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The Following is a Peer Reviewed Abstract About Cimavax
Pedro C Rodríguez, MD, Gryssell Rodríguez, MS, Gisela González, PhD, Agustín Lage, MD, PhD
Introduction CIMAvax EGF is a therapeutic anticancer vaccine developed entirely in Cuba and licensed in Cuba for use in adult patients with stage IIIB/IV non-small-cell lung cancer (NSCLC). The vaccine is based on active immunotherapy by which an individual’s immune response is manipulated to release its own effector antibodies (Abs) against the epidermal growth factor (EGF).
Objective Review pre-clinical and clinical research conducted during development of CIMAvax EGF, primarily studies published by Cuban investigators in international peer-reviewed scientific journals.
Methods An automated search for “vaccine” and “EGF” was conducted in PubMed, resulting in 17 articles published by Cuban authors between January 1, 1994 and September 30, 2009. Main findings were described and discussed, along with unpublished preliminary findings of an initial ongoing phase III clinical trial.
Results Articles reviewed describe five phase I/II and one phase II clinical trials conducted in Cuba in 1995–2005. A non-controlled 1995–1996 study resulted in the earliest published scientific evidence of the feasibility of inducing an immune response against autologous EGF in patients with different advanced stage tumors. Subsequent controlled, randomized trials included patients with advanced stage (IIIB/IV) NSCLC. The 2nd and 3rd phase I/II trials differentiated immunized patients as poor antibody responders (PAR) and good antibody responders (GAR), according to their anti-EGF antibody response, and confirmed greater immunogenicity with Montanide ISA 51 adjuvant in the vaccine formulation, as well as the benefits of low-dose cyclophosphamide treatment 72 hours before the first immunization. The 4th phase I/II trial found increased immunogenicity with an increased dose divided in 2 anatomical sites and also established correlation between Ab titers, serum EGF concentration and length of survival. In the first 4 phase I/II trials and the phase II trial, vaccine was administered after chemotherapy (ChTVV schedule). In the 5th phase I/II trial, longer survival and increased immunogenicity were achieved using a VChTV schedule and dividing the vaccine dose in 4 anatomical sites. The phase II clinical trial confirmed results of earlier studies as well as the mild-to-moderate adverse event profile associated with CIMAvax EGF. Longer survival was observed in all vaccinated patients compared to controls, and the difference was significant (p <0.05) in the group aged <60 years.
Conclusions CIMAvax EGF’s benefits in earlier NSCLC stages and in other tumor locations, as well as in patients unfit for chemotherapy, need to be evaluated. Evidence of the vaccine’s safety for chronic use also needs to be systemized.
Key words: Epidermal growth factor, EGF receptor, non-small-cell lung carcinoma, lung cancer, vaccine therapy, immunotherapy, cancer vaccines
In Cuba, cancer is the second leading cause of death and the primary cause of years of potential life lost, making a significant impact on life expectancy at birth. Lung cancer is the malignant disease with highest incidence and also the leading cause of cancer mortality in the country. In 2005–2007, an average of 4234 new lung cancer cases and 4601 deaths from the disease were reported annually, for a crude mortality rate of 54.3 men and 27.3 women per 100,000 population. If present demographic trends and risk factors persist, lung cancer incidence and mortality can be expected to rise significantly in Cuba in the next five years.
