Clinical immunogenic profile of Heberon® alpha

Hugo Nodarse Cuni; Cimara Hortensia Bermúdez-Badell; Idrian García García; Iraldo Bello Rivero; Pedro López-Saura

Perfil de inmunogenicidad clínica del Heberon® alpha

ARTÍCULO ORIGINAL

Clinical immunogenicity profile for Heberon^® Alpha

Perfil de inmunogenicidad clínica del Heberon^® alpha

Hugo Nodarse-Cuní¹
Cimara Hortensia Bermúdez-Badell¹
Idrian Garcia Garcia¹
Iraldo Bello Rivero¹
Pedro López-Saura²

¹ Dirección de Investigaciones Clínicas del Centro de Ingeniería Genética y Biotecnología (CIGB). La Habana, Cuba.
² Instituto de Ciencias Básicas y Preclínicas Victoria de Girón. La Habana, Cuba.

ABSTRACT

Introduction: Interferons are a heterogeneous family of natural proteins described as agents capable of interfering with viral replication. The production of interferon as a drug began by recombinant DNA in microorganisms since the seventies of the 20^th century. Patients who receive interferon products, the result of genetic engineering, develop neutralizing antibodies for the biological activity of this protein and, therefore, are able to prevent or reverse treatment response.
Objectives: To analyze the immunogenicity profile of recombinant human interferon alfa-2b produced in the Center for Genetic Engineering and Biotechnology in Havana, Cuba.
Methods: The analysis of the immunogenicity profile of recombinant human interferon alfa-2b was performed in 952 patients from 23 clinical studies, performed with the lyophilized formulation with albumin. The experimental design was started with a "sandwich" type enzyme-linked immunosorbent assay (ELISA) for detecting the presence of the antibody and its confirmation of specificity (IFN alpha-2b/sample/conjugate protein A-peroxidase), and then we investigated, through a biological assay in cells, the ability of the antibody to neutralize the antiviral activity of interferon alpha.
Results: The development of antibodies with neutralizing capacity for the antiviral action occurred in 22 patients, representing 2.3 % of those evaluated. This percentage of immunogenicity of recombinant human interferon alfa-2b, produced by the Center for Genetic Engineering and Biotechnology, is below the reported 2.7 % for recombinant human interferon alfa-2b and 25.7 % for interferon alfa-2a in the international market.
Conclusions: Recombinant human interferon alfa-2b produced at the Center for Genetic Engineering and Biotechnology can be used as a safe drug for the treatment of all diseases included in its therapeutic indications.

Keywords: Immunogenicity; interferon alpha; anti-IFN antibodies.

RESUMEN

Introducción: Los interferones son una familia heterogénea de proteínas naturales descritas como agentes capaces de interferir con la replicación viral. El interferón como medicamento, comenzó su producción por la vía del ADN recombinante en microorganismos desde los años 70 del siglo XX. Los pacientes que reciben productos de interferón, fruto de la ingeniería genética, desarrollan anticuerpos neutralizantes de la actividad biológica de esta proteína y por tanto, son capaces de impedir o revertir la respuesta al tratamiento.
Objetivos: Analizar el perfil de inmunogenicidad del interferón alfa-2b humano recombinante producido en el Centro de Ingeniería Genética y Biotecnología de la Habana, Cuba.
Métodos: El análisis del perfil de inmunogenicidad del interferón alfa-2b humano recombinante fue realizado en 952 pacientes provenientes de 23 estudios clínicos, ejecutados con la formulación liofilizada con albúmina. El diseño experimental se inició con un ensayo inmunoenzimático (ELISA) tipo "sandwich" para detectar la presencia del anticuerpo y su confirmación de especificidad (IFN alfa-2b/muestra/ conjugado proteína A-peroxidasa), para luego investigar, a través de un ensayo biológico en células, la capacidad del anticuerpo para neutralizar la actividad antiviral del interferón alfa.
Resultados: El desarrollo de anticuerpos con capacidad neutralizante de la acción antiviral ocurrió en 22 pacientes, lo que representa el 2,3 % de los evaluados. Este porcentaje de inmunogenicidad del interferón alfa-2b humano recombinante, producido por el Centro de Ingeniería Genética y Biotecnología, se encuentra por debajo del 2,7 % reportado para el interferón alfa-2b humano recombinante y del 25,7 % para el interferón alfa-2a del mercado internacional.
Conclusiones: El interferón alfa-2b humano recombinante producido en el Centro de Ingeniería Genética y Biotecnología, puede ser empleado como un fármaco seguro para el tratamiento de todas las enfermedades comprendidas en sus indicaciones terapéuticas.

