Idiotype-specific intravenous immunoglobulin (IVIG) for therapy of autoimmune diseases.

Intravenous immunoglobulin (IVIG) is used successfully for therapy of inflammatory and autoimmune diseases, especially in cases of conventional therapy resistance. Within the broad spectrum of immunomodulatory activities of IVIG in vitro and in vivo, the anti-idiotypic activity, neutralizing the autoimmune disease related idiotypes, is one of the main mechanism. We and others have proven that from the IVIG composition, diverse fractions of autoimmune disease specific IVIG can be affinity purified (sIVIG). This sIVIG was shown to be more efficient than the whole compound of IVIG in experimental animal models of autoimmune diseases.The affinity purification of disease sIVIG encompasses three stages. The first stage is to construct an autoantigen column for affinity purification of the autoantibodies. In the second stage the purified autoantibodies are used to construct a new column composed of the autoantibodies. The later is utilized for affinity purification of anti-autoantibodies (anti- idiotypes) IVIG defined as autoimmune disease specific IVIG- sIVIG.

Emerging antibody-based products.

Antibody-based products are not widely available to address many global health challenges due to high costs, limited manufacturing capacity, and long manufacturing lead times. There are now tremendous opportunities to address these industrialization challenges as a result of revolutionary advances in plant virus-based transient expression. This review focuses on some antibody-based products that are in preclinical and clinical development, and have scaled up manufacturing and purification (mg of purified mAb/kg of biomass). Plant virus-based antibody products provide lower upfront cost, shorter time to clinical and market supply, and lower cost of goods (COGs). Further, some plant virus-based mAbs may provide improvements in pharmacokinetics, safety and efficacy.

BiovaxID®: a customized idiotype vaccine for the treatment of B-cell lymphoma.

Most patients with B-cell lymphoma face an often incurable disease, particularly those diagnosed with an indolent subtype. The addition of passive immunotherapy to old and new chemotherapy regimens has improved both response rates and disease-free survival, leading in many cases to an extended overall survival. However, a cure remains elusive in most cases. For this reason, the patient- and tumor-specific idiotype, that is the collection of epitopes exclusively presented by the tumor clone's surface immunoglobulin, has been extensively studied as a privileged target for vaccine therapy, aiming at preventing disease re-occurrence after standard treatment. BiovaxID(®) (Biovest International, FL, USA), the most clinically advanced among such therapeutic vaccines, finds itself at a crucial turning point when it comes to further development. Both clinical trials in which it has been formally employed have shown intriguing results. Independent studies using slightly different versions of a conceptually identical vaccine provided all proofs of principle required to ascertain the vaccine's value - biological and clinical efficacy as well as clinical benefit. However, all these data have failed to bring an idiotype vaccine to the market owing to reasons that often have very little to do with the product itself. In fact, some successful studies were not conceived with this goal in mind, while others simply did not enroll enough patients to convincingly make their case for regulatory approval. It is likely that one or more new clinical trials will have to be successfully completed to reach the ultimate goal - that is, to make BiovaxID available to most patients and to adequately position it in the very crowded therapeutic algorithm of B-cell lymphoma.

Vaccination with patient-specific tumor-derived antigen in first remission improves disease-free survival in follicular lymphoma.

