Biomarkers for pancreatic cancer: Recent achievements in proteomics and genomics through classical and multivariate statistical methods

Table 2 Studies based on serum and tissue biomarkers through non-omics techniques

Ref.	Type of marker	Markers	Sample	Study group	Analytical methods	Statistical methods	Performance
41	P	CA 19-9	S	Pretreatment CA 19-9: 115 patients from 5 German centers; 73% treated within prospective clinical trials. Median TTP: 4.4 mo; median OS: 9.4 mo. CA 19-9 kinetics during chemotherapy: 69 patients (TTP) and 84 patients (OS)	Elecsys assay	Cox proportional hazards regression; for CA 19-9 kinetics, CA 19-9 was treated as a time-varying covariate	Univariate analysis: log (CA 19-9) associated with TTP (HR = 1.24; P < 0.001) and OS (HR = 1.16; P = 0.002). Multivariate analysis: results confirmed. Log(CA 19-9) kinetics during chemotherapy: significant predictor for TTP in univariate analyses (HR = 1.48; P < 0.001) and multivariate (HR = 1.45; P < 0.001) and for OS (univariate: HR = 1.34; P < 0.001; multivariate: HR =1.38; P < 0.001)
42	P	CA 19-9, CEA, CRP, LDH and bilirubin		291 patients; 253 patients (87 %) received treatment within prospective clinical trials. Median TTP: 5.1 mo. Median OS 9.0 mo	Elecsys assay	Kaplan Meier method and Cox proportional hazards regression	Univariate analysis: pre-treatment CA 19-9 (HR = 1.55), LDH (HR = 2.04) and CEA (HR = 1.89) significantly associated with TTP. Baseline CA 19-9 (HR = 1.46), LDH (HR = 2.07), CRP (HR = 1.69) and bilirubin (HR = 1.62) significant prognostic factors for OS. Multivariate analyses: pre-treatment log (CA 19-9) for TTP and log (bilirubin) and log (CRP) for OS had an independent prognostic value
44	P	IGFs	S and P	80 patients received treatment (40 Ganitumab; 40 placebo)	Immunoassays	Kaplan Meier method and Cox proportional hazards regression	Ganitumab associated with improved OS vs placebo (HR = 0.49; 95%CI: 0.28-0.87)
45	P	TROP2	T	197 patients; subgroup of 134 patients treated surgically	Immunohistochemistry	Kaplan Meier method and Cox proportional hazards regression	TROP2 overexpression observed in 109 (55%) patients and associated with decreased OS (P < 0.01). Univariate Analysis: TROP2 overexpression correlates with lymph node metastasis (P < 0.04) and tumor grade (P < 0.01). In the subgroup of patients treated surgically, TROP2 overexpression correlated with poor progression-free survival (P < 0.01). Multivariate analyses: TROP2 is an independent prognosticator
46	P	JAM-A	T	186 patients; subgroup of 83 patients treated surgically	Immunohistochemistry	Kaplan Meier method and Cox proportional hazards regression	Low expression of JAM-A observed in 79 (42 %) patients and associated with poor OS (P < 0.01). Univariate analysis: low expression of JAM-A correlates with positive lymph node status (P = 0.02), the presence of distant metastasis (P = 0.05), and tumor grade (P = 0.04). In the subgroup of patients with surgically resected PC, low expression of JAM-A correlated with decreased progression-free survival (P < 0.01). Multivariate analysis: JAM-A was an independent predictor of poor outcome
47	P	TBX4	T	77 stage II PDAC tumors	Immunohistochemistry	Kaplan Meier method and Cox proportional hazards regression	48 cases (62.3%) expressed TBX4 at a high level. No significant correlation between TBX4 expression and other clinicopathological parameters, except tumor grade and liver metastasis recurrence. Survival of patients with TBX4-high expression significantly longer than those with TBX4-low expression (P = 0.010). Multivariate analysis: low TBX4 expression independent prognostic factor for OS. TBX4 promoter methylation status frequently observed in PDAC and normal adjacent pancreas
48	P	HSP27	T	86 patients	Tissue microarray (TMA) analysis	Kaplan Meier method and Cox proportional hazards regression	HSP27 expression found in 49% of tumor samples. Univariate analyses: significant correlation between HSP27 expression and survival. Multivariate Cox-regression: HSP27 expression emerged as an independent prognostic factor. HSP27 expression also correlated inversely with nuclear p53 accumulation
