Vaccines are a major contribution to global health and their manufacturing can be complex due to their diversity and inherent properties. Such complexity is exacerbated in outbreaks and pandemics and thus the traditional development and manufacturing paradigm is no longer valid. In the last decade, outbreaks and pandemics of H1N1 Influenza, Ebola, MERS, Zika and now SARS-Cov2 (COVID19) have heavily influenced vaccine manufacturing and lessons learned have allowed to improve our general response to high surge of vaccines. Despite of that, there are remaining issues around process development, manufacturing processes and capacity. How can vaccine manufacturing evolve to increase efficiency, speed and global access? What does the vaccine facility of the future could look like and how platforming vaccines can help addressing future global health challenges?

In this lecture, I will discuss the recent situation of the vaccines against SARS-CoV-2.

Leishmaniasis is a neglected tropical disease caused by Leishmania protozoa transmitted by infected sand flies. Today this disease is spread in more than 88 countries and affecting more than 320 million people. There is no licensed vaccine available against human leishmaniasis. We have previously established a CRISPR genome-edited attenuated Leishmania major strain (LmCen-/-) as a second-generation leishmanization vaccine candidate. We evaluated the efficacy of LmCen-/- against VL caused by L.donovani infection in a pre-clinical hamster model as well as immunogenicity in the PBMCs isolated from human subjects living in an endemic region. Intradermal immunization of hamsters with LmCen-/- did not develop any lesion; while inducing a strong pro-inflammatory immune response. Immunized hamsters when challenged with L. donovani either by intradermal needle injection or by infected sand flies were protected as indicated by a lack of pathology and mortality. Spleen cells from LmCen-/- immunized and challenged hamsters produced significantly higher Th1-associated cytokines including IFN-γ and TNF-α, and significantly reduced expression of the anti-inflammatory cytokines IL-10 and IL-21, compared to non-immunized and challenged animals. We developed a GLP-grade LmCen-/- which is equally protective as laboratory-grade LmCen-/- parasites. GLP-grade LmCen-/- parasites induced a pro-inflammatory immune response in the PBMCs isolated from healed VL patients as well as healthy people living in the endemic region. Together, this study demonstrates that the LmCen-/- parasites are safe and efficacious against VL and can proceed to be tested as a vaccine in human clinical trials. These findings could help in the development and evaluation of effective Leishmania vaccines against Leishmaniasis.

Viral infections are an extremely common and predictable problem in organ and hematopoietic stem cell transplant patients. Antiviral drugs given either prophylactically or as early therapy for patients detected to be shedding virus appears to be an effective strategy for reducing herpes virus infections. However, long-term treatment with these drugs is associated with significant toxicity, expense and the appearance of drug resistance virus isolates ultimately resulting in treatment failure. Over the last few years, there is increasing evidence that cellular immune therapies can reverse the outgrowth of hematological malignancies and can also provide therapeutic benefit against lethal viral infections. While the expansion and adoptive transfer of virus-specific T-cells from the healthy original donor can be an effective strategy to control viral replication, this is not possible when donors are seronegative or are subsequently inaccessible. Studies from our group have demonstrated the successful expansion of human cytomegalovirus (CMV) and Epstein-Barr virus (EBV)-specific T-cells from solid organ and stem cell transplant recipients with active viral infection or disease and the long term reconstitution of protective anti-viral immunity following their adoptive transfer back into the patients. More recently, we have also developed a single platform technology to extend this immunotherapeutic strategy for multiple pathogens including CMV, EBV, adenovirus and BK polyoma virus. T cells directed towards multiple pathogens can be rapidly expanded and can be used for adoptive immunotherapy. Finally, in collaboration with an Australian biotech company, Cellestis Inc., we have also developed a novel T cell-based immune monitoring technology (QuantiFERON-CMVâ) which will allow us to identify high risk transplant patients who may develop virus-associated complications post-transplantation.

Novel coronavirus SARS-CoV-2 outbreaks have rapidly spread to multiple countries, highlighting the urgent necessity for fast, sensitive, and specific diagnostic tools for virus surveillance. We have previously demonstrated that Class 1 type I CRISPR, which is composed of Escherichia coli Cascade, Cas3, and programmable pre-crRNA, mediates distinct DNA cleavage activity in human cells. Here, the previously unknown collateral single-stranded DNA cleavage we observed with type I CRISPR-Cas3 highlights its potential for development as a Cas3-mediated rapid (within 40 min), low-cost, instrument-free detection method for SARS-CoV-2. This Cas3-based assay is comparable with Cas12- and real-time reverse-transcriptase PCR-based assays in its speed and sensitivity, but offers greater specificity for single-base-pair discrimination while negating the need for highly trained operators. These findings support the use of CRISPR diagnostics for point-of-care testing in patients with suspected SARS-CoV-2 infections.

