Prognostic Biomarkers in Pancreatic Ductal Adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers worldwide. Radical surgical resection combined with chemotherapy, the only potentially curative treatment, is possible in only a small proportion of patients. Although overall survival is poor, marked variation exists between patients of the same tumor stage. New biomarkers could be helpful in predicting prognosis. Despite considerable research on potential biomarkers, since the discovery of CA19-9, none has gained a role in clinical practice. Identification of new biomarkers to predict PDAC patient outcome more accurately and to enhance our knowledge of the molecular mechanisms behind the disease is crucial. Differential diagnosis between PDAC and chronic pancreatitis (CP) can be challenging. A pancreatic mass can prove to be benign or malignant. A clear preoperative diagnosis would be valuable for patients to avoid unnecessary and extensive surgery. The standard serum- based marker for diagnosis of PDAC, CA19-9, has diagnostic limitations because it can be normal in patients with localized disease or high in patients with benign pancreatic disease, including CP. The aim of this thesis was to explore, tissue expression of tumor biomarkers in PDAC. The prognostic significance of these biomarkers in patient survival was evaluated. In each study, we used different biomarkers: podocalyxin (PODXL), PROX1, β-catenin, UCHL5, and REG4. In the last study, we also evaluated the diagnostic significance of serum REG4 levels in patients with PDAC and in those with CP. Immunohistochemical expression of tumor markers was evaluated in 154 surgical specimens and serum REG4 level in 130 samples from PDAC patients treated between 2000 and 2011. The CP control group comprised 34 patients who underwent resection because of suspicion of malignancy. PODXL, PROX1, β-catenin, and UCHL5 were independent prognostic markers. High tissue expression of PODXL prognosticated poor survival among PDAC patients compared with low tissue expression, whereas high tissue expression of both PROX1 and β-catenin was associated with increased survival. Positive nuclear UCHL5 expression was an independent factor for favorable prognosis. REG4 failed to be an independent marker of prognosis in PDAC, but serum REG4 levels were higher in PDAC than in CP suggesting its utility in differential diagnosis. These studies provide novel knowledge of potential prognostic tumor markers in PDAC. Moreover, we identified a serum biomarker, REG4, that may be useful in differential diagnosis between PDAC and CP.Haiman duktaalinen adenokarsinooma on yksi huonoennusteisimmista syöpätaudeista. Suomessa haimasyöpä aiheuttaa kolmanneksi eniten syöpäkuolemia. Radikaali kirurgia yhdistettynä sytostaattihoitoon on ainoa mahdollinen parantava hoitomuoto, mutta vain pieni osa haimasyöpäpotilaista soveltuu tällaiseen hoitoon. Vaikka taudin yleinen ennuste on huono, saman levinneisyysasteen potilailla ennuste saattaa vaihdella paljonkin. Uudet biomarkkerit auttaisivat potilaiden ennusteen arvioinnissa. Vaikka lukuisia potentiaalisia markkereita on tutkittu vuosikausien ajan, yhtään markkeria ei ole otettu kliiniseen käyttöön CA 19-9:n löytämisen jälkeen. On välttämätöntä löytää uusia biomarkkereita, jotta haimasyöpäpotilaiden ennustetta voidaan arvioida tarkemmin ja täsmällisemmin. Tämä parantaisi myös tietämystä haimasyövän kehittymisen taustasta molekyylitasolla. Erotusdiagnostiikka haimasyövän ja kroonisen haimatulehduksen välillä voi ajoittain olla hankalaa. Haiman kuvantamistutkimuksissa nähdyt muutokset voivat olla pahan- tai hyvänlaatuisia. Tarkka diganoosi ennen mahdollista leikkausta hyödyttäisi potilaita, joita ei välttämättä tällöin tarvitsisi altistaa raskaalle kirurgiselle hoidolle. CA19-9, jota käytetään laajasti leikkausta edeltävässä diagnosoinnissa, on osin ongelmallinen, koska se voi olla normaalitasolla syövästä tai koholla hyvänlaatuisesta muutoksesta huolimatta. Tutkimuksemme tavoitteena oli tutkia kudosmarkkereiden ennusteellista arvoa haimasyövässä. Jokaisessa osatyössä tutkimme eri biomarkkereita. Tutkitut markkerit olivat podokalyksiini, PROX1, β-catenin, UCHL5 ja REG4. Viimeisessä osatyössä tutkimme myös REG4:n diagnostista arvoa haimasyövän ja kroonisen pankreatiitin välillä. Markkereiden immunohistokemiallinen ilmentyminen arviotiin 154 leikatusta haimasyöpäpotilaasta vuosina 2000- 2011. Kroonisen pankreatiitin kontrolliryhmään kuului 34 potilasta, jotka oli leikattu haiman pahanlaatuisen kasvainepäilyn takia. PODXL, PROX1, β-catenin ja UCHL5 olivat itsenäisiä ennustemarkkereita. Korkea PODXL:n kudosilmentyminen ennusti huonompaa selviytymistä syövästä verrattuna matalaan ilmentymiseen. Korkea PROX1:n ja β-cateninin kudosilmentyminen sen sijaan ennusti parempaa selviytymistä. Positiivinen UCHL5:n ilmentyminen liittyi myös parempaan ennusteeseen verrattuna negatiiviseen ilmentymiseen. REG4:n osalta taudin ennusteessa ei ollut merkittäviä eroja kudosilmentymisen mukaan, mutta REG4:n pitoisuudet olivat merkittävästi korkeammat haimasyövässä kuin kroonisessa haimatulehduksessa. Tutkimuksemme toi uutta tietoa potentiaalisista ennusteellisista biomarkkereista. Löysimme myös uuden mahdollisen biomarkkerin, REG4:n, jota voidaan mahdollisesti käyttää tulevaisuudessa apuna diagnostiikassa

