Serum levels of haptoglobin and SAA were determined in pretreatment samples from 75 subjects with confirmed PA, 32 patients with chronic pancreatitis, 42 patients with other benign disease (benign pancreatic neoplasm, serous cystadenoma or benign biliary stricture), and 150 healthy control subjects. Healthy control subjects were chosen such that two gender-matched and age-approximated cases were measured for each PA case. Characteristics of the study participants are presented in .
Both haptoglobin and SAA were significantly elevated in serum from PA subjects (). Haptoglobin () was significantly elevated in serum of PA subjects relative to serum from healthy controls (P < 0.0001), serum from chronic pancreatitis subjects (P = 0.0102) and serum from patients with other benign disorders (P = 0.0015). Serum haptoglobin levels were significantly higher in PA subjects with locally advanced or metastatic disease relative to PA subjects with disease localized to the pancreas (P = 0.003, data not shown) suggesting that serum haptoglobin levels increased with increasing tumor burden. Logistic regression models, adjusting for age and gender, revealed that the odds that haptoglobin predicts PA versus non-cancer controls were increased 6.2-fold for a one standard deviation increase in haptoglobin (P < 0.0001). SAA () was significantly elevated in serum of PA subjects relative to both normal control serum (P < 0.0001) and serum from chronic pancreatitis subjects (P = 0.0109) and approached significance compared to cases with other benign disorders (P = 0.0508). Serum SAA levels were not significantly different in PA subjects with localized vs. advanced disease (data not shown). A one standard deviation increase in log SAA levels was associated with a 3.6-fold increase in the odds of PA in age and gender corrected logistic regression model (P < 0.0001).
Fig. 1 Serum marker levels. Haptoglobin (A) and serum amyloid A (B) levels were determined in serum from healthy control subjects (CON, N = 150), patients with chronic pancreatitis (ChPT, N = 32), patients with benign neoplasm or other periampullary lesions (more ...)
Pretreatment serum haptoglobin and SAA levels did not significantly correlate with survival in PA subjects, although a trend towards significance was noted in which increased SAA associated with decreased survival (data not shown). Since pancreatic cancer is often associated with jaundice, we also examined the possibility that bilirubin levels could explain increased serum marker levels in PA subjects. Linear regression models revealed no apparent relationship between bilirubin and haptoglobin (P = 0.356) or SAA (P = 0.731) serum levels in pretreatment samples.
Receiver Operating Characteristic curve (ROC) analyses were used to investigate the sensitivity and specificity of haptoglobin and SAA as separate diagnostic tests for PA. depicts haptoglobin and SAA ROC curves for the discrimination of PA subjects prior to treatment (true positive cases) and non-PA control subjects, including normal control, chronic pancreatitis, benign neoplasms and cases with other benign periampullary lesions (true negative cases). ROC analysis estimates a curve, which describes the inherent tradeoff between sensitivity and specificity of a diagnostic test. Each point on the ROC curve is associated with a specific diagnostic criterion. If two ROC curves do not cross, then the diagnostic test corresponding to the higher of the two curves is uniformly better than the diagnostic test corresponding to the lower of the two curves, no matter what “cutoff” is chosen. The situation is more ambiguous if the ROC curves cross, as is the case for haptoglobin and SAA (). The area under the ROC curve (AUC) may be regarded as an average of the sensitivity over all possible specificities. The diagnostic measure with the higher AUC is therefore typically regarded as better. Thus, haptoglobin (AUC = 0.792) had greater accuracy than SAA (AUC = 0.691) over all possible cutoffs. Using classification tree analysis, single cutoffs were identified for haptoglobin (2687 μg/ml) and SAA (67.745 μg/ml) that minimized overall misclassification. These cutoffs yielded a sensitivity of 82.7 % and a specificity of 71.1% for haptoglobin and a sensitivity of 34.7% and 90.2% specificity for SAA when considering PA as true positive cases and all non-PA controls as true negative cases.
Fig. 2 Receiver operator characteristic curves for diagnosis of pancreatic adenocarcinoma versus non-cancer cases. Curves demonstrate the relative accuracy for the individual serum haptoglobin or SAA levels to discriminate between PA and control cases. Serum (more ...)
As individual markers, haptoglobin and SAA were less accurate than CA 19-9 for diagnosing PA versus non-cancer controls. However, at the commonly used cutoff of 37 U/ml, CA 19-9 still misclassified 37 of 299 cases examined (see ). We examined the possibility that combining haptoglobin and SAA with CA 19-9 in a panel diagnostic screen could improve classification. Haptoglobin, SAA and CA 19-9 serum levels from PA patients (true positive cases) and patients with chronic pancreatitis, benign neoplasms or other benign periampullary lesions (true negative cases) were used in a classification tree analysis with the stipulation that each marker be used only once. Marker levels from healthy controls were excluded from this tree analysis as these cases are much less likely to present with suspicion of PA. Thus, the resulting tree () represents an algorithm for distinguishing PA from the benign conditions. The accuracy of this algorithm for distinguishing PA from the individual classes as well as from all controls as a group is shown in . In each case the panel screen incorporating information from all three markers improved overall diagnostic accuracy over CA 19-9 alone. Except for the chronic pancreatitis group, the panel screen improved both sensitivity and specificity for discriminating PA from control groups.
Table 2 Diagnostic accuracy of the haptoglobin/SAA/CA 19-9 panel screen. Data presented as percentage. The number of accurate determinations of PA (sensitivity) or control (for specificity) per total possible cases is given in parentheses. For misclassification (more ...)
Fig. 3 Classification tree for discrimination of pancreatic adenocarcinoma (PA, N = 75) from chronic pancreatitis, benign neoplasm or other periampullary lesions (CON, N = 74). Healthy control cases were excluded from this analysis. The tree depicts an algorithm (more ...)