This presentation introduces the process of accelerated productization, pioneered by Diagnostics Development (DxD) Hub. DxD Hub is a Singapore national platform for diagnostics development and productization, funded by the National Research Foundation and hosted by the Agency for Science, Technology and Research (A*STAR). The key to accelerated productization lies in (i) end-to-end productization: understanding user requirements, and developing the regulatory framework of minimal viable product based on format of the end product; and (ii) agile iterative process: quick succession of iteration based on evolving market and user input. DxD Hub model is based on that of a public-private partnership involving R&D entities, clinical community and commercial parties. Using the example of RESOLUTE - a direct polymerase chain reaction (PCR) test, this presentation illustrates the productisation journey of a diagnostic test. RESOLUTE received provisional authorisation from the Health Sciences Authority in April 2020, and is a collaboration between Defence Science Organisation, Advanced Remanufacturing Technology Centre, and commercial partner Advanced Medtech.

The general focus of my lab is to characterize at the molecular level, genome editing proteins derived from the bacterial CRISPR-Cas system and apply them for reversing disease causing mutations. Using biophysical tools, we dissect the mechanism of action of Cas9 proteins with respect to nucleic acid interrogation and use this knowledge to design editors with high gene targeting specificity. We combine biochemical tools for producing recombinant Cas9 proteins, induced pluripotent stem cell (iPSC) technology for studying their efficacy in patient cells and single molecule experiments for evaluating their targeting specificity. Based on this broad theme, there are two key areas where I carry out research on:

1) We have extensively characterised a TypeII Cas9 from Francisella novicida (FnCas9) and demonstrated its ability in genome editing of the sickle cell anemia mutation in patient derived iPSCs and HSCs. FnCas9 is highly specific in target interrogation and produces negligibe background insertions/deletions in patient Hematopoietic stem cells. Currently we are working towards completing pre-clinical studies on FnCas9

2) We have repurposed the high specificity of FnCas9 towards a Point of Care diagnostic assay for detecting nucleotide variants in a target. This can perform with minimum equipment and technical know-how and is being currently deployed for rapid and accurate diagnosis of COVID- 19 positive cases. By tying up with industries, we are scaling up the production of assay components to enable testing on a large scale during the ongoing pandemic.

Current bottlenecks for improving accessibility and scalability of SARS-CoV-2 testing include diagnostic assay costs, complexity, and supply chain shortages. To resolve these issues, we developed SalivaDirect. The critical component of our approach is to use saliva instead of respiratory swabs, which enables non-invasive frequent sampling and reduces the need for trained healthcare professionals during collection. Furthermore, we simplified our diagnostic test by (1) not requiring nucleic acid preservatives at sample collection, (2) replacing nucleic acid extraction with a simple proteinase K and heat treatment step, and (3) testing specimens with a dualplex quantitative reverse transcription PCR (RT-qPCR) assay. We validated SalivaDirect with reagents and instruments from multiple vendors to minimize the risk for supply chain issues. Regardless of our tested combination of reagents and instruments from different vendors, we found that SalivaDirect is highly sensitive with a limit of detection of 6-12 SARS-CoV-2 copies/μL. When comparing paired nasopharyngeal swabs and saliva specimens using the authorized ThermoFisher Scientific TaqPath COVID-19 combo kit and our SalivaDirect protocol, we found high agreement in testing outcomes (>94%). Being flexible and inexpensive ($1.29-$4.37/sample), SalivaDirect is a viable and accessible option to help alleviate SARS-CoV-2 testing demands. SalivaDirect received Emergency Use Authorization from the FDA on August 15th, 2020. Details can be found on our website: covidtrackerct.com/about-salivadirect.

In this presentation Dr. Erfan Ibrahim will describe a few predictive models for COVID-19 infections and fatality rates that were developed for the US and India and how they compare with the actual statistics of the pandemic over the past 6 months. Dr. Ibrahim will also share a model he developed from empirical data on the actual fatality rate associated with COVID-19 and the average duration between date of infection and date of death of patients in severe cases.