Recognizing rising cancer incidence and mortality as a major public health problem, the Cuban Ministry of Public Health has implemented a Comprehensive Cancer Control Program (PICC, its acronym in Spanish), operating across all levels of the national public health system. This program constitutes a new therapeutic approach to the disease with biotechnology serving as a bridge between basic immunology research and public health. In Cuba, biotechnological research and development is conceived as a complete scientific cycle (closed loop), from concept to clinical application and marketing of scientific products. Income from product sales is invested in both sustaining use of immunotherapies in the national public health system and in research and development of new immunotherapeutic modalities.[2,3]
Although chemotherapies have become an indispensable arsenal for reducing tumor burden and extending survival, their impact on lung cancer is measurable only in months with the added burden of severe adverse reactions. Lung cancer patients generally confront an initial stage of diagnosis and oncological treatment, during which complete or partial remission is achieved, followed by a second stage, during which the disease inexorably progresses toward terminal illness and death. The mechanism of action of immunotherapy products, such as monoclonal antibodies (mAbs) and therapeutic vaccines, is much more selective toward tumor cells and may increase cancer patient survival with improved quality of life.[3–5]
The Epidermal Growth Factor Receptor (EGFR) is a well-known oncogene. Its overactivation can induce malignant transformation of a normal cell, signaling inhibition of apoptosis, cell proliferation, angiogenesis, metastasis and tumor-induced proinflammatory and immunosuppressive processes. The EGFR signaling and transduction pathway can be efficiently interrupted by EGF deprivation, direct specific mAb receptor inhibition, or low molecular weight molecules competing intracellularly with adenosine triphospate (ATP) for the receptor’s tyrosine kinase activity site, with negative repercussions on cell proliferation and, consequently, on tumor development.[6,7]
Inducing EGF deprivation by active immunotherapy is an emerging concept developed by Cuban researchers which involves manipulating an individual’s immune response to release its own effector antibodies (Abs) against EGF, thereby reducing tumor size or preventing its progression.[8–13]
CIMAvax EGF is a therapeutic anticancer vaccine developed entirely in Cuba. Its active pharmaceutical ingredients (API) are produced by the Center for Genetic Engineering and Biotechnology (CIGB, its Spanish acronym); the vaccine is formulated at the Molecular Immunology Center (CIM, its Spanish acronym); and clinical trials are conducted in hospitals meeting professional and technological Good Clinical Practice standards.[14,15] Proof of principle (POP) of this novel therapy’s clinical impact was made possible by the integration of Cuban biotechnological development in the public health system and collaboration between research institutes and hospitals.
Since 1995, CIMAvax EGF has undergone five phase I/II and one phase II clinical trials. Results of these investigations led the Government Center for Quality Control of Medicines (CECMED, its Spanish acronym), the Cuban regulatory authority, to license this therapeutic vaccine for use in adult patients with stage IIIB/IV non-small-cell lung cancer (NSCLC). Research results have also been published in various national and international scientific journals.
The objective of this article is to review the pre-clinical and clinical research conducted during development of this novel Cuban therapeutic anticancer vaccine, based primarily on studies published by Cuban investigators in international peer-reviewed scientific journals.
An automated search of the PubMed database (Medline) through WHO’s HINARI service was made using the key words “vaccine” and “EGF”. General selection criteria were articles published by Cuban authors in peer-reviewed international journals. Seventeen articles, published between January 1, 1994 and September 30, 2009, were found and reviewed for their coverage of concepts, clinical data and essential technological aspects of CIMAvax EGF development and potential impact on NSCLC. The main findings in each article were described and discussed, along with unpublished preliminary findings of an initial ongoing phase III clinical trial.
RESULTS AND DISCUSSION
Vaccine Formulation and Clinical Induction of Immune Response against Autologous EGF
When an individual is immunized with autologous EGF (from its own species), no anti-EGF antibody (Ab) response occurs. To prove the feasibility of manipulating an individual’s immune response to induce an anti-EGF Ab response, reducing EGF concentration in blood and depriving the tumor of this growth factor, a vaccine formulation was needed that would make EGF recognizable to the immune system, that is, render it immunogenic. Therefore, selection of an adequate immunopotentiator and adjuvant was required.In 1992, preclinical studies were begun to select the most effective vaccine formulation for inducing autologous EGF immunogenicity. In this stage, components of the vaccine formulation were outlined, and the possibility of inducing effective autologous EGF immunogenicity was demonstrated. Different vaccine formulations were tried, and 2 immunopotentiating proteins were selected: tetanus toxoid (TT) and Neisseria meningitidis P64k (P64k), both produced in Cuba (Finlay Institute and CIGB, respectively). Two possible adjuvants were also proposed for clinical evaluation: aluminum hydroxide (Superfos, Denmark) and Montanide ISA 51 (Seppic, France).[13–14]
Clinical development of CIMAvax EGF began in 1995 with a phase I/II open-label clinical trial (Pilot 1) at the Medical-Surgical Research Center (CIMEQ, its Spanish acronym) in Havana. A non-randomized design was used in 10 patients with histologically proven primary malignant tumors in different locations, previously treated with first-line chemotherapy (ChTVV therapeutic schedule) (Table 1). The main objective was to evaluate the immunogenicity of the incipient vaccine formulation.
