**1. Introduction**

Lymphoma is the most common neoplasm of the canine hemolymphatic system. It represents about 15% of all malignant neoplasms. It has a very poor prognosis, the mean survival with high grade non Hodgkin's lymphomas being two months without treatment [1]. Incidence is increasing.

Lymphomas in dogs, as in humans, can be divided into numerous types depending on the cell line involved and their immunophenotype [2]. T lymphomas have a worse prognosis than B lymphomas and late clinical stages obviously have a very short survival period.

Lymphomas are known in human medicine to respond to chemotherapy, and some of them can even be cured by complex chemotherapy protocols, although severe side effects are noted.

Chemotherapy protocols have also been developed for dogs in the last 40 years. They have proved to be effective for the overall survival of the treated animal. Although the order of drug administration and duration of the maintenance part of the protocol vary considerably, most oncologists agree that a doxorubicin-based (eg CHOP) combination chemotherapy protocol provides the longest period of disease control and overall survival [3].

Even with chemotherapy, survival is relatively short and adjuvant therapies have been developed to improve prognosis. Immunotherapy protocols are of particular interest for this purpose as they may arm immune system cells against the abnormal proteins synthesized by cancer cells [4]. It is a selective way of destroying cancer cells and a treatment with much fewer side effects than chemo- or radiotherapy.

The immune system can destroy cancer cells by different methods:

© 2013 Simonet et al.; licensee InTech. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2013 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


Several kinds of immunotherapy protocol are available both in human and veterinary medicine [4, 5, 6, 7]. Heat shock proteins (HSPs) such as gp96 or HSPs70 which are synthesized by the cells submitted to stress are advantageous vaccination adjuvants due to their chaperone properties and their role in antigen presentation [8]. As chaperone molecules, almost all the cell peptides are associated with these proteins and HSP purification provides a fingerprint of the cell's protein synthesis [9]. This is particularly useful for cancer cells which synthesize numerous abnormal proteins during their natural progress [10]. These cells being genetically unstable, their abnormal protein synthesis differs from patient to patient and during the course of the disease. The HSPs and their associated peptides (AAPs) have special receptors (CD91) on dendritic cells which allow the internalization of the AAPs and their modification in order to be expressed at the surface of class I HLA proteins on the cell membrane, triggering activation of the CD8 T cells if they are abnormal [11].

Cancer cells are stressed by the mechanical and metabolic characteristics of the tumour. They synthesize many HSPs [12]. We thus isolated these proteins in order to make an autologous vaccine against the tumour. The HSPs were purified using a hydroxylapatite powder (HA) column. The powder carrying the HSPs was then injected subcutaneously to stimulate the immune system's response to the tumour.

Purification of HSPs using classical way is long and tedious. The use of hydroxylapatite powder allows a much faster purification process. Hydroxylapatite chromatography has been described by Tiselius in the early seventies. Hydroxylapatite chromatography is an adsorption chromatography. The adsorption mechanism is very poorly understood. The surface of the material is occupied bu Ca++ and PO<sup>4</sup> - . These ions are supposed to interact with the chemical groups of the proteins. However, post synthesis treatment such as sintering or spray- drying process modify the physico-chemical properties of the material surface. Furthermore, the interaction of the material surface with biological fluids triggers epitaxial growth of carbonated apatite at the surface of the material [13].

It was decided that the proteins purified will be injected carried by the particles for several reasons: the hydroxylapatite particles have been described as vaccine adjuvant, they are phagocytosed by the APCs and can deliver the proteins directly in the APCs, they trigger an afflux of APCs at the injection site [14]. Most of the adjuvants used in antiinfectious vaccines are nano or microparticular.

The aims of this study was to check the feasibility of this protocol using HA-particles with dogs suffering of high grade lymphoma and to know if secondary effects were detected.