PURPOSE: Vaccination with hybridoma-derived autologous tumor immunoglobulin (Ig) idiotype (Id) conjugated to keyhole limpet hemocyanin (KLH) and administered with granulocyte-monocyte colony-stimulating factor (GM-CSF) induces follicular lymphoma (FL) -specific immune responses. To determine the clinical benefit of this vaccine, we conducted a double-blind multicenter controlled phase III trial. PATIENTS AND METHODS: Treatment-naive patients with advanced stage FL achieving complete response (CR) or CR unconfirmed (CRu) after chemotherapy were randomly assigned two to one to receive either Id vaccine (Id-KLH + GM-CSF) or control (KLH + GM-CSF). Primary efficacy end points were disease-free survival (DFS) for all randomly assigned patients and DFS for randomly assigned patients receiving at least one dose of Id vaccine or control. RESULTS: Of 234 patients enrolled, 177 (81%) achieved CR/CRu after chemotherapy and were randomly assigned. For 177 randomly assigned patients, including 60 patients not vaccinated because of relapse (n = 55) or other reasons (n = 5), median DFS between Id-vaccine and control arms was 23.0 versus 20.6 months, respectively (hazard ratio [HR], 0.81; 95% CI, 0.56 to 1.16; P = .256). For 117 patients who received Id vaccine (n = 76) or control (n = 41), median DFS after randomization was 44.2 months for Id-vaccine arm versus 30.6 months for control arm (HR, 0.62; 95% CI, 0.39 to 0.99; P = .047) at median follow-up of 56.6 months (range, 12.6 to 89.3 months). In an unplanned subgroup analysis, median DFS was significantly prolonged for patients receiving IgM-Id (52.9 v 28.7 months; P = .001) but not IgG-Id vaccine (35.1 v 32.4 months; P = .807) compared with isotype-matched control-treated patients. CONCLUSION: Vaccination with patient-specific hybridoma-derived Id vaccine after chemotherapy-induced CR/CRu may prolong DFS in patients with FL. Vaccine isotype may affect clinical outcome and explain differing results between this and other controlled Id-vaccine trials.

Stem cell transplant and idiotypic vaccination for B-cell malignancies.

Several types of B-cell malignancy, including but not limited to multiple myeloma and follicular lymphoma, are still considered incurable. In a substantial number of cases, patients must undergo either autologous or allogeneic stem cell transplantation as a standard of care procedure for their disease. Among experimental treatments for multiple myeloma and follicular lymphoma, idiotypic vaccination has been attempted over the last two decades with variable degrees of success. Few clinical trials have combined stem cell transplant procedures with idiotypic vaccination, and they are the subject of this review, which will also include some of our original data, as well as our overall evaluation of this field of clinical investigation. Although apparently at the opposite extremes of the therapeutic option array, toxicity-burdened stem cell transplantation and virtually innocuous idiotypic vaccination might well offer a sound curative opportunity to some patients with otherwise incurable B-cell malignancies, provided that the latter treatment first succeeds at obtaining regulatory approval.

Escherichia coli-based production of a tumor idiotype antibody fragment--tetanus toxin fragment C fusion protein vaccine for B cell lymphoma.

The unique immunoglobulin idiotype expressed on the surface of B lymphoma cells can be used as an effective antigen in tumor-specific vaccines when fused to immunostimulatory proteins and cytokines. A DNA vaccine encoding for an idiotype antibody single chain Fv (scFv) fragment fused to the Tetanus Toxin Fragment C (TTFrC) has been shown to induce protective anti-tumor responses. Protein-based strategies may be more desirable since they provide greater control over dosage, duration of exposure, and in vivo distribution of the vaccine. However, production of fusion protein vaccines containing complex disulfide bonded idiotype antibodies and antibody-derived fragments is challenging. We use an Escherichia coli-based cell-free protein synthesis platform as well as high-level expression of E. coli inclusion bodies followed by refolding for the rapid generation of an antibody fragment - TTFrC fusion protein vaccine. Vaccine proteins produced using both methods were shown to elicit anti-tumor humoral responses as well as protect from tumor challenge in an established B cell lymphoma mouse model. The development of technologies for the rapid production of effective patient-specific tumor idiotype-based fusion protein vaccines provides opportunities for clinical application.

[Idiotype vaccines in the treatment of follicular lymphoma: current status and future perspectives]. / Vacuna idiotípica en el tratamiento del linfoma folicular: situación actual y perspectivas futuras.