COVID-19 has presented unprecedented challenges for epidemiologists, as well as health care, the economy and society. Keeping up with the dizzying pace of change in the epidemic and producing consistently policy-relevant modelling predictions has been more challenging than ever before. In this context, a range of models have been used, and although they differ considerably in their construction, many of the messages have been consistent and accurate. Producing precise predictions of the epidemic trajectory has been more challenging for several reasons, including that the data available is surprisingly sparse. Months into the pandemic and with millions of cases worldwide, much of our understanding is still derived from small groups of well-described patients.

Summarize ten important trends of China’s healthcare system after the impact of COVID-19, including the increasing importance of public health, community care, healthcare professionals, internet-based healthcare, outpatient care and so on.

Dengue virus (DENV) is a major human pathogen infecting approximately 400 million people annually worldwide. However, currently there is no highly effective vaccine and therapeutics are non-existent. Dr Lok’s research reveals the differing particle morphology of DENV, when placed in the presence of four different serotypes. The changes in particle morphology, thus antigenicity, is due to the rearrangement of the surface viral envelope (E) proteins allowing DENV to evade host immunity. This in turn complicates the development of effective vaccines and therapeutics. Dr Lok will discuss the extensive studies that have been conducted to examine the different morphologies displayed by various DENV strains and how Lok and her team identify molecular determinants on the E proteins that lead to different structural changes. By studying the different morphological variants displayed by strains within each serotype, Dr Lok posits that we may be able to incorporate “representative morphologies” for each serotype to make a more effective vaccine with greater coverage against all DENV serotypes antigenic structures. Professor Lok will also discuss how highly potent human antibodies can neutralize all or some of the morphological variants of four DENV serotypes.

Infectious disease epidemiology plays an important role in understand the biology of a pathogen and disease progression at a population level. A number of intrinsic and extrinsic factors play role in spread of infectious diseases and some of these aspects will be deliberated in the context of an arbovirus, namely, chikungunya virus. Further, the impact of non-communicable diseases on the epidemiology of the current COVID-19 epidemic will be discussed.

Since the outbreak of COVID-19, 2-3% of COVID-19 patients died in short time. Immune responses may play critical role in this process. The numbers of T cells and NK cells were reduced in SARS-CoV-2-infected patients, which was probably due to the combinational interactions of these cells. Our study showed a decreased T cell proportion with down-regulated gene expression related to T cell activation and differentiation were occurred in COVID-19 severe patients, which may help to understand the immune response and provide effective treatment strategies for COVID-19.

Viral infections result in a wide spectrum of clinical outcomes, ranging from asymptomatic infection to severe and even life-threatening diseases. Such varied infection outcome applies also to the newly emerged SARS-CoV-2, the cause of COVID-19. While the correlates of severe disease have been explored for various infections, the pathophysiology that differentiates symptomatic from asymptomatic infection remains ill understood. We recently identified the molecular underpinnings of symptomatic infection, using the live attenuated yellow fever virus vaccine (YF17D) as a model. Using whole blood transcriptomic and metabolomic profiling, we found that individuals with adaptive endoplasmic reticulum (ER) stress and reduced tricarboxylic acid (TCA) cycle activity at baseline showed increased susceptibility to symptomatic outcome. Like most viral infections, YF17D infection induced translational and metabolic activities necessary, at least in part, to reproduce new viruses. Such increased activities exacerbated ER stress. The ensuing maladaptive ER stress response triggered downstream pro-inflammatory responses that cascaded into symptomatic outcome. Our findings thus suggest that ER stress response and immunometabolism underpin symptomatic yellow fever; similar processes could also extend to other viral infections, including COVID-19. I will summarize these findings and suggest how antiviral approaches could be applied to prevent disease development and progression.

The ongoing COVID-19 pandemic has driven unprecedented efforts in vaccine research and development against the etiologic agent SARS-CoV-2. The usual timeframe for vaccine development, 5-10 years, has been reduced to 12-18 months, and in the past 8 months, roughly 200 vaccine development projects have been initiated. 24 vaccines are in human clinical trials, and Phase III activity has started for 2-3 candidate vaccines and will soon start for several more. While many are cautiously optimistic that we may prove that at least one vaccine is safe and effective, we cannot guarantee the timeframe, the durability of protection, or the long-term safety of these regimens. We will review the major published data from vaccine clinical trials and put them into the context of what is known or hypothesized about relevant immune responses and potential safety concerns. Finally we will address issues of manufacturing, access, and uptake that may profoundly affect the ability of COVID-19 vaccines to return us to the “old” normal.