Palabras clave: Immunogenicidad; interferón alfa; anticuerpos anti-IFN.

INTRODUCTION

Interferons (IFN) are a family of natural heterogeneous proteins described as agents capable of interfering with viral replication. They are present in all animal species and their multifunctional influence allows various cellular processes, either as a component of the immune system during defensive response to viruses,¹ parasites² and certain tumors,³ or by proliferation inhibition or modulation of cellular differentiation.⁴

Interferon as medicine, began from the 70s of the XX century their production by means of recombinant DNA in microorganisms. This recombinant molecule, despite being human, has an unglycosylated structure which differs slightly from natural counterparts⁵ and is presented with structural differences between different pharmaceutical preparations which exist with alpha IFN as active principle. For these reasons and contrary to what happens with the native structure, patients receiving IFN products made by genetic engineering develop neutralizing antibodies that block biological activity of this protein and therefore prevent or reverse the response to treatment.⁶ It is also possible to detect antibodies against IFN which were not induced by treatment; in this case are known as auto-antibodies and it is recognized that although their frequency is sporadic, they may also have negative consequences on treatment⁷ or pathogenic implications.⁸

In 1981 came the first report of anti- interferon antibodies in a patient with nasopharyngeal carcinoma treated with beta IFN.⁹ Later it became known that prolonged exposure to doses of alpha IFN induces the formation of antibodies with possible neutralizing capacity.¹⁰ Experts have long recommended studying the influence on the phenomenon of immunogenicity that can exert various factors such as differences in antigenicity among commercial preparations of alpha IFN, the patient's underlying disease, the dosage administered, the route of administration and the treatment schedule.¹¹

The methodology to be used in studies of immunogenicity is one of the key issues and the one recommended by World Health Organization (WHO) is based on the neutralization of antiviral activity.However, this method is expensive and methodologically complex, that´s why it has resorted to immunochemical methods that offer the possibility to evaluate a large number of samples with a simple, rapid implementation, low cost, high specificity and sensitivity.¹²

The Center for Genetic Engineering and Biotechnology (CIGB) in Havana, Cuba produces since 1986 the recombinant human alpha-2b IFN, marketed as Heberon^® alpha by the Heber Biotec SA Company. The CIGB developed and validated a simple immunoenzymatic assay for detection of alpha anti-IFN antibodies by "sandwich" type (ELISA) and the verification of the specificity there of (alpha-2b IFN/sample/protein A-peroxidase conjugate).¹³ A bioassay was also implemented in cells, to assess the ability of antibodies to neutralize the antiviral activity of alpha interferon.¹⁴

METHODS

For evaluation and characterization of the profile of immunogenicity of recombinant human alpha-2b IFN (Heberon^® alpha) in its lyophilized variant with albumin, was monitored the use of the product in different medical indications for 952 patients, divided in 872 adults from 21 clinical trials and 80 children included in two trials. Active monitoring was extended for 24 years (1986-2010) and this investigation contains information about alpha anti-IFN antibody associated with the application of the product by intramuscular, subcutaneous, intralesional and intrathecal routes.

The database was generated from exploratory and confirmatory clinical investigations, phase IV studies of national extension, as well as reports of treated cases. The table 1 shows a distribution of patients evaluated during the therapeutic schemes with Heberon^® alpha covering between 3 and 60 consecutive months of treatment. Product doses administered ranged between 3 and 10 million IU (MUI) daily or three times a week. All clinical investigations referred to in this article were among its objectives the development and neutralizing activity of the alpha anti-IFN in subjects treated.

