Table 1.
Baseline characteristics of the NSCLC study population.
Fig 1.
Expression of CD13 in NSCLC tissues.
Examples for negative immunostaining are shown for squamous cell carcinoma (Fig. 1A) and for adenocarcinoma (Fig. 1B). Positive staining results are given for squamous cell carcinomas (stromal expression of CD13) (Fig. 1C and Fig. 1D) and for adenocarcinomas (tumor cell expression of CD13) (Fig. 1E and Fig. 1F). Co-staining of CD31 (green) and of CD13 (red) demonstrates CD13 expression in endothelial cells and perivascular stroma cells in a case of adenocarcinoma (Fig. 1G) and squamous cell carcinoma (Fig. 1H).
Table 2.
Correlations of clinicopathological variables with CD13 in NSCLC patients.
Table 3.
Univariate log rank test results for the association of CD13 with overall survival for the investigated study collective of n = 270 tissue microarray-embedded NSCLC patients (immunohistochemical analysis).
Fig 2.
Prognostic impact of CD13 expression in non-small cell lung cancer (NSCLC).
Univariate prognostic models (i.e. microarray-based mRNA expression) are shown for CD13 of all patients (Fig. 2A) and for stage I tumor patients (Fig. 2B) according to “The Kaplan-Meier plotter” database (www.kmplot.com [25]). With regard to the immuno-histochemical protein expression of CD13 in endothelial cells and vessel-associated stroma cells of the tumors prognostic analyses were performed for our complete NSCLC study collective (Fig. 2C), for squamous cell carinoma patients (Fig. 2D), for stage III NSCLC tumor patients (Fig. 2E), and for patients with pN2 lymph node status (Fig. 2F).
Table 4.
Univariate log rank test results for the association of CD13 with overall survival for the study collective of n = 1926 lung cancer patients as provided by “The Kaplan-Meier plotter” (KM plotter) database (transcriptomic data).
Table 5.
Overall survival: Exploratory prognostic factors in a Cox proportional hazards model.
Included variables: CD13 expression in stromal cells (negative expression (ref.) vs. positive expression), CD13 expression in NSCLC tumor cells (negative expression (ref.) vs. positive expression), tumor stage (stage I (ref.) vs. stage II vs. stage III), lymph node status (pN0 (ref.) vs. pN1 vs. pN2 vs. pN2), and tumor histology (squamous cell carcinoma vs. adenocarcinoma vs. large cell carcinoma).
Fig 3.
In vivo therapeutic activity of systemic tTF-NGR against CD13+ A549 tumor xenografts.
To investigate CD13 expression flow cytometry was performed with a monoclonal PE-labeled anti-CD13 antibody. CD13 expression was found in 47% of the A549 lung cancer cells (green, control; purple, CD13) (Fig. 3A). Following treatment with tTF-NGR (1 mg tTF-NGR/kg x4 (arrows); i.v.; n = 4 CD-1 nude mice) tumor growth of subcutaneous A549 xenotransplants was reduced as compared to the saline control group (n = 6) CD-1 nude mice (Fig. 3B). The CD13 expression in subcutaneous A549 xenotransplant is demonstrated by immunofluorescence; since the antibodies used for CD13 and CD31 staining are species-specific for human CD13 and CD31, vascular and perivascular staining were not assayed in the xenografts (Fig. 3C).