Results of this first clinical trial constituted the earliest published scientific evidence of the feasibility of inducing an immune response against autologous EGF in patients with different advanced stage tumors. Additionally, protein P64k was confirmed as the optimal immunopotentiator for EGF conjugation and the CIMAvax EGF formulation.
Selection of NSCLC Target Location and Advanced Development of Vaccine Formulation
A key factor in developing the final vaccine formulation was selection of the tumor location in which to introduce this novel therapeutic strategy. NSCLC was selected because of its frequency and because EGFR is overexpressed in tissues during development and progression of lung neoplasms in the following proportions: 62% of all NSCLC tumors, 89% of squamous cell tumors, 41% of adenocarcinomas and 80% of bronchoalveolar tumors. Magnitude of EGFR expression has been reported in the literature as a predictive factor of response to biological therapy in NSCLC patients.[6,8,11,16]
The next two phase I/II clinical trials (Pilot 2 and Pilot 3) were conducted jointly in overlapping periods (1997–1999 and 1998–2001, respectively) at CIMEQ and the National Oncology and Radiobiology Institute (INOR, its Spanish acronym), also in Havana (Table 1). Pilot 2 included 20 advanced stage NSCLC (IIIB/IV) patients; 10 were randomly immunized with EGF/P64k vaccine using aluminum hydroxide as adjuvant (EGF/P64k/AL), and 10 received EGF/P64k adjuvanted with Montanide ISA 51 (EGF/P64k/Montanide ISA 51).
The Pilot 3 study also included 20 advanced stage NSCLC (IIIB/IV) patients who received the same treatments used in Pilot 2 (EGF/P64k/AL and EGF/P64k/Montanide ISA 51) with a similar randomization in 2 groups, except they all received a cyclophosphamide dose (200 mg/m2 of body surface) 72 hours before vaccine treatment onset. Cyclophosphamide is a widely studied anticancer drug. Its immunomodulating effects have significant dose- and therapeutic schedule-related repercussions.[18,19] This pretreatment was introduced to disrupt immunologic tolerance to EGF and to induce immunogenicity toward this human molecule from the first dose.
The Pilot 2 and Pilot 3 clinical trials confirmed that vaccination induced a specific anti-EGF Ab response and enabled classification of immunized patients into 2 subpopulations: poor antibody responders (PAR)—those with poor anti-EGF Ab response—and good antibody responders (GAR)—those with an anti-EGF antibody response ≥1:4000 and at least 4 times (4x) their preimmunization levels. In both studies, survival was longest in the GAR group (mean, 12.41 months; median, 9.1 months), compared to mean, 5.47 months (median, 4.5 months), in PAR patients (Figure 1). The difference in survival was statistically significant (P <0.05). Six-month survival was achieved by 84% of GAR and 38% of PAR. Twelve-month survival was achieved by 37% of GAR and 4% of PAR. These studies also showed that patients with an anti-EGF Ab response ≥60 days survived longer than those whose response lasted <60 days.
Table 1: EGF Therapeutic Anticancer Vaccine Phase I/II Trials Conducted in Cuba, 1995–2005
Figure 1: Survival Functions for Good Antibody Responders (GAR) and Poor Antibody Responders (PAR) to EGF Vaccination in 2 Pilot Phase I/II Clinical Trials (pooled data).
Results of the Pilot 2 and Pilot 3 phase I/II clinical trials confirmed greater immunogenicity with Montanide ISA 51 adjuvant (EGF/P64k/Montanide ISA 51) in the vaccine formulation, as well as the benefits of low-dose cyclophosphamide treatment 72 hours before the first immunization.[17,20]
A 4th phase I/II clinical trial (Pilot 4), conducted at the Hermanos Ameijeiras Hospital (HAH) in Havana in 2000–2003, evaluated 2 dose levels of the therapeutic vaccine. Forty-three patients with advanced stage (IIIB/IV) NSCLC were randomized in 2 groups and received 71 µg or 142 µg of EGF (Table 1). The lower dose was applied in one deltoid region and the higher dose distributed between the 2 deltoid regions.
The Pilot 4 study established a correlation between vaccine dose, anti-EGF Ab titers, EGF serum concentrations, and patient survival. Survival in treated patients (mean, 9.83 months; median, 8 months) significantly exceeded (p <0.05) the historical control (mean, 6.2 months; median, 4.1 months) while positively correlating with GAR titers (≥1/4000) and lower serum EGF concentrations (<168 pg/ml). This clinical trial demonstrated for the first time that EGF serum concentration levels decrease when Ab titers rise.