TKTL1 as a Prognostic Marker in Pancreatic Ductal Adenocarcinoma and Its Correlation with FDG-PET-CT

Introduction: Glucose metabolism in cancer cells differs from noncancerous cells. The expression of transketolase-like protein 1 (TKTL1), a key enzyme in the glucose metabolism of cancer cells, predicts poor prognosis in several cancer types. We studied TKTL1 as a prognostic tool and whether TKTL1 expression correlates with 18F-FDG-PET-CT among patients with pancreatic ductal adenocarcinoma (PDAC). Methods: This retrospective study examined two PDAC patient cohorts: 168 patients operated on at Helsinki University Hospital between 2001 and 2011, and 20 patients with FDG-PET-CT results available from the Auria Biobank. We used immunohistochemistry for TKTL1 expression, combining results with clinicopathological data. Results: Five-year disease-specific survival (DSS) was slightly but not significantly better in patients with a high versus low TKTL1 expression, with DSS of 28.0 versus 17.3%, respectively (p = 0.123). TKTL1 served as a marker of a better prognosis in patients over 65 years old (p = 0.012) and among those with TNM class M1 (p = 0.018), stage IV disease (p = 0.027), or perivascular invasion (p = 0.008). Conclusions: Our study shows that TKTL1 cannot be used as a prognostic factor in PDAC with the exception of elderly patients and those with advanced disease. The correlation of TKTL1 with 18F-FDG-PET-CT requires further study in a larger patient cohort.Peer reviewe

Nuclear ubiquitin C-terminal hydrolase L5 expression associates with increased patient survival in pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma is a lethal disease with an overall 5-year survival of less than 5%. Prognosis among surgically treated patients is difficult and identification of new biomarkers is essential for accurate prediction of patient outcome. As part of one of the major cellular protein degradation systems, the proteasome plays a fundamental role in both physiological and pathophysiological conditions including cancer. The proteasome-associated deubiquitinating enzyme ubiquitin C-terminal hydrolase L5 (UCHL5)/Uch37 is a modulator of proteasome activity with cancer prognostic marker potential. Cytoplasmic and nuclear immunoexpression of UCHL5 was evaluated in 154 surgical specimens from pancreatic ductal adenocarcinoma patients treated at Helsinki University Hospital, Finland, in 2000-2011. UCHL5 expression in relation to clinicopathological parameters and the association between UCHL5 In this study, positive expression and patient survival were assessed. Positive nuclear UCHL5 expression was associated with increased patient survival (p = 0.005). A survival benefit was also detectable in these subgroups of patients: over 65 years (p <0.001), at tumor stages IIB to III (p = 0.007), or with lymph-node positivity (p = 0.006). In stages IIB to III disease, patients with positive nuclear UCHL5 expression showed a twofold increase in 5-year cancer-specific survival compared to those with negative expression. Multivariate analysis identified positive nuclear UCHL5 expression as an independent prognostic factor (p = 0.012). In conclusion, UCHL5 expression could function as a prognostic marker in pancreatic ductal adenocarcinoma, particularly at disease stages IIB to III. As UCHL5 is one of the few markers predicting increased survival, our results may be of clinical relevance.Peer reviewe

