A Cholecystokinin B Receptor-Specific Aptamer Does Not Activate Receptor Signaling

Targeted nanoparticles which deliver effective doses of chemotherapeutic drugs directly to pancreatic tumors could improve treatment efficacy without the toxicities associated with systemic drug administration. One protein on tumor cells that can be targeted by nanoparticles is a G-protein coupled cell surface receptor, the cholecystokinin B receptor (CCKBR). Previously, we had shown that attaching the CCKBR ligand gastrin to the surface of nanoparticles can enhance their up-take by tumors. The drawback of using gastrin is that it can also activate the receptor, causing tumor cell growth. This study shows that a DNA aptamer that binds to the CCKBR and enhances nanoparticle up-take by tumors does not activate this receptor. PANC-1 cells, a cultured human pancreatic cancer cell line, were treated for 24 h with CCKBR aptamer 1153. Cell lysates were run on Bis-Tris gels, transferred to membranes, blocked in 5% BSA and incubated overnight with primary antibodies, including antibodies directly against phosphorylated-Akt (Ser473), total Akt, and beta-actin, a protein loading control. Although the CCKBR aptamer 1153 is internalized by pancreatic cancer cells in a receptor-mediated fashion, it does not stimulate cell proliferation. Because of this, we anticipate that it will not activate CCKBR signaling. If aptamer 1153 does not activate downstream receptor signaling, our future work will test whether the aptamer could be used to specifically direct drug-containing nanoparticles to tumors, making chemotherapy treatments for pancreatic cancer patients more effective with fewer off-target effects and toxicity

Metastasis of hormone-independent breast cancer to lung and bone is decreased by α-difluoromethylornithine treatment

INTRODUCTION: Polyamines affect proliferation, differentiation, migration and apoptosis of cells, indicating their potential as a target for cancer chemotherapy. Ornithine decarboxylase converts ornithine to putrescine and is the rate-limiting step in polyamine synthesis. α-Difluoromethylornithine (DFMO) irreversibly inhibits ornithine decarboxylase and MDA-MB-435 human breast cancer metastasis to the lung without blocking orthotopic tumor growth. This study tested the effects of DFMO on orthotopic tumor growth and lung colonization of another breast cancer cell line (MDA-MB-231) and the effects on bone metastasis of MDA-MB-435 cells. METHODS: MDA-MB-231 cells were injected into the mammary fat pad of athymic mice. DFMO treatment (2% per orally) began at the day of tumor cell injection or 21 days post injection. Tumor growth was measured weekly. MDA-MB-231 cells were injected into the tail vein of athymic mice. DFMO treatment began 7 days prior to injection, or 7 or 14 days post injection. The number and incidence of lung metastases were determined. Green fluorescent protein-tagged MDA-MB-435 cells were injected into the left cardiac ventricle in order to assess the incidence and extent of metastasis to the femur. DFMO treatment began 7 days prior to injection. RESULTS: DFMO treatment delayed MDA-MB-231 orthotopic tumor growth to a greater extent than growth of MDA-MB-435 tumors. The most substantial effect on lung colonization by MDA-MB-231 cells occurred when DFMO treatment began 7 days before intravenous injection of tumor cells (incidence decreased 28% and number of metastases per lung decreased 35–40%). When DFMO treatment began 7 days post injection, the incidence and number of metastases decreased less than 10%. Surprisingly, treatment initiated 14 days after tumor cell inoculation resulted in a nearly 50% reduction in the number of lung metastases without diminishing the incidence. After intracardiac injection, DFMO treatment decreased the incidence of bone metastases (55% vs 87%) and the area occupied by the tumor (1.66 mm(2 )vs 4.51 mm(2), P < 0.05). CONCLUSION: Taken together, these data demonstrate that DFMO exerts an anti-metastatic effect in more than one hormone-independent breast cancer, for which no standard form of biologically-based treatment exists. Importantly, the data show that DFMO is effective against metastasis to multiple sites and that treatment is generally more effective when administered early