Return to Preclinical Research: Manipulating Immunopharmacological Variables and Fine Tuning the Therapeutic Schedule
The immunopharmacology of cancer vaccines is not yet fully understood, and there is scant data on immunopharmacological determinants influencing therapeutic anticancer vaccines.[22–28] Many variables must be evaluated, such as therapeutic schedules, administration route, dose, dosing interval, and optimal combination with already established therapies, among others. This process is even more complex when attempting to fine tune the therapy schedule of a vaccine based on a circulating self-molecule, such as autologous EGF, and induce an Ab response that suppresses it from circulation, thus depriving its receptor.
For CIMAvax EGF vaccine, more efficient strategies for inducing Ab response (priming) were needed, as well as for long-term response maintenance through reimmunization (boosting).[25–27] Investigators therefore returned to preclinical research using murine biomodels, defined as GAR (BALB/c mice) or PAR (C57BL6 mice), depending on their genetic background and antibody response when challenged with the vaccine. The animals were immunized with CIMAvax EGF vaccine (EGF/P64k/Montanide ISA 51), and some immunopharmacological variables (dose, number of immunizations, dosing interval) were manipulated during both the response induction phase (priming) and the reinforcement or reimmunization (boosting) phase, aimed at inducing an early, robust and prolonged anti-EGF Ab response, potentiating its active immune deprivation.
For priming, fractioning an apparently low dose (4 µg) into 4 parts administered by intramuscular injection in different anatomical sites increased maximal Ab titer levels and extended the duration of vaccine response. Shortening the interval between booster doses reduced persistence of anti-EGF Ab titers, whereas repeated boosting converted PAR status to GAR.
Results of this study led to the conclusion that the vaccine should be administered in a high but fractioned dose in multiple anatomical sites (such as the 2 deltoid and 2 gluteal regions), thereby bringing the EGF vaccine closer to regional lymph nodes and synergizing the immune response.
Return to Clinical Trials: Combining Therapeutic Vaccination (V) and Chemotherapy (ChT)
Tumor biology is the result of cell genome interaction with environment; in addition to mutual influence, this interaction has repercussions in both reconfiguration of cell metabolism and evolutionary selection of more efficient tumor mechanisms. The immune system is one of the critical elements involved in shaping cell phenotype.[30–34]
The complexity of mechanisms involved in malignant transformation of cells lends itself to a combination therapy approach aimed at, among other things, simultaneously controlling tumor immunoevasion and dissemination. In this context, immunotherapies emerge as an approach that ensures greater specificity and limited associated toxicity. Immunotherapies have been included in standard cytotoxic therapeutic regimens since they were first developed. Passive mAb immunotherapy, in particular, has confirmed the potential of these combinations.[35–39] In this context, it was hypothesized that potentially autoreactive lymphocyte clones, capable of generating an autologous anti-EGF antibody response, reemerge from chemotherapy-induced maximum lymphocyte depletion (lymphopenia nadir), amplified with homeostatic advantage.[40,41]
Based on these concepts and preclinical research results, a therapeutic schedule combining vaccine, chemotherapy and more vaccine (VChTV) was proposed. This schedule was evaluated in the 5th phase I/II clinical trial (Pilot 5) begun in 2001 in the Hermanos Ameijeiras Hospital (HAH) in Havana, using the CIMAvax EGF (EGF/P64k/Montanide ISA 51) formulation in 20 stage IIIB/IV NSCLC patients. This study corroborated the correlation between an increase in anti-EGF Ab titers and a decrease in serum EGF concentration. Correlation between antibody titers and EGF/EGFR binding inhibition capacity was also shown, and GAR patient sera immunodominance against loop B of the EGF molecule was established.
Another important finding in the Pilot 5 clinical trial was the differentiation of a new subpopulation of patients with anti-EGF antibody titers ≥1:64000, defined as super good antibody responders (sGAR). A correlation between Ab response and survival was also observed; sGAR patients survived significantly longer than patients classified as only GAR (Figure 2). This clinical finding tends to confirm the hypothesis that potentially autoreactive clones reemerge and amplify, possibly favoring active immunization with the VChTV therapeutic schedule.