biopreparatБИОпрепараты. Профилактика, диагностика, лечениеBiological Products. Prevention, Diagnosis, Treatment2221-996X2619-1156Scientific Centre for Expert Evaluation of Medicinal Products10.30895/2221-996X-2022-22-4-414-434biopreparat-437Research ArticleСтатьиДоклинические исследования противовирусной активности гибридного белка RPH-137 и молнупиравира в отношении COVID-19Preclinical studies of antiviral activity of the RPH-137 fusion protein and molnupiravir against COVID-19https://orcid.org/0000-0002-5953-984XШипаеваЕ. В.ShipaevaE. V.

Шипаева Елена Владимировна,  канд. мед. наук. 

Ленинский проспект, д. 111/1, Москва, 119421

Elena V. Shipaeva, Cand. Sci. (Med.)

111/1 Leninsky Ave, Moscow 119421

shipaeva@rpharm.ruhttps://orcid.org/0000-0002-8735-7429ФилонО. В.FilonO. V.

Филон Ольга Владимировна

Ленинский проспект, д. 111/1, Москва, 119421

Olga V. Filon

111/1 Leninsky Ave, Moscow 119421

ov.filon@rpharm.ruhttps://orcid.org/0000-0001-8148-5086ЗинченкоА. В.ZintchenkoA. V.

Зинченко Аркадий Владимирович,  Ph. D. 

Ленинский проспект, д. 111/1, Москва, 119421

Arkadi V. Zintchenko, Ph. D. 

111/1 Leninsky Ave, Moscow 119421

a.zinchenko@rpharm.ruhttps://orcid.org/0000-0003-4495-7050ШипуновГ. А.ShipunovG. A.

Шипунов Георгий Александрович

Ленинский проспект, д. 111/1, Москва, 119421

Georgy A. Shipunov

111/1 Leninsky Ave, Moscow 119421

shipunov@rpharm.ruhttps://orcid.org/0000-0003-4743-6189ДмитриеваА. А.DmitrievaA. A.

Дмитриева Анастасия Андреевна

Ленинский проспект, д. 111/1, Москва, 119421

Anastasia A. Dmitrieva

111/1 Leninsky Ave, Moscow 119421

aa.dmitrieva@rpharm.ruhttps://orcid.org/0000-0003-4793-7477ЛемакМ. С.LemakM. S.

Лемак Мария Степановна,  канд. биол. наук. 

Ленинский проспект, д. 111/1, Москва, 119421

Maria S. Lemak, Cand. Sci. (Biol.)

111/1 Leninsky Ave, Moscow 119421

lemak@rpharm.ruhttps://orcid.org/0000-0001-7299-660XГришинС. А.GrishinS. A.

Гришин Сергей Александрович, канд. мед. наук. 

Ленинский проспект, д. 111/1, Москва, 119421

Sergey A. Grishin, Cand. Sci. (Med.)

111/1 Leninsky Ave, Moscow 119421

sa.grishin@rpharm.ruhttps://orcid.org/0000-0003-3246-6457ТрофимецЕ. И.TrofimetsE. I.

Трофимец Екатерина Игоревна

Заводская ул., д. 3-245, Ленинградская обл., Всеволожский р-н, п. Кузьмоловский, 188663

Ekaterina I. Trofimets

3/245 Zavodskaya St., Kuzmolovsky urban-type settlement, Vsevolozhsky district, Leningrad region 188663

trofimets.ei@doclinika.ruhttps://orcid.org/0000-0003-1451-7716КрышеньК. Л.KryshenK. L.

Крышень Кирилл Леонидович,  канд. биол. наук. 

Заводская ул., д. 3-245, Ленинградская обл., Всеволожский р-н, п. Кузьмоловский, 188663

Kirill L. Kryshen, Cand. Sci. (Biol.)

3/245 Zavodskaya St., Kuzmolovsky urban-type settlement, Vsevolozhsky district, Leningrad region 188663

kryshen.kl@doclinika.ruhttps://orcid.org/0000-0002-3029-1035КозловскаяЛ. И.KozlovskayaL. I.

Козловская Любовь Игоревна,  канд. биол. наук. 

Поселение Московский, поселок Института полиомиелита, вл. 8, к. 1, Москва, 108819

Liubov I. Kozlovskaya, Cand. Sci. (Biol.)

8/1 Village of the Institute of Poliomyelitis, Moskovsky settlement, Moscow 108819

kozlovskaya_li@chumakovs.suhttps://orcid.org/0000-0003-4624-2593ЛунинА. С.LuninA. S.

