Abstract
A promising therapy for patients with B-cell lymphoma is based on vaccination with idiotype monoclonal antibodies (mAbs). Since idiotypes are different in each tumor, a personalized vaccine has to be produced for each patient. Expression of immunoglobulins with appropriate post-translational modifications for human use often requires the use of stable mammalian cells that can be scaled-up to reach the desired level of production. We have used a noncytopathic self-amplifying RNA vector derived from Semliki Forest virus (ncSFV) to generate BHK cell lines expressing murine follicular lymphoma-derived idiotype A20 mAb. ncSFV/BHK cell lines expressed approximately 2 mg/L/24 h of A20 mAb with proper quaternary structure and a glycosylation pattern similar to that of A20 mAb produced by hybridoma cells. A20 mAb purified from the supernatant of a ncSFV cell line, or from the hybridoma, was conjugated to keyhole limpet hemocyanin and used to immunize Balb/c mice by administration of four weekly doses of 25 µg of mAb. Both idiotype mAbs were able to induce a similar antitumor protection and longer survival compared to non-immunized mice. These results indicate that the ncSFV RNA vector could represent a quick and efficient system to produce patient-specific idiotypes with potential application as lymphoma vaccines.
Highlights
A promising therapy for patients with B-cell lymphoma is based on vaccination with idiotype monoclonal antibodies
In order to generate mammalian stable cell lines expressing the Id A20, we cloned the genes coding for its heavy (HC) and light chains (LC) into ncSFV-pac vector as described in “Materials and methods”
Since efficient production of the monoclonal antibodies (mAbs) will require an optimal ratio of LC:HC18, we cloned their coding genes into ncSFV-pac using four different configurations, generating the following vectors: (i) ncSFV-pac-A20sgPr, in which HC is followed by LC using independent viral subgenomic promoters, (ii) ncSFV-pac-A20HLC, in which LC is followed by HC using independent viral subgenomic promoters, (iii) ncSFV-pac-A20-2A, in which HC is fused to LC using the foot and mouth disease virus 2A autoprotease sequence as a linker, and (iv) ncSFV-pac-A20ires, having LC and HC separated by an IRES sequence (Fig. 1)
Summary
A promising therapy for patients with B-cell lymphoma is based on vaccination with idiotype monoclonal antibodies (mAbs). Both idiotype mAbs were able to induce a similar antitumor protection and longer survival compared to non-immunized mice These results indicate that the ncSFV RNA vector could represent a quick and efficient system to produce patient-specific idiotypes with potential application as lymphoma vaccines. The idiotype (Id) of a B-cell lymphoma refers to the unique amino acid sequences of the variable fragments of the heavy and light chains of the surface immunoglobulin expressed by tumor cells For this reason, the Id represents a patient- and tumor-specific antigen with great potential for lymphoma-targeted therapy[1,2]. Passive immunotherapy using anti-Id monoclonal antibodies (mAbs) has been shown to generate clinical responses, relapse is likely to occur due to the emergence of tumor escape m utants[3,4].
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