Analysis of circulating DNA and protein biomarkers to predict the clinical activity of regorafenib and assess prognosis in patients with metastatic colorectal cancer: a retrospective, exploratory analysis of the CORRECT trial.

BACKGROUND: Tumour mutational status is an important determinant of the response of metastatic colorectal cancer to targeted treatments. However, the genotype of the tissue obtained at the time of diagnosis might not accurately represent tumour genotype after multiple lines of treatment. This retrospective exploratory analysis investigated the clinical activity of regorafenib in biomarker subgroups of the CORRECT study population defined by tumour mutational status or plasma protein levels. METHODS: We used BEAMing technology to identify KRAS, PIK3CA, and BRAF mutations in DNA obtained from the plasma of 503 patients with metastatic colorectal cancer who enrolled in the CORRECT trial. We quantified total human genomic DNA isolated from plasma samples for 503 patients using a modified version of human long interspersed nuclear element-1 (LINE-1) quantitive real-time PCR. We also measured the concentration of 15 proteins of interest-angiopoietin 2, interleukin 6, interleukin 8, placental growth factor, soluble TIE-1, soluble VEGFR1, VEGF-A, VEGF-C, VEGF-D, VEGF-A isoform 121, bone morphogenetic protein 7, macrophage colony-stimulating factor, stromal cell-derived factor-1, tissue inhibitor of metalloproteinase 2, and von Willebrand factor-in plasma samples from 611 patients. We did correlative analyses of overall survival and progression-free survival in patient subgroups based on mutational status, circulating DNA concentration, and protein concentrations. The CORRECT trial was registered with ClinicalTrials.gov, number NCT01103323. FINDINGS: Tumour-associated mutations were readily detected with BEAMing of plasma DNA, with KRAS mutations identified in 349 (69%) of 503 patients, PIK3CA mutations in 84 (17%) of 503 patients, and BRAF mutations in 17 (3%) of 502 patients. We did not do correlative analysis based on BRAF genotype because of the low mutational frequency detected for this gene. Some of the most prevalent individual hot-spot mutations we identified included: KRAS (KRAS G12D, 116 [28%] of 413 mutations; G12V, 72 [17%]; and G13D, 67 [16%]) and PIK3CA (PIK3CA E542K, 27 [30%] of 89 mutations; E545K, 37 [42%]; and H1047R, 12 [14%]). 41 (48%) of 86 patients who had received anti-EGFR therapy and whose archival tumour tissue DNA was KRAS wild-type in BEAMing analysis were identified as having KRAS mutations in BEAMing analysis of fresh plasma DNA. Correlative analyses suggest a clinical benefit favouring regorafenib across patient subgroups defined by KRAS and PIK3CA mutational status (progression-free survival with regorafenib vs placebo: hazard ratio [HR] 0·52, 95% CI 0·35-0·76 for KRAS wild-type; HR 0·51, 95% CI 0·40-0·65 for KRAS mutant [KRAS wild type vs mutant, pinteraction=0·74]; HR 0·50, 95% CI 0·40-0·63 for PIK3CA wild-type; HR 0·54, 95% CI 0·32-0·89 for PIK3CA mutant [PIK3CA wild-type vs mutant, pinteraction=0·85]) or circulating DNA concentration (progression-free survival with regorafenib vs placebo: HR 0·53, 95% CI 0·40-0·71, for low circulating DNA concentrations; HR 0·52, 95% CI 0·40-0·70, for high circulating DNA concentrations; low vs high circulating DNA, pinteraction=0·601). With the exception of von Willebrand factor, assessed with the median cutoff method, plasma protein concentrations were also not associated with regorafenib activity in terms of progression-free survival. In univariable analyses, the only plasma protein that was associated with overall survival was TIE-1, high concentrations of which were associated with longer overall survival compared with low TIE-1 concentrations. This association was not significant in multivariable analyses. INTERPRETATION: BEAMing of circulating DNA could be a viable approach for non-invasive analysis of tumour genotype in real time and for the identification of potentially clinically relevant mutations that are not detected in archival tissue. Additionally, the results show that regorafenib seems to be consistently associated with a clinical benefit in a range of patient subgroups based on mutational status and protein biomarker concentrations. FUNDING: Bayer HealthCare Pharmaceuticals.

Disturbed balance between serum levels of receptor tyrosine kinases Tie-1, Tie-2 and angiopoietins in systemic sclerosis.

