Periodic Reporting for period 3 - IF-EBOla (Control of the Ebola Oubreak by both innovative Ultrasensitive Detection of EBOV and therapy)
Reporting period: 2016-11-01 to 2017-10-31
Ebola virus (EBOV) disease (EVD) is a hemorrhagic fever that can be fatal if left untreated. The 2014 outbreak in West Africa reached historic proportions and spread rapidly beyond Africa, emphasizing the lack of ultrasensitive diagnostics and efficient drugs against EBOV. Currently, there are no vaccines or antiviral drugs approved for prevention or treatment of EVD. As it can be transmitted to humans from infected animals, there is an imminent need for prompt diagnosis and protection strategies in EBOV endemic regions. In addition, better understanding of survival and pathogenesis factors are critical for the design of appropriate countermeasures. The EU-funded IF-EBOLA project was a collaborative effort among European, USA, Canada and African partners designed to control EBOV spreading during an outbreak. The key objective was to “set up both pragmatic and fundamental countermeasure strategies for efficient preparedness by local public health authorities to control future Ebola outbreaks.
The concept of the IF-EBOLA was to combine an early passive anti-Ebola immunotherapy prompted by timely EBOV-diagnosis, along with immune response monitoring. We developed different innovative technological tools including methods for the ultrasensitive detection of Zaire Ebola virus (ZEBOV) alongside a passive immunotherapy with polyclonal antibody strategy targeting ZEBOV designed to drastically improve their prognosis. Additionally, accurate monitoring of the efficiency of immune response in EVD survivors was performed to achieve a better assessing survival factors and improve the design novel therapies and vaccine strategies.
The concept of the IF-EBOLA was to combine an early passive anti-Ebola immunotherapy prompted by timely EBOV-diagnosis, along with immune response monitoring. We developed different innovative technological tools including methods for the ultrasensitive detection of Zaire Ebola virus (ZEBOV) alongside a passive immunotherapy with polyclonal antibody strategy targeting ZEBOV designed to drastically improve their prognosis. Additionally, accurate monitoring of the efficiency of immune response in EVD survivors was performed to achieve a better assessing survival factors and improve the design novel therapies and vaccine strategies.
Initially, the IF-EBOLA action (during “the emergency period”) included two main clinical monitoring phases: patients and convalescents immune-infectious profiling (i) without and (ii) with our specific treatment. Due to epidemic breakdown and major foreseen and unforeseen changes produced during this “non-emergency” period, we have been authorized to modify and extend for one year our program consisting now in three main actions: (i) Preclinical validation of the therapeutic equine anti-Ebola antibodies; (ii) Retrospective evaluation of the ultrasensitive detection EBOV in historical clinical samples (iii) profiling homeostasis (“immune-pathogenesis parameters”) of patients and survivors.
IF-EBOLA components of a novel integrated approach tackling EVD:
(i) The IF -EBOLA Fab’ntech’s therapy was based in the development of horse anti-EBOV polyclonal F(ab’)2 using a well-established production method (stated by EMA) of ZEBOV antigen that exhibiting very good pharmacological characteristics, and a high anti-viral potency in vitro and in vivo. The F(ab’)2-treated lethally-challenged animals (mice and guinea pigs with a 8-days life expectancy) reached 100% of survive and 100% rescued dying animals (treated 3 days after their lethal challenge). For preparedness, we have stored 900 liters of high-affinity anti-ZEBOV polyclonal serum ready to potentially treat over 20 000 EVD patients. An NHP model will be tested with the NIH (USA);
(ii) Undoubtedly, EBOV detection must be done at the early EVD phase (low viral load and symptoms), to apply supportive and specific treatments, to isolate patients to improve the epidemic control. Also, it is important to treat potential convalescent transmitters exhibiting residual viral loads. To detect residual viruses, we used an ApoH-coated magnetic-bead sample enrichment preparation to concentrate virus in patient samples and combine it with the FDA-approved USAMRIID real-time PCR method (as tested in Sierra Leone and in Liberia and at the USAMRIID in Frederick, US). We achieved an ApoH-dependent ultrasensitive EBOV detection to reveal the presence of residual EBOV from self-cured convalescents previously acutely infected. Using the same method of ultrasensitive enrichment detect bacteria (potentially involved in sepsis) as few as 1 bacterium/5 mL of whole blood and also generate more NGS reads for accurate metagenomic analysis.
(iii) To characterize the immune response against Ebola is key to assess immune and infectious diversity parameters. To this, state-of-the-art technologies were used such as next generation sequencing, metagenomics and RNA arrays. OIB has developed an extremely potent, new hardware and algorithm (RIGEL) dedicated for its efficient use in the field for rapid sequencing and metagenomic analyses; a new outstanding corrected gene bank with 22-times more data than the traditional Genebank. Revealed a prevalence close to 20% of Lassa virus, alongside of nosocomial bacteria and current local infections such as HIV, Mycobacterium tuberculosis and malaria. In addition, the RIGEL algorithm permitted to detect microbiological profiles associated to either survival or fatal outcomes. Future vaccines and therapeutic strategies depend on virus diversity and targeted viral components, such specific viral epitopes. Thus, deep cell and humoral immune response analyses were done by BGU in a survivor population from Liberia by permitting to determine new critical epitopes associating neutralizing capacity of antibodies and cell responses in survivors.