49	P	dCK	T	45 patients with resected PDAC received adjuvant gemcitabine based-therapy in multicenter phase 2 studies	Immunohistochemistry	Kaplan Meier method and Cox proportional hazards regression	Median follow-up: 19.95 mo (95%CI: 3.3-107.4 mo). Lymph node (LN) ratio and dCK protein expression significant predictors of DFS and OS in univariate analysis. Multivariate analysis: dCK protein expression the only independent prognostic variable (DFS: HR = 3.48, 95%CI: 1.66-7.31, P < 0.001, OS: HR = 3.2, 95%CI: 1.44-7.13, P < 0.004)
50	P	Notch3 and Hey-1	T	42 patients who underwent resection and 50 patients diagnosed with unresectable PDAC	Immunohistochemistry	Mann-Whitney U test, Wilcoxon test, Cox regression analysis, Kaplan-Meier analysis	All 3 Notch family members significantly elevated in tumor tissue. Significantly higher nuclear expression of Notch1, -3 and -4, HES-1, and HEY-1 (all P < 0.001) in locally advanced and metastatic tumors compared to resectable cancers. In survival analyses, nuclear Notch3 and HEY-1 expression significantly associated with reduced OS and DFS following tumor resection with curative intent
51	D and P	21 biomarkers	P	clinically defined cohort of 52 locally advanced (Stage II/III) PDAC cases and 43 age-matched controls	Proximity ligation assay	Combination of the PAM algorithm and logistic regression modeling. Biomarkers that were significantly prognostic for survival were determined using univariate and multivariate Cox survival models	CA19-9, OPN and CHI3L1 were found to have superior sensitivity for pancreatic cancer vs CA19-9 alone (93% vs 80%). CEA and CA125 have prognostic significance for survival (P < 0.003)
52	D	83 circulating proteins	S	333 PDAC patients; 144 controls (benign pancreatic conditions); 227 healthy controls. Samples from each group split randomly into training and blinded validation sets. Panels evaluated in validation set and in patients diagnosed with colon (33), lung (62) and breast (108) cancers	bead-based xMAP immunoassays	A Metropolis algorithm with Monte Carlo simulation (MMC) was used to identify discriminatory biomarker panels in the training set	Training set (160 PDAC, 74 Benign, 107 Healthy): panel of CA19–9, ICAM-1, and OPG discriminated PDAC from Healthy controls (SN/SP 88/90%), panel of CA 19–9, CEA, and TIMP-1 discriminated PDAC patients from Benign subjects (SN/SP = 76%/90%). Independent validation set (173 PDAC, 70 Benign, 120 Healthy): panel of CA 19–9, ICAM-1 and OPG demonstrated SN/SP of 78%/94%; panel of CA19–9, CEA, and TIMP-1 demonstrated SN/SP of 71%/89%. The CA19–9, ICAM-1, OPG panel is selective for PDAC and does not recognize breast (SP = 100%), lung (SP = 97%), or colon (SP = 97%) cancer
53	D and P	YKL-40, IL-6, and CA 19.9	P	559 patients with PC from prospective biomarker studies from Denmark (n = 448) and Germany (n = 111)	ELISA and chemiluminescent immunometric assay	Kaplan Meier method and Cox proportional hazards regression	Odds ratios (ORs) for prediction of PC significant for all biomarkers, with CA 19.9 having the highest AUC (CA 19.9: OR = 2.28, 95%CI: 1.97-2.68, P = 0.0001, AUC = 0.94; YKL-40: OR = 4.50, 3.99-5.08, P = 0.0001, AUC = 0.87; IL-6: OR = 3.68, 3.08-4.44, P = 0.0001, AUC = 0.87). Multivariate Cox analysis: high preoperative IL-6 and CA 19.9 independently associated with short OS (CA 19.9: HR = 2.51, 1.22–5.15, P = 0.013; IL-6: HR = 2.03, 1.11-3.70, P = 0.021). Multivariate Cox analysis of non-operable patients: high pre-treatment levels of each biomarker independently associated with short OS (YKL-40: HR = 1.30, 1.03-1.64, P = 0.029; IL-6: HR = 1.71, 1.33-2.20, P = 0.0001; CA 19.9: HR = 1.54, 1.06-2.24, P = 0.022). Patients with preoperative elevation of IL-6 and CA 19.9 had shorter OS (P = 0.005) compared to patients with normal levels (45% vs 92% alive after 12 mo)

Type of marker: P: Prognostic/predictive; D: Diagnostic; Sample: S: Serum; P: Plasma; T: Tissue; TTP: Time to progression.