The immunogenicity study, based on the detection of anti-IFN alpha antibodies was performed in serum samples of patients, obtained from blood extracted and processed by centrifugation for 5 minutes at 250 × g, according to the current standard procedure in the hospital clinical laboratory. Were required 2 mL of serum collected into labeled Eppendorf tubes and preserved in a temperature no greater than -20 °C initially in the hospital and subsequently translocated to the CIGB for stored until later analysis.

Sera were analyzed before starting treatment for the presence of auto-antibodies and in a quarterly frequency to assess the induction of antibodies after started receiving the IFN. In line with WHO recommendations, all blood drawings were taken 48 hours after administration of Heberon^® alpha, to prevent any exogenous alpha IFN that could mask the determination by neutralization of antibodies present.

The immunoenzymatic assay type ELISA from CIGB used as a positive control one serum of a patient with known high titers of IFN neutralizing antibodies while serum from healthy volunteers represents the negative control. In the experimental design ninety-six well polystyrene plates (Costar, Cambridge, MA, USA) were coated with alpha IFN, washed with 0.05 % polysorbate 20 (Sigma, USA) and incubated with the samples or controls (100 μL/well) for 2 h 37 °C. The samples were diluted in 2 % skim milk (Oxoid, Hampshire, England) in PBS, pH 7.2-7.4 (assay buffer). After 4 washings, a protein A -peroxidase conjugate (Immunotechnology and Diagnosis Division, CIGB) was added (100 µL/well) and incubated for 30 min at 37 °C. After 6 washings the reaction was developed for 15 min at room temperature with 5.5 mg of ο-phenylendiamine (Merck, Germany) and 5.5 mL of 0.015 % H₂O₂ (Caledon, Canada) in 11 mL of citrate-phosphate pH 5.0 (100 μL/well). Finally, the reaction was stopped with 50 μL/well of 2 M H₂SO₄. The absorption was read at 492 nm in a plate reader (PR 521, Tecnosuma, Havana) and sera with extinction values higher than 3 standard deviations above the average of the negative controls in each plate (cut-off) were considered to be potential antibody binding and then analyzed in the confirmatory test for specificity through pre-incubated at a 1:20 dilution with 100 μg/mL of IFNα-2b for 1 h at 37 ºC and were confirmed to be positive when the fall of the absorption value after this pre-incubation was higher than the absolute cut-off value.

The subsequent step of evaluating the neutralizing capacity of the antiviral activity of IFN was performed as recommended by the International Society for Research on IFN and WHO,¹⁵ where the effects was measured by the inhibition of cytopathogenic effect for Mengo virus on Hep-2 cells (human laryngeal carcinoma)¹⁶ and using an IFN preparation calibrated in antiviral activity against the WHO international standard Gxa 01-901-305.¹⁷ Were evaluated sera samples classified as positive for the detection and confirmation of binding antibody alpha anti-IFN and considered as neutralizing those serum capable of reducing by 50 % antiviral protection of cells, which is considered equivalent to reduce the activity of alpha IFN from 10 IU to 1 IU.

RESULTS

The administration of 3 MUI of Heberon^® alpha, three times a week (tpw), was the therapeutic regimen with the highest number (445.47 %) of patients evaluated, followed by 10 MUI three tpw (133.14 %) and 3 MUI two tpw (109.11 %). Seven hundred fifty-one (79 %) of the evaluated patients were treated for 48 or more weeks. There were 383 patients (40 %) who received ≥10 MUI as total doses weekly; while others 215 (23 %) received 30 MUI (See table I).

Table 1 shows the results of immunogenicity accumulated during 24 years of clinical trials with the IFN alpha-2b produced by CIGB. Into the population analyzed and representing the main therapeutic indications of the product, there are 575 (60 %) patients come from hepatology, hematology and dermatology uses of Heberon^® alpha. In total this report includes information from 952 patients.