Лунин Александр Сергеевич

Поселение Московский, поселок Института полиомиелита, вл. 8, к. 1, Москва, 108819

Aleksandr S. Lunin

8/1 Village of the Institute of Poliomyelitis, Moskovsky settlement, Moscow 108819

lunin_as@chumakovs.suhttps://orcid.org/0000-0002-9978-222XАполоховВ. Д.ApolokhovV. D.

Аполохов Василий Даниилович

Поселение Московский, поселок Института полиомиелита, вл. 8, к. 1, Москва, 108819

Vasiliy D. Apolokhov

8/1 Village of the Institute of Poliomyelitis, Moskovsky settlement, Moscow 108819

apolohov_vd@chumakovs.suБарбашовС. Ф.BarbashovS. F.

Барбашов Сергей Федорович, кандидат биологических наук

ул. Кост-Бульвар-Саус, д. 505, офис 102, Ла-Хойя, г.Сан-Диего, штат Калифорния, 92037

Sergey F. Barbashov, Cand. Sci. (Biol.)

505 Coast Boulevard South, Suite 102, La Jolla, CA 92037

barbashov@r-pharm.comЛавровскийЯ. В.LavrovskyYa. V.

Лавровский Ян Вадимович, кандидат биологических наук

ул. Кост-Бульвар-Саус, д. 505, офис 102, Ла-Хойя, г.Сан-Диего, штат Калифорния, 92037

Yan V. Lavrovsky, Cand. Sci. (Biol.)

505 Coast Boulevard South, Suite 102, La Jolla, CA 92037

lavrovsky@r-pharm.comhttps://orcid.org/0000-0001-7721-5059СамсоновМ. Ю.SamsonovM. Yu.

Самсонов Михаил Юрьевич, кандидат медицинских наук

Ленинский проспект, д. 111/1, Москва, 119421

Mikhail Yu. Samsonov, Cand. Sci. (Med.)

111/1 Leninsky Ave, Moscow 119421

samsonov@rpharm.ruАкционерное общество «Р-Фарм»РоссияR-Pharm JSCRussian FederationАкционерное общество «Научно-производственное объединение «ДОМ ФАРМАЦИИ»РоссияResearch-and-manufacturing company “HOME OF PHARMACY”Russian FederationФедеральное государственное автономное научное учреждение "Федеральный научный центр исследований и разработки иммунобиологических препаратов им. М.П. Чумакова РАН" (Институт полиомиелита)РоссияChumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences (Institute of Poliomyelitis)Russian FederationР-Фарм Оверсиз инкСоединённые Штаты АмерикиR-Pharm Overseas, Inc.United States202208122022224Разработка биологических лекарственных препаратов для профилактики, диагностики и лечения COVID-19414434Copyright © Шипаева Е.В., Филон О.В., Зинченко А.В., Шипунов Г.А., Дмитриева А.А., Лемак М.С., Гришин С.А., Трофимец Е.И., Крышень К.Л., Козловская Л.И., Лунин А.С., Аполохов В.Д., Барбашов С.Ф., Лавровский Я.В., Самсонов М.Ю., 20222022Шипаева Е.В., Филон О.В., Зинченко А.В., Шипунов Г.А., Дмитриева А.А., Лемак М.С., Гришин С.А., Трофимец Е.И., Крышень К.Л., Козловская Л.И., Лунин А.С., Аполохов В.Д., Барбашов С.Ф., Лавровский Я.В., Самсонов М.Ю.Shipaeva E.V., Filon O.V., Zintchenko A.V., Shipunov G.A., Dmitrieva A.A., Lemak M.S., Grishin S.A., Trofimets E.I., Kryshen K.L., Kozlovskaya L.I., Lunin A.S., Apolokhov V.D., Barbashov S.F., Lavrovsky Y.V., Samsonov M.Y.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://www.biopreparations.ru/jour/article/view/437

Поиск эффективных и безопасных лекарственных средств для борьбы с коронавирусной инфекцией, вызванной вирусом SARS-CoV-2, является актуальной задачей. RPH-137 – оригинальный гибридный белок-ловушка вируса SARS-CoV-2, состоящий из внеклеточного домена ангиотензинпревращающего фермента 2 типа и Fc-фрагмента IgG1 человека.

Цель работы

Цель работы: доклиническая оценка эффективности RPH-137 и молнупиравира в отношении инфекции, вызванной SARS-CoV-2.