(iv) We have generated compounds able to inhibit enzymes involved in vascular leakage (a hallmark of EVD) resulting from exacerbated inflammatory processes. We published the in vitro proof-of-concept and now we are preparing an in vivo proof-of-concept in a vascular leakage model.
Due to changes and issues faced by the consortium, the communication and dissemination activities have essentially been done through presentation in international conferences, press releases and 2 peer-reviewed publications. A recently submitted manuscript, on the F(ab’)2 efficacy precedes seven other manuscripts in preparation with data generated by OIB (2), BGU (2), Fabentech (1) and IRD (2) (cf. section 6).
IF-EBOLA components of a novel integrated approach tackling EVD:
(i) The IF -EBOLA Fab’ntech’s therapy was based in the development of horse anti-EBOV polyclonal F(ab’)2 using a well-established production method (stated by EMA) of ZEBOV antigen that exhibiting very good pharmacological characteristics, and a high anti-viral potency in vitro and in vivo. The F(ab’)2-treated lethally-challenged animals (mice and guinea pigs with a 8-days life expectancy) reached 100% of survive and 100% rescued dying animals (treated 3 days after their lethal challenge). For preparedness, we have stored 900 liters of high-affinity anti-ZEBOV polyclonal serum ready to potentially treat over 20 000 EVD patients. An NHP model will be tested with the NIH (USA);
(ii) Undoubtedly, EBOV detection must be done at the early EVD phase (low viral load and symptoms), to apply supportive and specific treatments, to isolate patients to improve the epidemic control. Also, it is important to treat potential convalescent transmitters exhibiting residual viral loads. To detect residual viruses, we used an ApoH-coated magnetic-bead sample enrichment preparation to concentrate virus in patient samples and combine it with the FDA-approved USAMRIID real-time PCR method (as tested in Sierra Leone and in Liberia and at the USAMRIID in Frederick, US). We achieved an ApoH-dependent ultrasensitive EBOV detection to reveal the presence of residual EBOV from self-cured convalescents previously acutely infected. Using the same method of ultrasensitive enrichment detect bacteria (potentially involved in sepsis) as few as 1 bacterium/5 mL of whole blood and also generate more NGS reads for accurate metagenomic analysis.
(iii) To characterize the immune response against Ebola is key to assess immune and infectious diversity parameters. To this, state-of-the-art technologies were used such as next generation sequencing, metagenomics and RNA arrays. OIB has developed an extremely potent, new hardware and algorithm (RIGEL) dedicated for its efficient use in the field for rapid sequencing and metagenomic analyses; a new outstanding corrected gene bank with 22-times more data than the traditional Genebank. Revealed a prevalence close to 20% of Lassa virus, alongside of nosocomial bacteria and current local infections such as HIV, Mycobacterium tuberculosis and malaria. In addition, the RIGEL algorithm permitted to detect microbiological profiles associated to either survival or fatal outcomes. Future vaccines and therapeutic strategies depend on virus diversity and targeted viral components, such specific viral epitopes. Thus, deep cell and humoral immune response analyses were done by BGU in a survivor population from Liberia by permitting to determine new critical epitopes associating neutralizing capacity of antibodies and cell responses in survivors.
(iv) We have generated compounds able to inhibit enzymes involved in vascular leakage (a hallmark of EVD) resulting from exacerbated inflammatory processes. We published the in vitro proof-of-concept and now we are preparing an in vivo proof-of-concept in a vascular leakage model.
Due to changes and issues faced by the consortium, the communication and dissemination activities have essentially been done through presentation in international conferences, press releases and 2 peer-reviewed publications. A recently submitted manuscript, on the F(ab’)2 efficacy precedes seven other manuscripts in preparation with data generated by OIB (2), BGU (2), Fabentech (1) and IRD (2) (cf. section 6).
Collectively, from a rationally-designed preparedness strategy, the European-US-Canadian-African (Sierra Leone & Liberia MoH partners) IF-EBOLA consortium has generated outstanding tools not only to drastically pave the way for new therapeutic and diagnostic approaches, but also to modify strategies to contain outbreaks. These IF-EBOLA products have fueled involved European and USA companies from the consortium to remain very active to extend their works to move to the international market. Thus, Fab’ntech is applying for funding through partnerships in the US including the NIH to go through an NHP validation of anti-Ebola F(ab’)2 before the clinical trial to be ready to propose thousands of therapeutic doses for patients of next Ebola outbreaks and extending this approach for other VHF. The success of algorithms for metagenomic NGS analyses developed by OIB are now being developed and used by the DHS from the US and the European Union for different surveillance purposes.
The successes of proof-of-concepts done for each one of the IF-EBOLA actions permitted establishing a rich “preparedness IF-EBOLA toolbox”. These tools and concepts are being promoted through different ongoing European projects (PANDORA-ID-NET (2018-2022) & EPIRISK-EBOV (2019-2021)) “extending” the IF-EBOLA action (the consortium staying very active including other ongoing proposals in the USA) as part of our integrated approach: “A Total One-Health Approach for Restauration Impacts”.
The successes of proof-of-concepts done for each one of the IF-EBOLA actions permitted establishing a rich “preparedness IF-EBOLA toolbox”. These tools and concepts are being promoted through different ongoing European projects (PANDORA-ID-NET (2018-2022) & EPIRISK-EBOV (2019-2021)) “extending” the IF-EBOLA action (the consortium staying very active including other ongoing proposals in the USA) as part of our integrated approach: “A Total One-Health Approach for Restauration Impacts”.