The Heberon^® alpha administration 2 or 3 tpw for ≥ 48 weeks were the frequencies of administration related with the appearance of antibodies. In 16 different diseases and 930 patients, the treatment was satisfactorily completed without incidents in immunogenicity against the product. The detection of alpha anti-IFN antibodies occurred with positive neutralizing antiviral activity in 22 patients from five diseases, representing 2.3 % of population evaluated. This percentage obtained for recombinant human alpha-2b IFN from CIGB is lower than 2.7 % reporting for the other alpha-2b IFN and also inferior than 25.7 % described for the alpha-2a IFN in the international market.¹⁸

Both competing products on the market are also more immunogenic than the Heberon^® alpha produced at CIGB in Cuba for patient populations from other parts of the world with similar diseases to those studied in this report (Recurrent Respiratory Papillomatosis: 5.3 % and 73.7 % vs. 5.3 %, Hepatitis C: 3.0 % and 32.5 % vs. 1.4 %).^19-22 It cannot be concluded any association between the occurrence of neutralizing anti-IFN activity and any of the listed diseases, because the samples were not taken at random. From the clinical viewpoint, an interesting finding was that all patients with neutralizing titers of alpha anti-IFN were non-responders to treatment. Despite this complete coincidence and although this work does not show the data on the antibody binding, there are evidences that non-response to treatment is not an exclusive outcome of patients with neutralizing antibodies; for example, there were 23 patients with chronic hepatitis C without response to alpha IFN treatment and with non-neutralizing anti-IFN alpha antibodies detected.

DISCUSSION

The analysis of alpha interferon immunogenicity and its formulations is complex and requires continuous research since many questions on the causes and factors involved are not yet clarified. It is not easy to perform controlled studies in humans with this purpose. The data presented in this paper are supported by 24 years of clinical observation in 952 individuals, including children and adults with use of different formulations of recombinant human alpha-2b IFN applied by various routes of administration in 21 recognized medical indications for this product.

Immunogenicity screening is usually performed using immunoassays which are based on a variety of formats and detection systems. All screening procedures detect antigen-antibody interaction (binding) but may differ in their underlying scientific/ technical principles. Reports in the literature,^23,24 about the determinations of alpha anti-IFN antibodies by Enzyme-Linked Immunosorbent Assay (ELISA) has been characterized by using "sandwich" systems recognized as very useful to differences between negative and positive serum in a random group of samples. ELISA developed by the CIGB to measure antibody titers involved standardization, validation and comparison against the technique recommended by WHO, process where no differences were found in the results compared against radioimmunoassay or bioassay and that made it possible to consider this ELISA a satisfactory option in terms of sensitivity, precision and high specificity.¹³

A strategy for immunogenicity assessment has established confirmatory assays as second step for eliminating any false positive results following the initial screening assay. A common approach for confirming antibodies is addition of an excess of antigen to the sample followed by a comparison of spiked and unspiked sample in the binding assay. This should result in a reduction of positive signal for true positives in the spiked sample; following this methodology, the absorbance values of the positive samples in this report were substantially reduced by preincubation with soluble IFN alpha-2b as proof of assay specificity.

The third step for distinguishing neutralizing from non-neutralizing antibodies was satisfactorily fulfilled because the samples confirmed as positive for binding antibodies were evaluated in the assay of neutralizing of antiviral activity (NAA) test recommended by WHO and the International Society for Research on interferon.¹⁵For the analysis, was used an universally employed, quantitative standardized virus-induced cytopathic-effect assay based on IFN induction of antiviral resistance, which is considered to represent the reference standard for measurement of NAA²⁵ and where antibody titers were determined by a neutralization test against 10 IU/mL of IFN with using the Kawade method and expressed as ten-fold reduction units (TRU/mL), that is, the dilution of serum that reduces 10 IU/mL of IFN to 1 IU/mL.²⁶