Материалы и методы

Материалы и методы: RPH-137 получали в стабильной линии клеток китайского хомячка. В работе использовали субстанцию молнупиравира. Изучение ингибирования вирус-индуцированной цитотоксичности проводили в культуре клеток Vero. В исследовании эффективности in vivo сирийских хомячков заражали интраназально SARS-CoV-2 (вариант ПИК35) в дозе 5 lg ТЦД50. Оценивали массу тела, массовый коэффициент и гистологическую картину легких. В орофарингеальных мазках измеряли содержание вирусной РНК методом ОТ-ПЦР по показателю порогового цикла амплификации Ct. Статистическая обработка: однофакторный и двухфакторный дисперсионный анализ (ANOVA), t-тест Стьюдента, критерий Манна–Уитни.

Результаты

Результаты: RPH-137 и молнупиравир ингибировали цитопатическое действие вируса SARS-CoV-2 в культуре клеток Vero: для RPH-137 EC50=4,69 мкг/мл (21,3 нМ) и 16,24 мкг/мл (73,8 нМ) для доз 50 ТЦД50 и 200 ТЦД50 соответственно, для молнупиравира EC50=0,63 мкг/мл (1900 нМ) для обеих доз вируса. RPH-137 при внутримышечном введении в дозах 30 и 80 мг/кг не оказывал влияния на развитие инфекции у сирийских хомячков. RPH-137 при внутрибрюшинном введении в дозе 100 мг/кг показал статистически значимый эффект по ряду параметров по сравнению с животными контрольной группы (контроль заражения), в том числе вызывая снижение воспалительного процесса и общей площади поражения легких на 7 сут на 27 и 30% соответственно. Молнупиравир при пероральном введении в дозе 300 мг/кг 2 раза в сутки значимо подавлял развитие инфекции, вызванной SARS-CoV-2.

Выводы

Выводы: RPH-137 и  молнупиравир ингибируют цитопатическое действие вируса SARS-CoV-2 в культуре клеток Vero. У сирийских хомячков введение молнупиравира демонстрировало более выраженное подавление инфекции, вызванной SARS-CoV-2, по сравнению с RPH-137. Однако RPH-137 проявлял статистически значимое действие по ряду параметров, что открывает перспективы для его дальнейшего изучения. 

Finding effective and safe medicines to fight SARS-CoV-2 infection is an urgent task. RPH-137 is an original trap fusion protein against SARS-CoV-2 virus. It comprises the angiotensin-converting enzyme type 2 extracellular domain and the human IgG1 Fc fragment.

The aim of the study was to carry out a preclinical evaluation of the efficacy of RPH-137 and molnupiravir against SARS-CoV-2 infection.

Materials and methods

Materials and methods: the authors analysed RPH-137 expressed in a stable CHO cell line and molnupiravir used as an active pharmaceutical ingredient. Drug-mediated inhibition of virus-induced cytotoxicity was assessed in Vero cell culture. In vivo efficacy assessments were performed in Syrian hamsters. The animals were infected intranasally with SARS-CoV-2 (PIK35 clinical isolate) in the dose of 5 log TCID50. The authors evaluated body weight measurements, lung–body weight ratios, and lung histopathology findings and determined viral RNA levels in oropharyngeal swabs by RT-PCR using the amplification cycle threshold (Ct). The statistical analyses involved one- and two-way ANOVA, Student's t-test, and Mann–Whitney test.

Results

Results: RPH-137 and molnupiravir inhibited the cytopathic effect of SARS-CoV-2 in Vero cells; the EC50 values of RPH-137 amounted to 4.69 μg/mL (21.3 nM) and 16.24 μg/mL (73.8 nM) for 50 TCID50 and 200 TCID50, respectively, whereas the EC50 values of molnupiravir were 0.63 μg/mL (1900 nM) for both doses. Intramuscular RPH-137 (30 and 80 mg/kg) had no effect on the infection process in Syrian hamsters. The comparison with the challenge control group showed that intraperitoneal RPH-137 (100 mg/kg) had statistically significant effects on a number of parameters, including a 27% reduction in inflammation and a 30% reduction in the total lesion area of the lungs by Day 7. Intragastric molnupiravir (300 mg/kg twice daily) significantly inhibited SARS-CoV-2 infection.

Conclusions

Conclusions: both RPH-137 and molnupiravir inhibited the cytopathic effect of SARS-CoV-2 in Vero cells. In Syrian hamsters, molnupiravir demonstrated a more pronounced inhibition of SARS-CoV-2 infection than RPH-137. However, RPH-137 had statistically significant effects on a range of parameters. This offers additional perspectives for further research.

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The authors declare that there are no conflicts of interest present.