Verapamil inhibits tumor progression of chemotherapy-resistant pancreatic cancer side population cells.

Tumor side population (SP) cells display stem-like properties that can be modulated by treatment with the calcium channel blocker verapamil. Verapamil can enhance the cytotoxic effects of chemotherapeutic drugs and multidrug resistance by targeting the transport function of the P-glycoprotein (P-gp). This study focused on the therapeutic potential of verapamil on stem-like SP tumor cells, and further investigated its chemosensitizing effects using L3.6pl and AsPC-1 pancreatic carcinoma models. As compared to parental L3.6pl cells (0.9±0.22%), L3.6pl gemcitabine-resistant cells (L3.6plGres) showed a significantly higher percentage of SP cells (5.38±0.99%) as detected by Hoechst 33342/FACS assays. The L3.6plGres SP cells showed stable gemcitabine resistance, enhanced colony formation ability and increased tumorigenicity. Verapamil effectively inhibited L3.6plGres and AsPC-1 SP cell proliferation in vitro. A pro-apoptotic effect of verapamil was observed in L3.6pl cells, but not in L3.6plGres cells, which was linked to their differential expression of P-gp and equilibrative nucleoside transporter-1 (ENT-1). In an orthotopic pancreatic cancer mouse model, both low and high dose verapamil was shown to substantially reduce L3.6plGres-SP cell tumor growth and metastasis, enhance tumor apoptosis, and reduce microvascular density.

In vivo monitoring of sorafenib therapy effects on experimental prostate carcinomas using dynamic contrast-enhanced MRI and macromolecular contrast media.

PURPOSE: To investigate dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) with macromolecular contrast media (MMCM) to monitor the effects of the multikinase inhibitor sorafenib on subcutaneous prostate carcinomas in rats with immunohistochemical validation. MATERIALS AND METHODS: Copenhagen rats, implanted with prostate carcinoma allografts, were randomized to the treatment group (n = 8) or the control group (n = 8). DCE-MRI with albumin-(Gd-DTPA)35 was performed at baseline and after 1 week using a clinical 3-Tesla system. The treatment group received sorafenib, 10 mg/kg body weight daily. Kinetic analysis yielded quantitative parameters of tumor endothelial permeability-surface area product (PS; ml/100 ml/min) and fractional blood volume (Vb, %). Tumors were harvested on day 7 for immunohistochemical analysis. RESULTS: In sorafenib-treated tumors, PS (0.62 ± 0.20 vs 0.08 ± 0.09 ml/100 ml/min; P < 0.01) and Vb (5.1 ± 1.0 vs 0.56 ± 0.48%; P < 0.01) decreased significantly from day 0 to day 7. PS showed a highly significant inverse correlation with tumor cell apoptosis (TUNEL; r = -0.85, P < 0.001). Good, significant correlations of PS were also observed with tumor cell proliferation (Ki-67; r = 0.67, P < 0.01) and tumor vascularity (RECA-1; r = 0.72, P < 0.01). MRI-assayed fractional blood volume Vb showed a highly significant correlation with tumor vascularity (RECA-1; r = 0.87, P < 0.001) and tumor cell proliferation (Ki-67; r = 0.82, P < 0.01). CONCLUSION: Results of DCE-MRI with MMCM demonstrated good, significant correlations with the immunohistochemically assessed antiangiogenic, antiproliferative, and proapoptotic effects of a 1-week, daily treatment course of sorafenib on experimental prostate carcinoma allografts.

Dynamic contrast-enhanced computed tomography imaging biomarkers correlated with immunohistochemistry for monitoring the effects of sorafenib on experimental prostate carcinomas.

OBJECTIVES: To investigate dynamic contrast-enhanced computed tomography (DCE-CT) for monitoring the effects of sorafenib on experimental prostate carcinomas in rats by quantitative assessments of tumor microcirculation parameters with immunohistochemical validation. MATERIAL AND METHODS: Prostate carcinoma allografts (MLLB-2) implanted subcutaneously in male Copenhagen rats (n=16) were imaged at baseline and after a 1-week treatment course of sorafenib using DCE-CT with iopromide (Ultravist 370, Bayer Pharma, Berlin, Germany) on a dual-source 128-slice CT (Somatom Definition FLASH, Siemens Healthcare, Forchheim, Germany). Scan parameters were as follows: detector width, 38.4 mm; contrast agent volume, 2 mL/kg bodyweight; injection rate, 0.5 mL/s; scan duration, 90 seconds; and temporal resolution, 0.5 seconds. The treatment group (n=8) received daily applications of sorafenib (10 mg/kg bodyweight) via gavage. Quantitative parameters of tumor microcirculation (plasma flow, mL/100 mL/min), endothelial permeability-surface area product (PS, mL/100 mL/min), and tumor vascularity (plasma volume, %) were calculated using a 2-compartment uptake model. DCE-CT parameters were correlated with immunohistochemical assessments of tumor vascularity (RECA-1), cell proliferation (Ki-67), and apoptosis (TUNEL). RESULTS: Sorafenib significantly (P < 0.05) suppressed tumor perfusion (25.1 ± 9.8 to 9.5 ± 6.0 mL/100 mL/min), tumor vascularity (15.6% ± 11.4% to 5.4% ± 2.1%), and PS (8.7 ± 4.5 to 2.7 ± 2.5 mL/100 mL/min) in prostate carcinomas during the treatment course. Immunohistochemistry revealed significantly lower tumor vascularity in the therapy group than in the control group (RECA-1; 181 ± 24 vs. 314 ± 47; P < 0.05). In sorafenib-treated tumors, significantly more apoptotic cells (TUNEL; 7132 ± 3141 vs. 3722 ± 1445; P < 0.05) and significantly less proliferating cells (Ki-67; 9628 ± 1.298 vs. 17,557 ± 1446; P < 0.05) were observed than those in the control group. DCE-CT tumor perfusion correlated significantly (P < 0.05) with tumor cell proliferation (Ki-67; r=0.55). DCE-CT tumor vascularity correlated significantly (P < 0.05) with immunohistochemical tumor cell apoptosis (TUNEL; r=-0.59) and tumor cell proliferation (Ki-67; r=0.68). DCE-CT endothelial PS correlated significantly (P < 0.05) with immunohistochemical tumor cell apoptosis (TUNEL; r=-0.6) and tumor vascularity (RECA-1; r=0.53). While performing corrections for multiple comparisons, we observed a significant correlation only between DCE-CT tumor vascularity (RECA-1) and tumor cell proliferation (Ki-67). CONCLUSION: Sorafenib significantly suppressed tumor perfusion, tumor vascularity, and PS quantified by DCE-CT in experimental prostate carcinomas in rats. These functional CT surrogate markers showed moderate correlations with antiangiogenic, antiproliferative, and proapoptotic effects observed by immunohistochemistry. DCE-CT may be applicable for the quantification of noninvasive imaging biomarkers of therapy response to antiangiogenic therapy.