The state of art suggests that the type of the disease affects the generation of antibodies; the higher incidence has been reported in oncological²⁷ and viral²⁸ etiologies. The results of this study confirm the above approach, since they were found anti-interferon antibodies positive in patients infected with the papilloma and hepatitis C virus, as well as in cancer treatment for hemangioma of childhood, lung cancer and mycosis fungoid. These five diseases have in common the use of alpha IFN in prolonged treatment regimens, in which the higher dose and frequent the administration are factors that increase the possibility of developing antibodies.²⁹ This study confirms the occurrence of this potential risk, because immunogenicity was found positive in patients receiving 3 or 2 tpw for 48 or more weeks of IFN treatment. However, it should be considered low the incidence of this phenomenon for Heberon^® alpha since there were 447 patients free of antibodies in the same five diseases.

Many other factors related to the patient could influence the immunogenicity, one of them is related to the presence in blood of residual therapeutic product can establish complex with induced antibody and hence reduce the amount of antibody detectable by assays.³⁰ In this case the problem can be overcome by appropriate spacing of the timing between administration of product and sampling for antibody assessment i.e. allowing time for the product to be cleared from the circulation before sampling; this solution was implemented in this study by sampling at 48 hours after administration of Heberon^® alpha.

The direct relationship between the response to treatment and antibody formation has not been clearly verified. Nevertheless, the determination of neutralizing antibodies is of a high practical importance since they could be a cause of the lack of response to the treatment with this molecule and they could also trigger autoimmune processes. Certain authors have not found clinical implications due to the generation of these antibodies, it is possible that, in these cases, the affinity of the IFN receptors for the IFN was higher than the antibodies present in serum.³¹ Alternatively, patients could have responded before developing the neutralizing antibodies.³² None of the patients with neutralizing antibodies in this study had response to treatment with alpha IFN. Although the number of occurred events is very small and does not allow any statistical analysis, the coincidence is clinically interesting, especially by the association that arises between their presence and no therapeutic response.³³

Product-related factors that influence the immunogenicity include the manufacturing process, formulation, and stability characteristics.³⁴ This article show a low frequency of occurrence of neutralizing alpha anti-IFN antibodies after using the lyophilized formulation with albumin produced at CIGB. This percentage is lower than the immunogenicity of the other reported formulations of alpha IFN in the international market. The comparison between different products of the same biological molecule has been controversial because it suggests that the absence of equality in strains of origin and production processes there may be differences in the composition of products especially post-translational processing of the molecule and the impurities that may determine a greater or lesser degree of development of antibodies to exogenous IFN administered. While the latter is possible, it should not be the case of Heberon^® alpha from CIGB, because the molecule is well known, with well-defined and proven structure; it is not glycosylated, so posttranslational changes are minimal and verifiable by physic-chemical characterization and the production of recombinant human alpha-2b IFN, expressed in bacteria (Escherichia coli) yields a product considered as high purity.

Another issue to discuss is the presence of human serum albumin (HSA) in the composition of lyophilized Heberon^® alpha formulation, because the formation of complexes between IFN alpha molecules and HSA could increase immunogenicity.³⁵ The previous information from animal models with comparison of four formulations of human recombinant IFN alpha-2b showed that neutralizing antibodies titers were significantly higher in those lyophilized with HSA. Likewise, both elements seem to have influence in the formation of antibodies in mice and increase the immunogenicity of alpha IFN by small proportions of HSA-IFN aggregates that may form during storage of the product.^36,37 Due to ethical and practical reasons it is difficult to carry out such studies in humans under controlled conditions and although the results cannot be extrapolated to patients receiving alpha IFN, they can be taken as indicators.

