Preclinical development of human granulocyte-macrophage colony-stimulating factor-transfected melanoma cell vaccine using established canine cell lines and normal dogs

Abstract

Tumor vaccines and gene therapy have received significant attention as means of increasing cellular and humoral immune responses to cancer. We conducted a pilot study of seven research dogs to determine whether intradermal injection of canine tumor cells transfected via the Accell particle-mediated gene transfer device with the cDNA for human granulocyte-macrophage colony-stimulating factor (hGM-CSF) would generate biologically relevant levels of protein and result in demonstrable histological changes at sites of vaccination. Tumor cell vaccines of 10 7 irradiated canine melanoma cells were nontoxic, safe, and well tolerated. No significant alterations in blood chemistry values or hematological profiles were detected. A histological review of control vaccine sites revealed inflammatory responses predominated by eosinophils, whereas vaccine sites with hGM-CSF-transfected tumor cells had an influx of neutrophils and macrophages. Enzyme-linked immunosorbent assays of skin biopsies from vaccine sites had local hGM-CSF production (8.68 -16.82 ng/site of injection) at 24 hours after injection and detectable levels (0.014 -0.081 ng/site) for Յ2 weeks following vaccination. Flow cytometric analysis of hGM-CSF-transfected cells demonstrated Յ25% transfection efficiency, and hGM-CSF levels obtained during time-course assays demonstrated biologically relevant levels for both irradiated and nonirradiated samples. These data demonstrate the in vivo biological activity of irradiated hGM-CSF-transfected canine tumor cells and help provide evidence for a valid translational research model of spontaneous tumors.

Osteosarcoma cells as carriers to allow antitumor activity of canine oncolytic adenovirus in the presence of neutralizing antibodies

“…In canine cells, it has been shown that a radiation dose between 15 and 100 Gy is sufficient to block completely the cloning capacity of the tumor cells. 23 We confirmed that a single dose of 30 Gy on D17 and Abrams cell lines inhibited cell proliferation (data not shown). Consequently, canine cell lines were inactivated with a single dose of 30 Gy after infection and viral replication was assessed.…”

“…Biodistribution analysis was performed as previously described. 23 Briefly, frozen tissues were ground to a fine powder using a pestle and mortar and processed according to manufacturer instructions (Promega, Madison, WI). Luciferase enzyme activity in supernatants was assessed using luciferase assay reagent (Promega) and normalized to total protein content.…”

Cancer immunology and canine malignant melanoma: A comparative review

“…Note the multiple roles of the tyrosinase enzyme. different vaccination strategy and injected healthy dogs with lethally irradiated CMM cells transfected with human GM-CSF resulting in increased numbers of macrophages, which may act as APCs, at the vaccination site compared to non-transfected tumour cell vaccines (Hogge et al, 1999). In a further trial, Finocchiaro and Gilkin combined autologous/allogeneic formolised tumour cells as a vaccine injected concomitantly with lethally irradiated xenogeneic cells producing human IL-2 and human GM-CSF to treat CMM patients .…”

Enhancing antimelanoma immune responses through apoptosis

“…Three independent trials have examined the delivery of IL-2 or GM-CSF DNA with histoincompatible cells, 5 in combination with staphylococcal enterotoxin B, 2 or in autologous tumor vaccines. 37,38 The objective response rates for these studies were 50, 33, and 20%, respectively. The first two approaches led to median survival times that exceeded the control group or the anticipated survival based on historical data (39 and 24 weeks, respectively).…”