Additive clinical impact of epidermal growth factor receptor and podocalyxin-like protein expression in pancreatic and periampullary adenocarcinomas

The outcome of periampullary adenocarcinomas remains poor with few treatment options. Podocalyxin-like protein (PODXL) is an anti-adhesive protein, the high expression of which has been shown to confer a poor prognosis in numerous malignancies. A correlation and adverse prognostic synergy between PODXL and the epidermal growth factor receptor (EGFR) has been observed in colorectal cancer. Here, we investigated whether this also applies to periampullary adenocarcinomas. We analyzed the immunohistochemical expression of PODXL and EGFR in tissue microarrays with tumors from two patient cohorts; (Cohort 1, n=175) and (Cohort 2, n=189). The effect of TGF-beta -induced expression and siRNA-mediated knockdown of PODXL and EGFR, were investigated in pancreatic cancer cells (PANC-1) in vitro. We found a correlation between PODXL and EGFR in these cancers, and a synergistic adverse effect on survival. Furthermore, silencing PODXL in pancreatic cancer cells resulted in the down-regulation of EGFR, but not vice versa. Consequently, these findings suggest a functional link between PODXL and EGFR, and the potential combined utility as biomarkers possibly improving patient stratification. Further studies examining the mechanistic basis underlying these observations may open new avenues of targeted treatment options for subsets of patients affected by these particularly aggressive cancers.Peer reviewe

Prognostic and diagnostic value of REG4 serum and tissue expression in pancreatic ductal adenocarcinoma

Expression of regenerating islet-derived protein 4 (REG4), a secretory protein involved in cell differentiation and proliferation, is upregulated in inflammatory bowel diseases and in many gastrointestinal malignancies. The prognostic significance of its expression in pancreatic ductal adenocarcinoma is unknown. Our aim was to investigate tumor tissue and serum REG4 expression in pancreatic ductal adenocarcinoma patients. We also evaluated as a control the diagnostic value of serum REG4 level in patients with chronic pancreatitis. Immunohistochemical expression of REG4 was evaluated in 154 surgical specimens and serum REG4 level in 130 samples from pancreatic ductal adenocarcinoma patients treated at Helsinki University Hospital, Finland, in 2000–2011. REG4 tissue and serum expression was assessed in relation to clinicopathological parameters and patient survival. A chronic pancreatitis control group comprised 34 patients who underwent pancreatic resection because of suspicion of malignancy. Significant survival differences were detectable in subgroups: in tumor stages IA–IIA, high serum REG4 level predicted worse survival (p=0.046). In patients with grade I tumor, positive tissue REG4 expression predicted better survival (p=0.006). In multivariate analysis, neither tissue nor serum REG4 expression was independent prognostic factors. Serum REG4 levels were higher in pancreatic ductal adenocarcinoma than in chronic pancreatitis (p=0.002), with diagnostic sensitivity of 45% and specificity of 91%. In logistic regression analysis, a multivariate model with REG4, CA19-9, and age provided sensitivity of 82% and specificity of 79%. REG4 tissue expression is a prognostic marker in subgroups of pancreatic ductal adenocarcinoma patients. Serum REG4 level might be useful in differential diagnosis between pancreatic ductal adenocarcinoma and chronic pancreatitis.</p

PROX1 and beta-catenin are prognostic markers in pancreatic ductal adenocarcinoma