Heberon^® alpha is presented by Center for Genetic Engineering and Biotechnology as a lyophilized powder with 1.5 mg/mL of HAS in its composition, while INTRON A (IFN alfa-2b from Shering-Plough) is also lyophilized with 1.0 mg/mL of HAS. Considering the lower percentage of immunogenicity obtained for Cuban alpha IFN and that in the studied population there was a large group patient with unfavorable treatment and demonstrated negative neutralizing antibodies, it was concluded that highest albumin content exerts no influence contrary to immunological product safety. For the specific case of alpha anti-IFN antibodies even when there is no data corroborating clinically, the lack of albumin in the formulations is considered an advantage that reduces the risk of its occurrence.³⁸ In line with the above, Heberon^® alpha was much less immunogenic than Roferon A (IFN alfa-2a from Roche) which it is a lyophilized formulation with 5.0 mg/mL of HAS.

This study contributes to the characterization of the biological properties of Cuban recombinant IFN, because evaluate the inmunogenicity of Heberon^® alpha in several of its most important clinic uses. The results of this study are consistent with previous reports of other authors.^39,40

As conclusion, considering the large number of clinical studies and subjects of this study and the low percentage of patients with reports of immunogenicity with the ability to block the antiviral activity, it can be stated that alpha-2b interferon produced in CIGB has a rate lower immunogenicity compared to their similar and can be used as a safe drug for the treatment of all diseases included in their therapeutic indications.

ACKNOWLEDGEMENTS

The authors thank Heber Biotec S.A. for the supply of the investigational product. Authors are employees of the Centre for Biological Research, which is part of the Center for Genetic Engineering and Biotechnology (CIGB), Havana where Heberon^® alpha is produced. None authors have conflict of interests.

CONFLICTO DE INTERESES

Los autores declaran que no presentan conflicto de intereses.

REFERENCES BIBLIOGRAPHICAL

1. Hastings AK, Erickson JJ, Schuster JE, Boyd KL, Tollefson SJ, Johnson M, et al. Role of type I interferon signaling in human metapneumovirus pathogenesis and control of viral replication. J Virol. 2015 Apr;89(8):4405-20.

2. Troegeler A, Mercier I, Cougoule C, Pietretti D, Colom A, Duval C, et al. C-type lectin receptor DCIR modulates immunity to tuberculosis by sustaining type I interferon signaling in dendritic cells. Proc Natl Acad Sci U S A. 2017;114(4):E540-E549.

3. Wonganan O, He YJ, Shen XF, Wongkrajang K, Suksamrarn A, Zhang GL, et al. 6-Hydroxy-3-O-methyl-kaempferol 6-O-glucopyranoside potentiates the anti-proliferative effect of interferon α/β by promoting activation of the JAK/STAT signaling by inhibiting SOCS3 in hepatocellular carcinoma cells. Toxicol Appl Pharmacol. 2017;336:31-9.

4. Pylaeva E, Lang S, Jablonska J. The Essential Role of Type I Interferons in Differentiation and Activation of Tumor-Associated Neutrophils. Front Immunol. 2016;7: 629.

5. Kuriakose A, Chirmule N, Nair P. Immunogenicity of Biotherapeutics: Causes and Association with Posttranslational Modifications. J Immunol Res. 2016;2016:1298-473.

6. Scagnolari C, Trombetti S, Soldà A, Milella M, Gaeta GB, Angarano G, et al. Development and specificities of anti-interferon neutralizing antibodies in patients with chronic hepatitis C treated with pegylated interferon-α. Clin Microbiol Infect. 2012;18(10):1033-9.

7. Xue L, Clements-Egan A, Amaravadi L, Birchler M, Gorovits B, Liang M, et al. Recommendations for the Assessment and Management of Pre-existing Drug-Reactive Antibodies During Biotherapeutic Development. AAPS J. 2017;19(6):1576-86.

8. Gupta S, Tatouli IP, Rosen LB, Hasni S, Alevizos I, Manna ZG, et al. Distinct Functions of Autoantibodies Against Interferon in Systemic Lupus Erythematosus: A Comprehensive Analysis of Anticytokine Autoantibodies in Common Rheumatic Diseases. Arthritis Rheumatol. 2016;68(7):1677-87.

9. Vallbracht A, Treuner J, Flehmig B, Joester KE, Niethammer D. Interferon-neutralizing antibodies in a patient treated with human fibroblast interferon. Nature. 1981;289(5797):496-7.