Follicular lymphoma is the second most prevalent non-Hodgkin lymphoma, representing 20% of all lymphomas. Follicular lymphoma is an indolent disease with a slow progression in which, although exhibiting a good response to treatment, relapse is very frequent and complete remission is not easy to maintain. Therefore, the disease is regarded as incurable. The search for new therapeutic strategies, together with a better understanding of the immune system, has led to the emergence of a new treatment named immunotherapy. Follicular lymphoma is a malignancy suitable for this kind of treatment given the fact that it is characterized by presenting a unique tumour-specific antigen: the idiotype of the monoclonal immunoglobulin displayed on the membrane of tumour cells. Several studies have been conducted to test immunotherapy as complementary to conventional treatment. In a previous study by our group, a clear benefit was evident is obtained after idiotypic vaccination, when an adequate immunization of the patient is obtained, in comparison to chemotherapy alone. In this sense, analysis is needed of whether idiotypic vaccination can produce not only long-lasting and complete remission, but even cure. It would be of great interest to consider an optimisation of the experimental design of clinical trials, an improvement of vaccine production, and the study of the molecular mechanisms of the tumour cell which modify the target immunoglobulin.

Tumor-specific recombinant idiotype immunisation after chemotherapy as initial treatment for follicular non-Hodgkin lymphoma.

Tumor-specific variable regions of the clonal immunoglobulin (idiotype, Id) expressed by B cell non-Hodgkin lymphoma (NHL) can be targeted by active immunotherapy. We conducted a phase I/II trial to determine the safety and immunogenicity of a patient-specific, recombinant, mammalian cell-derived Id protein conjugated to keyhole limpet hemocyanin (Id-KLH; MyVax personalised immunotherapy) in 22 patients with follicular NHL in first remission after chemotherapy. Subjects received five subcutaneous immunisations with MyVax plus locally administered granulocyte-macrophage colony-stimulating factor (GM-CSF). Among 21 evaluable patients, 62% mounted Id-specific immune responses. Evoked anti-Id antibodies recognised both recombinant Id and native Id, and could specifically stain autologous tumor cells. At median follow-up of more than 6 years, median progression-free survival is 38 months. Immunisation of follicular lymphoma patients with MyVax Id-KLH is safe and patients often mount tumor-specific immune responses. These results form the basis of a pivotal phase 3 trial of MyVax in follicular NHL.

Prolonged idiotypic vaccination against follicular lymphoma.

During the last 2 decades, idiotypic vaccination has provided proof of principle of biological efficacy, clinical efficacy and clinical benefit in small follicular lymphoma trials. However, with the exception of anecdotal reports, most patients have received no more than 10 doses of their customised idiotype (Id) vaccine. Therefore, it is not known whether prolonged usage of idiotypic vaccination is safe. Since 2002, 18 previously treated patients with follicular lymphoma have received extended idiotypic vaccination at our institution outside clinical trials. Vaccination was provided as a compassionate alternative to no further treatment, and was meant to be stopped only upon complete consumption of the available patient- and tumor-specific vaccine [Id-keyhole limpet hemocyanin + granulocyte-macrophage colony-stimulating factor (Id-KLH + GM-CSF)], or in case of disease relapse or any serious non-local toxicity. So far, 18 patients have received an average of 18 doses of Id vaccine (median: 17; mean: 18; range: 10-31). Eleven patients are still actively receiving idiotypic vaccination: some of them are now over more than 6 years. Toxicity has been systematically negligible and mostly local. No patient has abandoned the vaccination program because of toxicity. Prolonged idiotypic vaccination with the soluble protein Id-KLH + GM-CSF formulation is safe and well tolerated.

Plant-produced idiotype vaccines for the treatment of non-Hodgkin's lymphoma: safety and immunogenicity in a phase I clinical study.

Plant-made vaccines have been the subject of intense interest because they can be produced economically in large scale without the use of animal-derived components. Plant-made therapeutic vaccines against challenging chronic diseases, such as cancer, have received little research attention, and no previous human clinical trials have been conducted in this vaccine category. We document the feasibility of using a plant viral expression system to produce personalized (patient-specific) recombinant idiotype vaccines against follicular B cell lymphoma and the results of administering these vaccines to lymphoma patients in a phase I safety and immunogenicity clinical trial. The system allowed rapid production and recovery of idiotypic single-chain antibodies (scFv) derived from each patient's tumor and immunization of patients with their own individual therapeutic antigen. Both low and high doses of vaccines, administered alone or co-administered with the adjuvant GM-CSF, were well tolerated with no serious adverse events. A majority (>70%) of the patients developed cellular or humoral immune responses, and 47% of the patients developed antigen-specific responses. Because 15 of 16 vaccines were glycosylated in plants, this study also shows that variation in patterns of antigen glycosylation do not impair the immunogenicity or affect the safety of the vaccines. Collectively, these findings support the conclusion that plant-produced idiotype vaccines are feasible to produce, safe to administer, and a viable option for idiotype-specific immune therapy in follicular lymphoma patients.