Background: The Wnt/beta-catenin pathway has a key role in regulating cellular processes and its aberrant signaling can lead to cancer development. The role of beta-catenin expression in pancreatic ductal adenocarcinoma is somewhat controversial. Transcription factor PROX1 is a target of Wnt/beta-catenin signaling and it is involved in carcinogenesis through alterations in its expression. The actions can be either oncogenic or tumor suppressive depending on the tissue. The aim of this study was to investigate PROX1 and beta-catenin expression in pancreatic ductal adenocarcinoma (PDAC). Methods: Expression of PROX1 and beta-catenin were evaluated in 156 patients by immunohistochemistry of tissue microarrays. Associations between tumor marker expression and clinicopathological parameters were assessed by the Fischer's exact-test or the linear-by-linear association test. The Kaplan-Meier method and log-rank test were used for survival analysis. Uni- and multivariate survival analyses were carried out by the Cox regression proportional hazard model. Results: High PROX1 expression was seen in 74 (48 %) tumors, and high beta-catenin expression in 100 (65 %). High beta-catenin expression was associated with lower tumor grade (p = 0.025). High PROX1 and beta-catenin expression associated significantly with lower risk of death from PDAC in multivariate analysis (HR = 0.63; 95 % CI 0.42-0.95, p = 0.026; and HR = 0.54; 95 % CI 0.35-0.82, p = 0.004; respectively). The combined high expression of PROX1 and beta-catenin also predicted lower risk of death from PDAC (HR = 0.46; 95 % CI 0.28-0.76, p = 0.002). Conclusion: In conclusion, high PROX1 and beta-catenin expression were independent factors for better prognosis in pancreatic ductal adenocarcinoma.Peer reviewe

Podocalyxin Is a Marker of Poor Prognosis in Pancreatic Ductal Adenocarcinoma

Aim of the Study Podocalyxin-like 1 is a transmembrane glyco-protein whose overexpression associates in many cancers with poor prognosis and unfavorable clinicopathological characteristics. Until now, its prognostic value has never been studied in pancreatic ductal adenocarcinoma (PDAC). The aim of this study was to investigate podocalyxin expression in PDAC by a novel monoclonal antibody and a commercially available polyclonal antibody. Patients and Materials With tissue microarrays and immuno-histochemistry, podocalyxin expression evaluation involved 168 PDAC patients. The associa-tions of the podocalyxin tumor expression with clinicopathological variables were explored by Fisher's exact test and the linear-by-linear test. Survival analyses were by Kaplan-Meier anal-ysis and the Cox proportional hazard model. Results The polyclonal antibody revealed membranous podocalyxin expression in 73 (44.0%) specimens and the monoclonal antibody was highly expressed in 36 (21.8%) cases. Membranous expression by the polyclonal antibody was associated with T classification (p=0.045) and perineural invasion (p=0.005), and high expression by the mono-clonal antibody with poor differentiation (p=0.033). High podocalyxin expression associated significantly with higher risk of death from PDAC by both the polyclonal antibody (hazard ratio (HR) = 1.62; 95% confidence interval (CI) 1.12-2.33; p=0.01) and the monoclonal antibody (HR = 2.10, 95% CI 1.38-3.20; p=/<20%), and perivascular invasion (respectively as HR = 2.03; 95% CI 1.32-3.13, p=0.001; and as HR = 2.36; 95% CI 1.47-3.80, p Conclusion We found podocalyxin to be an independent factor for poor prognosis in PDAC. To our knowledge, this is the first such report of its prognostic value.Peer reviewe

Membranous PODXL expression by polyclonal antibody HPA 2110 is a marker of poor prognosis in pancreatic ductal adenocarcinoma.

<p>Cancer-specific survival analysis according to the Kaplan-Meier method for membranous PODXL expression by the polyclonal antibody HPA 2110 in pancreatic ductal adenocarcinoma. Log-rank test was used here.</p

Cox uni- and multivariate analysis of relative risk of death from pancreatic ductal adenocarcinoma by PODXL expression.

<p>Abbrevations CI = Confidence interval, HR = Hazard ratio. Multivariate analysis included adjustment for age, gender, stage (IA-IIA, IIB and III), lymph node ratio (≥/< 20%), and perivascular invasion.</p><p>Cox uni- and multivariate analysis of relative risk of death from pancreatic ductal adenocarcinoma by PODXL expression.</p