10. Pineda C, Castañeda Hernández G, Jacobs IA, Alvarez DF, Carini C. Assessing the Immunogenicity of Biopharmaceuticals. BioDrugs. 2016;30(3):195-206.

11. Wadhwa M, Knezevic I, Kang HN, Thorpe R. Immunogenicity assessment of biotherapeutic products: An overview of assays and their utility. Biologicals. 2015;43(5):298-306.

12. Richards S, Amaravadi L, Pillutla R, Birnboeck H, Torri A, Cowan KJ, et al. 2016 White Paper on recent issues in bioanalysis: focus on biomarker assay validation (BAV): (Part 3 - LBA, biomarkers and immunogenicity). Bioanalysis. 2016;8(23):2475-96.

13. González-Cabañas R, Ferrero-Bibilonia J, Morales-Menéndez MG, Aguilera-Rodríguez A, López-Saura P. Antigenicidad del interferón alfa-2b recombinante (Heberon alfa R). Detección de anticuerpos mediante un ensayo inmunoenzimático y neutralización de actividad antiviral. Biotecnol Apl. 1998;15(2):71-6.

14. Ferrero J, Ochagavia ME Aguilera-Rodríguez A, López-Saura P. Titulación de la actividad antiviral de interferon utilizando el sistema de equipo SUMA. Biotecnol Apl. 1994;11:34-42.

15. Grossberg SE. Human antibodies to interferons. Report of a National Institutes of Health Workshop held on 26-26 July 1988 in Bethesda, MD. J interferon Res. 1988;8(6):6-7.

16. Herrera L, De la Fuente J, Quiñones Y, Padrón G, Agraz A, Vega J, et al. Biological production by recombinant DNA technology in Cuba. Dev Biol Stand. 1989;70:257-69.

17. National Institute of Alergy and Infectious Disease. Research Reference Reagent note No 31. Freeze-died Reference Human Recombinant alpha-2 interferon. Catalog number Gxa 01-901-535. National Institute Of Health Bethesda, Maryland. 1984;1-5.

18. Nodarse-Cuni H, Lopez PA. Cuban interferon alpha-2b. Thirty years as an effective and safe drug. Biotecnol Apl. 2017;34:1211-7.

19. Nurmukhametov RKh, Onufrieva EK, Soldatskiĭ IuL, Mezentseva MV, Kasianova NV, Tsvetnova MV, et al. Assessment of the formation of interferon neutralizing antibodies and their influence on the effectiveness of interferon therapy in children with juvenile respiratory papillomatosis. Vestn Otorinolaringol. 2000;(4):22-5.

20. Nodarse-Cuni H, Iznaga N, Viera D, Rodriguez H, Fernández H, Blanco Y, et al. Interferon alpha-2b as adjuvant treatment of recurrent respiratory papillomatosis in Cuba: National Programme (1994-1999 report). J Laryngol Otol. 2004;118(9):681-7.

21. Milella M, Antonelli G, Santantonio T, Currenti M, Monno L, Mariano N, et al. Neutralizing antibodies to recombinant alpha-interferon and response to therapy in chronic hepatitis C virus infection. Liver. 1993;13(3):146-50.

22. Bordens R, Grossberg SE, Trotta PP, Nagabhushan TL. Molecular and biologic characterization of recombinant interferon-alpha2b. Semin Oncol. 1997;24(3 Suppl 9):S9-41-S9-51.

23. Partridge MA, Purushothama S, Elango C, Lu Y. Emerging Technologies and Generic Assays for the Detection of Anti-Drug Antibodies. J Immunol Res. 2016;1-6.

24. Ingenhoven K, Kramer D, Jensen PE, Hermanrud C, Ryner M, Deisenhammer F, et al. Development and Validation of an Enzyme-Linked Immunosorbent Assay for the Detection of Binding Anti-Drug Antibodies against Interferon Beta. Front Neurol. 2017;8:305.

