Selective sigma-2 ligands preferentially bind to pancreatic adenocarcinomas: applications in diagnostic imaging and therapy - PubMed (original) (raw)

doi: 10.1186/1476-4598-6-48.

Jonathan E McDunn, Peter O Simon Jr, Peter S Goedegebuure, Jinbin Xu, Lynne Jones, Katherine Chang, Fabian Johnston, Kathryn Trinkaus, Richard S Hotchkiss, Robert H Mach, William G Hawkins

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Selective sigma-2 ligands preferentially bind to pancreatic adenocarcinomas: applications in diagnostic imaging and therapy

Hiroyuki Kashiwagi et al. Mol Cancer. 2007.

Abstract

Background: Resistance to modern adjuvant treatment is in part due to the failure of programmed cell death. Therefore the molecules that execute the apoptotic program are potential targets for the development of anti-cancer therapeutics. The sigma-2 receptor has been found to be over-expressed in some types of malignant tumors, and, recently, small molecule ligands to the sigma-2 receptor were found to induce cancer cell apoptosis.

Results: The sigma-2 receptor was expressed at high levels in both human and murine pancreas cancer cell lines, with minimal or limited expression in normal tissues, including: brain, kidney, liver, lung, pancreas and spleen. Micro-PET imaging was used to demonstrate that the sigma-2 receptor was preferentially expressed in tumor as opposed to normal tissues in pancreas tumor allograft-bearing mice. Two structurally distinct sigma-2 receptor ligands, SV119 and WC26, were found to induce apoptosis to mice and human pancreatic cancer cells in vitro and in vivo. Sigma-2 receptor ligands induced apoptosis in a dose dependent fashion in all pancreatic cell lines tested. At the highest dose tested (10 muM), all sigma-2 receptor ligands induced 10-20% apoptosis in all pancreatic cancer cell lines tested (p < 0.05). In pancreas tumor allograft-bearing mice, a single bolus dose of WC26 caused approximately 50% apoptosis in the tumor compared to no appreciable apoptosis in tumor-bearing, vehicle-injected control animals (p < 0.0001). WC26 significantly slowed tumor growth after a 5 day treatment compared to vehicle-injected control animals (p < 0.0001) and blood chemistry panels suggested that there is minimal peripheral toxicity.

Conclusion: We demonstrate a novel therapeutic strategy that induces a significant increase in pancreas cancer cell death. This strategy highlights a new potential target for the treatment of pancreas cancer, which has little in the way of effective treatments.

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Figures

Figure 1

Figure 1

Structure of sigma ligands utilized in this paper. Panel A: Haloperidol is a non-specific sigma-1 and sigma-2 receptor ligand. Panel B: Pentazocine is a sigma-1 specific ligand. Panel C: Sigma-2 receptor-specific ligands SV119 and WC26. Panel D: Chemically-labeled sigma-2 receptor-specific ligands K05-138 (fluorescein-labeled), RHM-1 (3H-labeled), and RHM-4 (18F-labeled).

Figure 2

Figure 2

Sigma-2 ligands preferentially bind to cancer as opposed to normal tissues. (A): Single cell suspensions were prepared from liver, lung, pancreas, brain, kidney, spleen and pancreatic tumors (panc-02 bearing mice). Single cell suspensions were incubated for 1 hour with KO5-138 (a fluorescent labeled sigma-2 receptor ligand, at 50 nM (thin doted line) or 100 nM (thick solid line) of ligand, or left unstained (thin solid line). FACS histograms demonstrate 100 fold more florescence in tumors compared to normal tissues. (B): Human pancreatic cancer lines (CFPAC-1, Panc-1, AsPC-1) demonstrate similar high degree of sigma-2 ligand binding to when compared to our murine model (Panc-02). Fluorescein signal peaks were shown in unstained control (thin solid line), 50 nM of ligand (dotted line), or 100 nM of ligand (thick solid line). The experiment was repeated twice with identical results. (C): Representative Micro PET/CT images of pancreas adenocarcinoma (Panc-02) bearing C57BL/6 mice administered RHM-4 (18F-labled sigma-2 ligand). The tumor indicated by arrows was approximately 1 cm3. The additional hot spot represents metastatic tumor in a regional lymph node.

Figure 3

Figure 3

Scatchard analysis of [3H]RHM-1 binding to the sigma-2 receptors in membrane homogenates from Panc-02 tumor allografts. (A): Representative saturation binding experiments which show the total bound, non-specific bound and specific bound. (B): Representative Scatchard plots which were used to determine _K_d and _B_max values.

Figure 4

Figure 4

Sigma-2 receptor ligands induce apoptosis of pancreatic cancer cells via a caspase 3 dependent pathway. (A): Treatment with WC26, SV119 and haloperidol (Halo.) caused dose-dependent apoptosis in all pancreatic cancer cell lines tested. At the highest dose tested, all sigma-2 receptor ligands resulted in significant apoptosis (25–35%, p < 0.05). (B): Sigma-2 related apoptosis was prevented by pan-caspase inhibitor (Z-VAD-fmk) and caspase-3/7 inhibitor (DEVD-CHO), while the caspase-1 inhibitor (YVAD-CHO) had no effect (Panc-02 cells).

Figure 5

Figure 5

Systemic administration of Sigma-2 receptor ligand induces substantial tumor apoptosis without major toxicity. (A): Systemic administration of the sigma-2 receptor ligand WC26 induces considerable apoptosis in Panc-02 tumor allografts (up to 50% of tumor cells were active caspase-3 positive following a single 2 mg dose of WC26, p < 0.0001). Data reported as percent specific apoptosis (percent specific apoptosis = observed apoptosis – tissue specific background apoptosis). (B): The mice appeared normal and no apparent toxicity was noted in serum biochemical analysis or by immunohistochemistry (data not shown). Reference ranges for the blood chemistry panel in C57Bl/6 mice: ALT: 18–184 U/L, AST: 55–251 U/L, glucose: 174–335 mg/dL, amylase: 2595 +/- 212 U/L, BUN: 34–58 mg/dL, creatinine < 1.1 mg/dL.

Figure 6

Figure 6

Systemic administration of sigma-2 receptor ligand slows tumor growth and improves survival. Mice with established pancreas tumors were treated with 5 days of WC26 in one of two dose ranges (2 mg/day, n = 9; Dash line or 1 mg/day n = 10; dashed line) or vehicle control (20% DMSO n = 10; solid line). (A): Mice treated with WC26 had smaller tumors compared to animals vehicle control (p < 0.0001). The two treatment groups were not statistically different. The double-headed arrow denotes the treatment period (5 days). **(B):** Survival of mice treated with WC26 compared favorably to the mice treated vehicle control (p= 0.002). Survival endpoints were defined as tumor diameter >15 mm or tumor ulceration.

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