Expressions 1982

This fifth edition of EXPRESSIONS is the culmination of student efforts in the 1982 Creative Writing Contest, Campus Chronicle Photography Contest, Art and Commercial Art courses at Des Moines Area Community College. Journalism students did the design, typography and lay-out work necessary to bring these efforts together in this 1982 edition of EXPRESSIONS.https://openspace.dmacc.edu/expressions/1004/thumbnail.jp

Utilizing Peptide Ligand GPCRs to Image and Treat Pancreatic Cancer

It is estimated that early detection of pancreatic ductal adenocarcinoma (PDAC) could increase long-term patient survival by as much as 30% to 40% (Seufferlein, T. et al., Nat. Rev. Gastroenterol. Hepatol. 2016, 13, 74&ndash;75). There is an unmet need for reagents that can reliably identify early cancerous or precancerous lesions through various imaging modalities or could be employed to deliver anticancer treatments specifically to tumor cells. However, to date, many PDAC tumor-targeting strategies lack selectivity and are unable to discriminate between tumor and nontumor cells, causing off-target effects or unclear diagnoses. Although a variety of approaches have been taken to identify tumor-targeting reagents that can effectively direct therapeutics or imaging agents to cancer cells (Liu, D. et al., J. Controlled Release 2015, 219, 632&ndash;643), translating these reagents into clinical practice has been limited, and it remains an area open to new methodologies and reagents (O&rsquo;Connor, J.P. et al., Nat. Rev. Clin. Oncol. 2017, 14, 169&ndash;186). G protein&ndash;coupled receptors (GPCRs), which are key target proteins for drug discovery and comprise a large proportion of currently marketed therapeutics, hold significant promise for tumor imaging and targeted treatment, particularly for pancreatic cancer

Expression of the Human Gastrin Receptors CCKBR and CCKCR in PANCO2 Murine Pancreatic Cancer Cells

Pancreatic cancer is the fourth leading cause of cancer mortality. The gastrointestinal hormone, gastrin, stimulates human pancreatic cancer growth via its receptor, CCKBR, and a splice-variant of CCKBR (termed CCKCR) detected only in cancer cells. This cancer-associated variant retains the fourth intron and exhibits increased cell signaling. Recently, a single nucleotide polymorphism (SNP; C\u3eA) in the fourth intron was identified. Presence of SNP(A) was correlated with CCKCR protein expression in pancreatic cancer specimens and decreased patient survival. We hypothesize expression of human CCKCR in murine pancreatic cancer cells will increase tumor growth and metastasis relative to expression of human CCKBR. The aim of this study was to engineer PANC02 murine pancreatic cancer cells to express hCCKBR or hCCKCR (each SNP form). While vectors encoding hCCKBR and hCCKCR-SNP(C) existed, a version mutated to form the SNP(A) required sub-cloning into a vector capable of transfection and selection in mammalian cells. The gene was excised and ligated into pCAGEN.neo. Resulting clones were screened by restriction analysis to confirm insertion and correct orientation of the gene. DNA sequencing confirmed the status of the SNP in each vector. Next, hCCKBR, hCCKCR-SNP(C), hCCKCR-SNP(A) and empty-vector (pCAGEN.neo) were transfected in parallel into PANC02 cells. Resulting neomycin-resistant clones were isolated and RNA was harvested. Clones were screened for up-regulation of corresponding receptor mRNA by endpoint RT-PCR (GAPDH , loading control). Up-regulation was evident in most clones: hCCKBR (11 of 11 clones), hCCKCR-SNP(C) (7 of 10), and hCCKCR-SNP(A) (11 of 11). Real-time RT-PCR analysis is currently ongoing to more precisely quantify mRNA expression relative to empty-vector controls. The results of this study will permit in vivo tumor growth and immunotherapy studies in an immune-competent syngeneic murine model