25. Grossberg SE, Kawade Y. The expression of potency of neutralizing antibodies for interferons and other cytokines. Biotherapy 1997;10:93-8.

26. Kawade Y. Quantitation of neutralization of interferon by antibody. Methods Enzymol. 1986;119:558-73.

27. Kintzing JR, Filsinger Interrante MV, Cochran JR. Emerging Strategies for Developing Next-Generation Protein Therapeutics for Cancer Treatment. Trends Pharmacol Sci. 2016;37(12):993-1008.

28. Matsuda F, Torii Y, Enomoto H, Kuga C, Aizawa N, Iwata Y, et al. Anti-interferon-α neutralizing antibody is associated with nonresponse to pegylated interferon-α plus ribavirin in chronic hepatitis C. J Viral Hepat. 2012;19(10):694-703.

29. Schellekens H, Casadevall N. Immunogenicity of recombinant human proteins: causes and consequences. J Neurol. 2004;251 Suppl 2:II4-9.

30. Ratanji KD, Derrick JP, Dearman RJ, Kimber I. Immunogenicity of therapeutic proteins: influence of aggregation. J Immunotoxicol. 2014;11(2):99-109.

31. Zhong ZD, Clements-Egan A, Gorovits B, Maia M, Sumner G, Theobald V, et al. Drug Target Interference in Immunogenicity Assays: Recommendations and Mitigation Strategies. AAPS J. 2017;19(6):1564-75.

32. Schellekens H, Casadevall N. Immunogenicity of biopharmaceuticals. The European perspective. Dev Biol (Basel). 2003;112:23-8.

33. Enomoto H, Nishiguchi S. Factors associated with the response to interferon-based antiviral therapies for chronic hepatitis C. World J Hepatol. 2015;7(26):2681-7.

34. van Beers MM, Bardor M. Minimizing immunogenicity of biopharmaceuticals by controlling critical quality attributes of proteins. Biotechnol J. 2012;7(12):1473-84.

35. Obrezanova O, Arnell A, de la Cuesta RG, Berthelot ME, Gallagher TR, Zurdo J. Aggregation risk prediction for antibodies and its application to biotherapeutic development. MAbs. 2015;7(2):352-63.

36. García García I, Prats Pérez PA, Cervantes Llanos M, Valenzuela Silva C, López-Saura P. Factors that Influence on the Immunogenicity of Human Recombinant Interferon Alpha-2b in mice. Biotecnol Apl. 2002;19(1-2):15-8.

37. Garcia I, Bobillo H, Soria Y, Cervantes M, Bermúdez C, Díaz G, et al. Influence of albumin content and freeze-drying on the immunogenicity of human recombinant Interferon alpha-2b in mice. Biotecnol Apl. 2009;26(4):328-32.

38. Braun A, Kwee L, Labow MA, Alsenz J. Protein aggregates seem to play a key role among the parameters influencing the antigenicity of interferon alpha (IFN-alpha) in normal and transgenic mice. Pharm Res. 1997;14(10):1472-8.

39. Torres B, Miranda A, García I, Porreo J, Nodarse-Cuní H. Inmunogenicidad del interferon alfa. Revista Habanera de Ciencias Médicas. 2005;4(5):1-3.

40. Arus E, Infante M, Padron GJ, Morales MG, Grá B, Soto G, López P. Interferon alfa-2b recombinante en hepatitis C crónica: resultados del tratamiento y determinación de anticuerpos anti-IFN. Biotecnol Apl. 1997;14:242-47.

Recibido: 7 de marzo de 2018.
Aprobado: 7 de abril de 2018.

Hugo Nodarse-Cuni. Center for Genetic Engineering and Biotechnology, Clinical Trials Division. PO Box 6332, Havana, Cuba. Telephone: +53 7 2080428 ext. 107.
Email: hugo.nodarse@cigb.edu.cu

Enlaces refback

No hay ningún enlace refback.

Esta obra está bajo una licencia de Creative Commons Reconocimiento-NoComercial 4.0 Internacional.