In Vitro and in Vivo oncolytic Activity of Lasota Strain of Newcastle Disease Virus on a Lymphoma B-Cell Line and a Canine Cutaneous T-Cell Lymphoma

Oncolytic viruses, either naturally or genetically modified, possess the ability to kill cancer cells. Newcastle Disease Virus (NDV), a type I avian paramyxovirus, has demonstrated a selective ability to kill cancer cells directly as well as through immunostimulation. NDV is able to infect more than 250 species of birds, particularly poultry and is considered a zoonosis, primarily causing conjunctivitis. Importantly, no infections have been reported in mammalian species nor is there evidence of human to human transmission. We use the LaSota strain which is lentogenic (less pathogenic). We seek to study the in vitro oncolytic (lympholytic) potential of NDV on healthy mononuclear and malignant lymphocytes and the in vivo oncolytic therapeutic potential in dogs with lymphoma.

LaSota vaccine strain (LSV) was propagated in pathogen –free 9 day old chicken embrios. Viral titers were reported as mean infective dose. Healthy human and canine mononuclear cells as well as a human diffuse large B cell lymphoma cell line (DHL4 cell line) were exposed to medium alone, LSV at a multiplicity of infection (MOI) of 0.1, 10 and 100 or CHOP (cyclophosphamide, doxorubicin, vincristine, prednisone) chemotherapy. Apoptosis was assessed by flow cytometry using annexin V/FITC/IT. Cell counts were carried out at 48 and 72 hs. Data was analyzed by FlowJo 9 and Prisma 6 Softwares. Interestingly, cells at the highest MOI (100) had the same rate of apoptosis as those exposed to chemotherapy, both significantly higher than negative controls.

The in vivo response and toxicity to intravenous and intratumoral inyection of LSV in a dog with naturally occurring aggressive, multicentric cutaneous T-cell lymphoma is reported. The patient had a 4 week history of paraplegia due to lymphomatous infiltration of the spinal canal and a 3 week history of rapid development of multiple elevated cutaneous lesions ranging from (from 0.5 by 0.5 cm to 15 cm in diameter and 2 cm in thickness). After informed consent, LSV was given in a single occasion with 1x10⁶ mean (50%) infective dose in tissue culture (TCDI 50) in only one lesion, as well as intravenously with a TCDI 50 of 1x10¹² in order to assess response of distant non-virally infiltrated lesions. At four weeks of follow up, no new lesions have appeared. All tumors are responding with complete flattening of the lesions and decrease in diamenter of all lesions. Spontaneous bleeding from the lesions was seen from days 4 through 7 after treatment. All laboratory work up is within normal limits except for indirect hiperbilirrubinemia (2x above normal) in the 4th week (Coombs results are pending). Consecutive samples for histology, electron microscopy and viral load are being evaluated. The patient has had an excellent performance status with no evident clinical toxicity and has begun to wag his tail and show slow movement of the posterior limbs with gain in sensitivity.

We have demonstrated the specific in vitro, oncolytic/lympholytic activity of NCV (LaSota) on human lymphoma- derived cells with minimal tropism towards healthy cells. Response to treatment of the first patient (dog) is encouraging. Longer follow up as well as histologic evaluation and viral shedding are being evaluated.