AGR2, a unique tumor-associated antigen, is a promising candidate for antibody targeting.

Anterior gradient 2 (AGR2), a protein disulfide isomerase, shows two subcellular localizations: intracellular (iAGR2) and extracellular (eAGR2). In healthy cells that express AGR2, the predominant form is iAGR2, which resides in the endoplasmic reticulum. In contrast, cancer cells secrete and express eAGR2 on the cell surface. We wanted to test if AGR2 is a cancer-specific tumor-associated antigen. We utilized two AGR2 antibodies, P3A5 and P1G4, for in vivo tumor localization and tumor growth inhibition. The monoclonal antibodies recognized both human AGR2 and mouse Agr2. Biodistribution experiments using a syngeneic mouse model showed high uptake of P3A5 AGR2 antibody in xenografted eAgr2+ pancreatic tumors, with limited uptake in normal tissues. In implanted human patient-derived eAGR2+ pancreatic cancer xenografts, tumor growth inhibition was evaluated with antibodies and Gemcitabine (Gem). Inhibition was more potent by P1G4 + Gem combination than Gem alone or P3A5 + Gem. We converted these two antibodies to human:mouse chimeric forms: the constructed P3A5 and P1G4 chimeric mVLhCκ and mVHhCγ (γ1, γ2, γ4) genes were inserted in a single mammalian expression plasmid vector, and transfected into human 293F cells. Expressed human:mouse chimeric IgG1, IgG2 and IgG4 antibodies retained AGR2 binding. Increase in IgG yield by transfected cells could be obtained with serial transfection of vectors with different drug resistance. These chimeric antibodies, when incubated with human blood, effectively lysed eAGR2+ PC3 prostate cancer cells. We have, thus, produced humanized anti-AGR2 antibodies that, after further testing, might be suitable for treatment against a variety of eAGR2+ solid tumors.University of Washington CoMotion FundNCI-EDRN Biomarker Developmental Lab grant U01CA111244, and DoD W81XWH-16-1-0614

Perineural invasion in pancreatic cancer: proteomic analysis and in vitro modelling.

Perineural invasion (PNI) is a common and characteristic feature of pancreatic ductal adenocarcinoma (PDAC) that is associated with poor prognosis, tumor recurrence, and generation of pain. However, the molecular alterations in cancer cells and nerves within PNI have not previously been comprehensively analysed. Here, we describe our proteomic analysis of the molecular changes underlying neuro-epithelial interactions in PNI using liquid chromatography-mass spectrometry (LC-MS/MS) in microdissected PNI and non-PNI cancer, as well as invaded and non-invaded nerves from formalin-fixed, paraffin-embedded PDAC tissues. In addition, an in vitro model of PNI was developed using a co-culture system comprising PDAC cell lines and PC12 cells as the neuronal element. The overall proteomic profiles of PNI and non-PNI cancer appeared largely similar. In contrast, upon invasion by cancer cells, nerves demonstrated widespread plasticity with a pattern consistent with neuronal injury. The upregulation of SCG2 (secretogranin II) and neurosecretory protein VGF (non-acronymic) in invaded nerves in PDAC tissues was further validated using immunohistochemistry. The tested PDAC cell lines were found to be able to induce neuronal plasticity in PC12 cells in our in vitro established co-culture model. Changes in expression levels of VGF, as well as of two additional proteins previously reported to be overexpressed in PNI, Nestin and Neuromodulin (GAP43), closely recapitulated our proteomic findings in PDAC tissues. Furthermore, induction of VGF, while not necessary for PC12 survival, mediated neurite extension induced by PDAC cell lines. In summary, here we report the proteomic alterations underlying PNI in PDAC and confirm that PDAC cells are able to induce neuronal plasticity. In addition, we describe a novel, simple, and easily adaptable co-culture model for in vitro study of neuro-epithelial interactions

Polymers imprinted with three REG1B peptides for electrochemical determination of Regenerating Protein 1B, a urinary biomarker for pancreatic ductal adenocarcinoma

Three peptides (each containing 13–18 amino acids) were synthesized and used as templates for molecular imprinting and epitope recognition of the Regenerating Protein 1B (REG1B), which is one of the urinary biomarkers for pancreatic ductal adenocarcinoma (PDAC). Poly(ethylene-co-vinyl alcohol)s were employed as the host for molecular imprinting of the peptides. Following their preparation, the molecularly imprinted polymers (MIP) were examined by cyclic voltammetry. The electrochemical responses of a screen-printed gold substrate coated with the MIP were measured at a working voltage of 300 mV (vs. Ag/AgCl); the entire protein and the peptides gave similar responses at concentrations of <1.0 pg⋅mL−1, with detection limits as low as 0.1 pg⋅mL−1. Urine samples from healthy and PDAC patients were then analyzed by using this modified gold electrode, and the results are in agreement with data obtained with ELISA

Rhinitis, Asthma and Respiratory Infections among Adults in Relation to the Home Environment in Multi-Family Buildings in Sweden

Risk factors for rhinitis, asthma and respiratory infections in the home environment were studied by a questionnaire survey. Totally 5775 occupants (&gt;= 18 years old) from a stratified random sample of multi-family buildings in Sweden participated (46%). 51.0% had rhinitis in the last 3 months (current rhinitis); 11.5% doctor diagnosed asthma; 46.4% respiratory infections in the last 3 months and 11.9% antibiotic medication for respiratory infections in the last 12 months. Associations between home environment and health were analyzed by multiple logistic regression, controlling for gender, age and smoking and mutual adjustment. Buildings constructed during 1960-1975 were risk factors for day time breathlessness (OR = 1.53, 95%CI 1.03-2.29). And those constructed during 1976-1985 had more current rhinitis (OR = 1.43, 95%CI 1.12-1.84) and respiratory infections (OR = 1.46, 95%CI 1.21-1.78). Cities with higher population density had more current rhinitis (p = 0.008) and respiratory infections (p&lt;0.001). Rented apartments had more current rhinitis (OR = 1.23, 95%CI 1.07-1.40), wheeze (OR = 1.20, 95%CI 1.02-1.41), day time breathlessness (OR = 1.31, 95%CI 1.04-1.66) and respiratory infections (OR = 1.13, 95%CI 1.01-1.26). Living in colder parts of the country was a risk factor for wheeze (p = 0.03) and night time breathlessness (p = 0.002). Building dampness was a risk factor for wheeze (OR = 1.42, 95%CI 1.08-1.86) and day time breathlessness (OR = 1.57, 95%CI 1.09-2.27). Building dampness was a risk factor for health among those below 66 years old. Odor at home was a risk factor for doctor diagnosed asthma (OR = 1.49, 95%CI 1.08-2.06) and current asthma (OR = 1.52, 95%CI 1.03-2.24). Environmental tobacco smoke (ETS) was a risk factor for current asthma (OR = 1.53, 95%CI 1.09-2.16). Window pane condensation was a risk factor for antibiotic medication for respiratory infections (OR = 1.41, 95%CI 1.10-1.82). In conclusion, rhinitis, asthma and respiratory infections were related to a number of factors in the home environment. Certain building years (1961-1985), building dampness, window pane condensation and odor in the dwelling may be risk factors