, which may indicate a relative low pro-angiogenic potential.[38]However, comparison of tumor-induced angiogenesis might be less important because PDAC progression and prognosis have been reported to be indeed angiogenesis independent.[43]

Tumorigenicity

Tumorigenicityin vivois an eff i cient way for evaluation of the tumor formation and metastatic potential of cancer cells, and is commonly measured by several parameters such as tumor mass, tumor size, rate of growth and metastasis. However, the tumor formation abilities varied among different methods, such as subcutaneous cancer cell injection, intraperitoneal cancer cell injection, orthotopic tumorimplantation and orthotopic cancer cell injection models.[38]Compared with other methods, orthotopic injection of cancer cell better ref l ects the clinical microenvironment and provides more convincing data. In this model, both Panc-1 and MIA PaCa-2 presented high intra-pancreatic tumorigenicity, local inf i ltration and distant metastasis potential.[44,45]However, data directly comparing the orthotopic tumorigenicity of the two PDAC cell lines are not yet given in this model.

By using immunohistochemical analysis, Panc-1 and MIA PaCa-2 tumors demonstrated quite similar morphology, mucin accumulation, cytokeratin (CK) prof i le (CK7/CK19, CK8/CK18 and CK20), trans (vimentin, chr-A and α1-chym) and dedifferentiation (pdx-1, shh and ptc) patterns.[46]

Chemo- and/or radiotherapy resistance

Chemo- and/or radiotherapy promote cancer cell death primarily by the induction of apoptosis. A number of studies have proven abundant evidence that Panc-1 tolerates much higher half maximal inhibitory concentration (IC50) values for 5-f l uorouracil (5-FU) and gemcitabine, and radiation than MIA PaCa-2.[47-49]In addition, MIA PaCa-2 was more sensitive than Panc-1 to oncolytic therapy and presented a dramatic increase in apoptosis.[47]The possible reason was that the expressions of anti-apoptotic proteins were much higher in Panc-1 than those in MIA PaCa-2.[48,49]Ribonucleotide reductase M2 subunit (RRM2), a gemcitabine-resistant enzyme, was also found to have signif i cantly higher expression in Panc-1 than MIA PaCa-2.[50]Taken together, pancreatic head cancer (Panc-1) is more chemo- and/or radio-resistant than pancreatic body/tail cancer (MIA PaCa-2).

Genetic alterations and expression patterns

Genetic alterations in PDAC are common, both at the cell and tissue levels. Cancer progression through the accumulation of genetic alterations results in a gain of cell growth and proliferation, and subsequently in increased dissemination and metastatic potential. The genetic basis of PDAC is usually elucidated using a candidate gene approach, which has identif i ed the four most frequent mutations. Genetic defects exist inKRAS,TP53andCDKN2A/p16genes but not inSMAD4/DPC4in both cell lines.[37,38,45]

A meta-analysis including a consensus set of 2984 genes from four independent studies strictly compared the gene expression prof i les between PDAC and normal pancreatic tissue, and found 62 differentially expressed genes already known to associate with tumorigenesis of PDAC.[51]Using this database,[51]we compared the PDAC-associated gene prof i les (http://www. pancreasexpression.org/index.html) between Panc-1 and MIA PaCa-2, and found 52 differentially expressed genes which had at least a 2-fold change in expression. Fourteen out of the 52 genes have already been described in detail (related PDAC cells function) and are presented in Table 1.

Table 1.Differentially expressed PDAC-related genes in Panc-1 compared with MIA PaCa-2

Table 2.Differentially expressed PDAC-related miRNAs in Panc-1 compared with MIA PaCa-2

MicroRNA prof i les

Studies have shown specif i cally altered miRNAs between PDAC cell lines and human pancreatic ductal epithelium control cell lines, and between subgroups of PDAC cell lines with different invasion or metastasis properties.[52-54]From these studies covering a large panel of miRNAs, we could fi nd several differentially expressed miRNAs between Panc-1 and MIA PaCa-2, such as miR-10b, miR-15b, miR-18a, miR-21, miR-22, miR-125, miR-155, miR-181 and miR-196a.[52,54]Other studies focusing on a limited number of miRNAs also showed differentially expressed miRNAs between Panc-1 and MIA PaCa-2 such as miR-10a and miR-217.[55,56]The differentially expressed PDAC-related miRNAs as elucidated to date between Panc-1 and MIA PaCa-2 are shown in Table 2.

Conclusions

So far, not enough attention has been paid to the molecular diversity between pancreatic head and body/ tail cancers and the overall information is limited. The current clinical data support the higher incidence, easier detection and better prognosis of pancreatic head cancer compared with pancreatic body/tail cancer. However, for tumors at the early stage, pancreatic head cancer may be associated with a lower survival compared with pancreatic body/tail cancer. Although previous reports describe large cohorts of patients, the evidence is still not so convincing because there is a lack of strictly case-matched comparison between the two subtypes of PDAC. In addition, race or environment may also affect the diversity because of the different results from Western and Eastern countries.

The frequently used PDAC cell lines Panc-1 and MIA PaCa-2, which are matched by donor age (±10 years), tumor stage, histological differentiation and ultrastructural features, to some extent might represent pancreatic head and body/tail cancers. Compared with MIA PaCa-2, Panc-1 is more chemo- and/or radioresistant, which is consistent with the clinical fi ndings that tumor located at the head is associated with poor prognosis after chemotherapy. In addition, Panc-1 has a greater potential of ECM adhesion and cell invasion than MIA PaCa-2, and exhibits a more 'oncogenic' prof i le.

In summary, we might emphasize that diversity exists between pancreatic head and body/tail cancers. This fi nding conf i rms the importance of subsite division and supports the development of individual treatment strategies. Further pioneering studies on strictly matched patient tumor samples are needed for a deep understanding of molecular diversity. Furthermore, genetic animal models may be established in the future to better consider specif i c topological differences in pancreatic tissue microenvironment.

Contributors:ZSS and KH proposed the study conception and design. LQ and XX drafted the manuscript. KH made a critical revision of manuscript. ZSS is the guarantor.

Funding:This work was supported by grants from the Foundation of Zhejiang Educational Committee (20110443), the Health Bureau of Zhejiang provine (201233263), and the Pancreatic Cancer Consortium Kiel (DFG).

Ethical approval:Not needed.

Competing interest:No benef i ts in any form have been received or will be received from a commercial party related directly or indirectly to the subject of this article.

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Received March 7, 2013

Accepted after revision July 6, 2013

AuthorAff i liations:Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Aff i liated Hospital, Zhejiang University School of Medicine; Key Lab of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou 310003, China (Ling Q, Xu X and Zheng SS); Institute for Experimental Cancer Research, Comprehensive Cancer Center North, UK S-H, Campus Kiel, 24105, Germany (Ling Q and Kalthoff H)

Prof. Holger Kalthoff, Institute for Experimental Cancer Research, Comprehensive Cancer Center North, UK S-H, Campus Kiel, 24105, Germany (Tel: 49-431-5971938; Fax: 49-431-5971939; Email: hkalthoff@email.uni-kiel.de); Prof. Shu-Sen Zheng, Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Aff i liated Hospital, Zhejiang University School of Medicine; Key Lab of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou 310003, China (Tel/Fax: 86-571-87236567; Email: zyzss@zju.edu.cn)

? 2013, Hepatobiliary Pancreat Dis Int. All rights reserved.

10.1016/S1499-3872(13)60076-4