Idiotype vaccination in patients with myeloma reduced circulating myeloma cells (CMC).

BACKGROUND: Circulating myeloma cells (CMC), exhibiting the same immunoglobulin heavy-chain gene rearrangements as the plasma cells, are part of the myeloma clone. In this study, we evaluated the effect of idiotype (Id) vaccination on CMC. PATIENTS AND METHODS: Eleven patients were immunized with the autologous Id in combinations with granulocyte-macrophage colony-stimulating factor and interleukin 12, and followed for CMC by quantitative real-time allele-specific PCR. Id-specific T cells were monitored by proliferation assay, enzyme-linked immunospot (interferon-gamma) assay, and quantitative real-time PCR for cytokines. Regulatory T (T(reg)) cells were analyzed by flow cytometry. RESULTS: CMC were detected in 9 of 11 patients at start of vaccination. In four patients, CMC declined and two had a complete molecular remission. Further two patients had stable levels of CMC during follow-up, while in three patients CMC progressively increased. Six patients had a vaccine-induced Id-specific T-cell response. A significant correlation was observed between reduced/stable levels of CMC and the Id-specific T cells (P < 0.02). The frequency of T(reg) cells was decreased in immune responders, but increased in immune nonresponders (P < 0.05). No significant change in the serum M-protein concentration was, however, observed in any patient. CONCLUSION: Id vaccination reduced CMC, which correlated with vaccine-induced Id-specific T cells. Further studies are warranted to analyze the clinical significance of CMC and clinical effects of Id vaccination.

Long-term idiotype vaccination combined with interleukin-12 (IL-12), or IL-12 and granulocyte macrophage colony-stimulating factor, in early-stage multiple myeloma patients.

PURPOSE AND EXPERIMENTAL DESIGN: Twenty-eight patients with immunoglobulin G myeloma stages I to II were immunized i.d. over 110 weeks with autologous M protein combined with interleukin-12 (IL-12; n = 15) or with IL-12 and granulocyte macrophage colony-stimulating factor (GM-CSF; n = 13). Idiotype-specific T-cell responses were assessed by [(3)H]thymidine incorporation, enzyme-linked immunospot assay, and delayed-type hypersensitivity reaction. RESULTS: Based on these three assays, idiotype-specific immune responses were noted in 5 of 15 (33%) patients in the IL-12 group and in 11 of 13 (85%) patients in the GM-CSF/IL-12 group (P < 0.01). Immune response was seen only in patients with M-component concentration of <50 g/L. Three of 16 (19%) responders showed a gradually increasing idiotype-specific T-cell response, whereas 11 of 16 (69%) patients showed initial response, which then disappeared rapidly; the latter pattern was frequently associated with subsequent progressive disease. Immune nonresponse was associated with an increase in the numbers of CD4(+)/CD25(+) cells (regulatory T cells), which was absent in responding patients. Median time to progression for immune responders (n = 16) was 108 weeks compared with 26 weeks for nonresponders (n = 12; P = 0.03). CONCLUSIONS: These results indicate that idiotype immunization of myeloma patients with GM-CSF and IL-12 may induce specific T-cell response more frequently than with IL-12 alone and that immune response may correlate with time to progression and nonresponse with increased numbers of regulatory T cells.

Targeting pemphigus autoantibodies through their heavy-chain variable region genes.

