The clinical features of all 60 PACs were described in Table 1. The mean age was 64.6 years, and 38 cases were male. The lesions were located in the pancreatic head in 30 cases and body/tail in 30 cases. The mean size was 36.0 mm (range, 21.0–60.5 mm). Clinically, 48 (80.0%) cases received gemcitabine-based chemotherapy, 2 cases received mFOLFIRINOX, and the remaining 10 cases did not receive any chemotherapy.
Table 1
Clinical features of included PAC.
Age (years, mean ± SD) | 64.6 ± 9.1 |
Gender (n, %) | |
Female | 22 (36.7%) |
Male | 38 (63.3%) |
BMI (mean ± SD) | 22.3 ± 2.8 |
Location (n, %) | |
Head | 30 (50.0%) |
Body/tail | 30 (50.0%) |
Lesion size (mm, mean ± SD) | 36.0 ± 8.7 |
Dilation of Bile duct (n, %) | |
Negative | 48 (80.0%) |
Positive | 12 (20.0%) |
Dilation of Pancreatic duct (n, %) | |
Negative | 24 (40.0%) |
Positive | 36 (60.0%) |
Serum Liver test | |
Albumin (g/L, mean ± SD) | 42.3 ± 4.3 |
Blood urea nitrogen (µmol/L, mean ± SD) | 4.8 ± 1.3 |
Lactic dehydrogenase (U/L, mean ± SD) | 172.3 ± 58.2 |
Fasting blood glucose (mmol/L, mean ± SD) | 6.6 ± 2.7 |
Uric acid (µmol/L, mean ± SD) | 267.8 ± 73.3 |
Serum CA199 (n, %) | |
Normal | 9 (15.0%) |
Elevation | 51 (85.0%) |
Therapy (n, %) | |
Gemcitabine-based chemo | 48 (80.0%) |
mFOLFIRINOX | 2 (3.3%) |
Other | 10 (16.7%) |
In LC-MS analysis, 2,550 features and 2,307 features were obtained at the electrospray ionization negative and positive (ESI− and ESI+) ionic modes. A total of 587 metabolites were identified and annotated based on compound molecular weight and peak intensity. For metabolites with multiple items, the average level was calculated, and 402 unique metabolites were finally included in the analysis.
Collectively, all patients with PAC were followed up for at least 1 year. The median follow-up time was 352 days, ranging from 0 day to 762 days. Forty-five patients died at the time of the last follow-up (January 15, 2021), which included all the ten cases without any chemotherapy. The median survival time was 390 days, ranging from 324 days to 492 days. Initially, 16 metabolites were identified by univariate analysis (Fig. 1). Further multivariate analysis indicated that cholesterol glucuronide (HR = 1.181, 95%CI = 1.092–1.276, p < 0.001) and taurocholic acid 3-sulfate (HR = 1.132, 95%CI = 1.058–1.210, p < 0.001) were the independent predictors for overall survival (OS). As shown in the human metabolome database, both cholesterol glucuronide and taurocholic acid 3-sulfate were associated with bile acid metabolism. In PAC, the head tumor always induced the obstructive dilation of the bile duct and cholestasis instead of the body/tail tumor. As shown in Supplemental Fig. 1, the level of taurocholic acid 3-sulfate was slightly higher in cancers with head location and bile duct dilation, than body/tail location and bile duct non-dilation. As for cholesterol glucuronide, no any significant difference in such two conditions.
In accordance with the coefficient of prognostic metabolites, the risk score was calculated and stratified into the high-risk (n = 15) and low-risk (n = 45) groups. The C-index of the risk score was 0.64 (95% CI: 0.70–0.59). The areas under the curve at 1 and 2 years were 0.685 and 0.588, respectively (Fig. 2A). As shown in Fig. 2B, the levels of the two metabolites were higher in the high-risk group than in the low-risk group (p < 0.001 for cholesterol glucuronide, p < 0.05 for taurocholic acid 3-sulfate). The Kaplan–Meier plot indicated that the high-risk group had significantly worse prognosis than the low-risk group (p < 0.001), with median survival times of 195 and 480 days, respectively (Fig. 2C).
Further, clinical parameters were incorporated into the Cox model with risk score to construct the predictive model. With univariate and multivariate analyses, therapy (HR = 0.270, 95% CI = 0.097–0.747, p = 0.012; Fig. 3) and risk score (HR = 1.983, 95% CI = 1.362–2.887, p < 0.001; Fig. 3) were found to be the independent predictors for prognosis. The predictive model was depicted in the nomogram (Fig. 4), with a C-index of 0.67 (95% CI: 0.73–0.62). A credible predictive model based on the metabolites was finally constructed for PAC prognosis.
Subsequently, we attempted to investigate the metabolic profile in different risk groups. As shown in the volcano plot (Fig. 5A), 29 metabolites showed higher levels in the high-risk group than in the low-risk group, and the difference was statistically significant (p < 0.05). The detailed level of significant metabolites is shown in the heatmap (Fig. 5B).
Finally, we performed WGCNA to cluster the metabolites and identified three distinct modules, namely, turquoise, blue, and grey (Figs. 6A and 6B). All the three showed no significant association with clinical parameters (Supplemental Fig. 2). A total of 243 metabolites were clustered into the turquoise module and majorly enriched in the glucose-alanine cycle. A total of 150 metabolites were clustered into the blue module and majorly enriched in α-linolenic acid metabolism. Only nine metabolites were clustered into the grey module, and enriched in no metabolic pathway. Details are shown in Figs. 6C and 6D.