OBJECTIVE: The aim of this study was to determine the characteristic factors for vascular development and maintenance levels as well as correlation between Tie-1 receptors, Tie-2 receptors and the corresponding ligands--angiopoietins--in systemic sclerosis (SSc) patients. MATERIALS AND METHODS: Serum levels of Tie-1, Tie-2, Ang-1 and Ang-2 were measured in 25 SSc patients and healthy controls. RESULTS: There was a statistically significant difference in serum Tie-1 (p = 0.009) and Ang-2 (p = 0.001) levels in SSc patients compared with healthy controls. Significant correlations between Tie-1 and Tie-2 (ρ = 0.70, p = 0.0001) and between Tie-1 and Ang-2 (ρ = -0.92, p = 0.002) were found in the SSc group. Serum levels of Tie-2 were positively associated with esophagus changes (U = 2.03, p = 0.041) and Ang-1 was negatively correlated with duration of Raynaud's phenomenon (ρ = -0.75, p = 0.00008). CONCLUSION: The increase in serum concentration of Tie-1 and Ang-2 in patients with SSc may confirm a molecular imbalance between receptor tyrosine kinases Tie and their ligands.

Serum Tie-1 is a Valuable Marker for Predicting the Progression and Prognosis of Cervical Cancer.

Objective: To investigate whether serum Tie-1 (sTie-1) is a valuable marker for predicting progression and prognosis of cervical cancer. Methods: Enzyme-linked immunosorbent assay (ELISA) was used to detect serum sTie-1 concentrations in 75 cervical cancer patients, 40 cervical intraepithelial neoplasia (CIN) patients, and 55 healthy controls without cervical lesions, and sTie-1 levels were compared between the groups. Receiver operating characteristic curves was used to evaluate the diagnostic value of sTie-1. The relationship between sTie-1 concentrations in patients with cervical cancer and clinicopathological features and prognosis were analyzed, and the risk factors for postoperative recurrence were determined using univariate and multivariable Cox proportional hazards regression. Results: We found that sTie-1 concentrations gradually increased according to lesion severity (i.e., cancer vs. CIN; p < 0.05) and were significantly elevated in adenocarcinoma compared with healthy controls. sTie-1 levels strongly distinguished between cervical cancer patients and the healthy controls (area under the curve = 0.846; cut-off value = 1,882.64 pg/ml; sensitivity = 74.6%; specificity = 96.4%). Moreover, sTie-1 levels in cervical cancer patients were significantly associated with tumor size, advanced tumor stage, lymph node metastasis, and reduced 4-years progression-free survival. Cervical cancer patients with high sTie-1 concentrations had a 3.123-fold [95% confidence interval (CI): 1.087-8.971, p = 0.034] higher risk for tumor recurrence. Conclusions: Elevated sTie-1 levels in patients with cervical carcinoma were associated with tumor progression and poor prognosis, indicating that sTie-1 may be a valuable marker for predicting progression and prognosis of cervical cancer.

Clinical significance of serum soluble Tie1 levels in patients with systemic sclerosis.

Tie1 is an endothelial cell-specific tyrosine kinase receptor, which maintains vascular integrity and regulates angiogenesis via modulating angiopoietin/Tie2 signaling. Since the altered angiogenesis is closely related to the developmental process of systemic sclerosis (SSc), we herein investigated the clinical significance of serum soluble Tie1 (sTie1) levels and the expression levels of Tie1 in dermal microvascular endothelial cells (DMECs) in patients with SSc. Although serum sTie1 levels were comparable among total SSc, diffuse cutaneous SSc (dcSSc), limited cutaneous SSc (lcSSc), and healthy controls, SSc patients with decreased serum sTie1 levels had significantly shorter disease duration than those with serum sTie1 levels not decreased. In SSc patients with disease duration of >6 years, the prevalence of clinical symptoms associated with proliferative vasculopathy, such as digital ulcers, scleroderma renal crisis, and elevated right ventricular systolic pressure, was significantly higher in patients with decreased serum sTie1 levels than in those with serum sTie1 levels not decreased. In immunohistochemistry, Tie1 expression was reduced in DMECs of SSc patients with disease duration of <3 years compared with those of healthy controls. Collectively, in SSc patients with short disease duration, decreased serum sTie1 levels may reflect the down-regulation of Tie1 in DMECs. The decrease in serum sTie1 levels may serve as a marker of proliferative vasculopathy in SSc with disease duration of >6 years.