Pemphigus vulgaris (PV) is a potentially fatal blistering disease characterized by autoantibodies against cell surface adhesion proteins desmoglein (Dsg) 3 and Dsg1. Previous studies using phage display to clone Dsg-reactive monoclonal antibodies from a PV patient demonstrated that a limited number of antibody variable region genes encode the autoantibody repertoire, with different genes for pathogenic and non-pathogenic mAbs. Here, we investigated the feasibility of specific autoantibody targeting in pemphigus. We produced rabbit anti-idiotypic antibodies against two pathogenic and two non-pathogenic PV mAbs. Antisera inhibited binding of the immunizing mAb to Dsgs by ELISA as well as pathogenicity against cultured human keratinocytes. Antisera also inhibited other mAbs using the same variable region heavy chain (V(H)) genes, despite different light chains or somatic mutations. Additionally, peptide phage display identified peptide sequences that bound PV mAbs in a V(H)-specific manner. To evaluate the therapeutic potential of V(H) gene-targeted reagents, preimmune sera and antisera were used to adsorb pathogenic antibodies from PV sera. Pooled antisera significantly reduced pathogenic activity from the original PV patient's serum and bound pathogenic antibodies from two other PV sera, suggesting shared autoantibody V(H) gene usage among PV patients. Together, these data suggest novel V(H) gene-targeted approaches toward PV treatment.

BiovaxID: a personalized therapeutic cancer vaccine for non-Hodgkin's lymphoma.

The clonal immunoglobulin molecule, idiotype (ID), expressed on the surface of B-cell malignancies can function as a tumor-specific antigen. BiovaxID is a patient-specific therapeutic cancer vaccine composed of the tumor idiotype conjugated to a carrier protein, keyhole limpet hemocyanin (KLH). In a Phase II clinical trial, administration of ID-KLH vaccine together with granulocyte-macrophage colony-stimulating factor to follicular lymphoma patients in complete remission induced tumor-specific cellular and humoral immunity and molecular remissions, and was associated with prolonged disease-free survival. A randomized, double-blind, Phase III clinical trial is ongoing to definitively determine the clinical benefit of BiovaxID plus granulocyte-macrophage colony-stimulating factor vaccination in patients with follicular lymphoma.

Cell-free production of scFv fusion proteins: an efficient approach for personalized lymphoma vaccines.

The unique immunoglobulin (Ig) idiotype on the surface of each B-cell lymphoma represents an ideal tumor-specific antigen for use as a therapeutic vaccine. We have used an Escherichia coli-based, cell-free protein-expression system to produce a vaccine within hours of cloning the Ig genes from a B-cell tumor. We demonstrated that a fusion protein consisting of an idiotypic single chain Fv antibody fragment (scFv) linked to a cytokine (GM-CSF) or to an immunostimulatory peptide was an effective lymphoma vaccine. These vaccines elicited humoral immune responses against the native Ig protein displayed on the surface of a tumor and protected mice against tumor challenge with efficacy equal to that of the conventional Ig produced in a mammalian cell and chemically coupled to keyhole limpet hemocyanin. The cell-free E coli system offers a platform for rapidly generating individualized vaccines, thereby allowing much more efficient application in the clinic.

Effect of linkage of transduction domain sequences to a lymphoma idiotype DNA vaccine on vaccine effectiveness.

Patient idiotype-specific vaccines for treatment of non-Hodgkin's lymphoma have shown promise in clinical trials, encouraging efforts to enhance the effectiveness of idiotype vaccines further. It has previously been found that for some other types of experimental vaccines, the addition of transduction domains has improved vaccine immunogenicity. Transduction domains are short amino acid sequences that are capable of increasing transport through cellular membranes. In this study, we tested murine B cell 38C13 lymphoma idiotype DNA vaccines with human immunodeficiency virus (HIV) Tat-derived transduction sequences for efficacy against 38C13 challenge. The rate of tumor onset was similar for the idiotype and transduction domain-conjugated idiotype vaccine groups. At days 22-23 postchallenge, the number of surviving mice was significantly higher in the group that had received a DNA vaccine consisting of the 38C13 idiotype sequence plus modified Tat transduction sequence, in comparison with the group that received idiotype-only vaccines. Although the overall survival difference was not statistically significant following day 24, a trend toward an increased survival rate for mice receiving idiotype plus Tat-derived transduction domains was maintained through day 106 postchallenge. Thus, the addition to idiotype vaccines of specific sequences that facilitate intracellular transport may have potential to improve the effectiveness of such vaccines.