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Maria Candida M. Mellado

Bio: Maria Candida M. Mellado is an academic researcher from Universidade Nova de Lisboa. The author has contributed to research in topics: Membrane & Gel electrophoresis. The author has an hindex of 8, co-authored 11 publications receiving 692 citations. Previous affiliations of Maria Candida M. Mellado include Federal University of Rio de Janeiro.

Papers
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Journal ArticleDOI
TL;DR: This article focuses on the essential role of VLP technology in new-generation vaccines against prevalent and emergent diseases and the implications of large-scale VLP production in the context of process control, monitorization and optimization.
Abstract: Virus-like particles (VLPs) are multiprotein structures that mimic the organization and conformation of authentic native viruses but lack the viral genome, potentially yielding safer and cheaper vaccine candidates. A handful of prophylactic VLP-based vaccines is currently commercialized worldwide: GlaxoSmithKline's Engerix (hepatitis B virus) and Cervarix (human papillomavirus), and Merck and Co., Inc.'s Recombivax HB (hepatitis B virus) and Gardasil (human papillomavirus) are some examples. Other VLP-based vaccine candidates are in clinical trials or undergoing preclinical evaluation, such as, influenza virus, parvovirus, Norwalk and various chimeric VLPs. Many others are still restricted to small-scale fundamental research, despite their success in preclinical tests. This article focuses on the essential role of VLP technology in new-generation vaccines against prevalent and emergent diseases. The implications of large-scale VLP production are discussed in the context of process control, monitorization and optimization. The main up- and down-stream technical challenges are identified and discussed accordingly. Successful VLP-based vaccine blockbusters are briefly presented concomitantly with the latest results from clinical trials and the recent developments in chimeric VLP-based technology for either therapeutic or prophylactic vaccination.

669 citations

Journal ArticleDOI
TL;DR: DLP and TLP were found to coexist under certain conditions as determined from all reaction products analyzed by capillary electrophoresis, and these results contribute to the design of new strategies for the improvement of TLP yield and quality by reducing the VP7 detachment from TLP.
Abstract: Virus-like particles constitute potentially relevant vaccine candidates. Nevertheless, their behavior in vitro and assembly process needs to be understood in order to improve their yield and quality. In this study we aimed at addressing these issues and for that purpose triple- and double-layered rotavirus-like particles (TLP 2/6/7 and DLP 2/6, respectively) size and zeta potential were measured using dynamic light scattering at different physicochemical conditions, namely pH, ionic strength, and temperature. Both TLP and DLP were stable within a pH range of 3-7 and at 5-25 degrees C. Aggregation occurred at 35-45 degrees C and their disassembly became evident at 65 degrees C. The isoelectric points of TLP and DLP were 3.0 and 3.8, respectively. In vitro kinetics of TLP disassembly was monitored. Ionic strength, temperature, and the chelating agent employed determined disassembly kinetics. Glycerol (10%) stabilized TLP by preventing its disassembly. Disassembled TLP was able to reassemble by dialysis at high calcium conditions. VP7 monomers were added to DLP in the presence of calcium to follow in vitro TLP assembly kinetics; its assembly rate being mostly affected by pH. Finally, DLP and TLP were found to coexist under certain conditions as determined from all reaction products analyzed by capillary electrophoresis. Overall, these results contribute to the design of new strategies for the improvement of TLP yield and quality by reducing the VP7 detachment from TLP.

30 citations

Journal ArticleDOI
15 Feb 2010-Talanta
TL;DR: It is demonstrated that insect cell mass spectrum fingerprints are characteristic of each viral protein/particle production and correlate with the intracellular viral protein content determined by Western blot.

26 citations

Journal ArticleDOI
TL;DR: The SDS-CGE method proved to be fast, consistent and reproducible, representing a feasible alternative to the laborious conventional electrophoresis for the characterization of RLPs, candidate vaccines against rotavirus infection.

24 citations

Journal ArticleDOI
TL;DR: The efficacy of the proposed purification strategy is confirmed, that now enables RLPs assembly studies and yielded high level of purity (>90%).

22 citations


Cited by
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Journal ArticleDOI
TL;DR: This article focuses on the essential role of VLP technology in new-generation vaccines against prevalent and emergent diseases and the implications of large-scale VLP production in the context of process control, monitorization and optimization.
Abstract: Virus-like particles (VLPs) are multiprotein structures that mimic the organization and conformation of authentic native viruses but lack the viral genome, potentially yielding safer and cheaper vaccine candidates. A handful of prophylactic VLP-based vaccines is currently commercialized worldwide: GlaxoSmithKline's Engerix (hepatitis B virus) and Cervarix (human papillomavirus), and Merck and Co., Inc.'s Recombivax HB (hepatitis B virus) and Gardasil (human papillomavirus) are some examples. Other VLP-based vaccine candidates are in clinical trials or undergoing preclinical evaluation, such as, influenza virus, parvovirus, Norwalk and various chimeric VLPs. Many others are still restricted to small-scale fundamental research, despite their success in preclinical tests. This article focuses on the essential role of VLP technology in new-generation vaccines against prevalent and emergent diseases. The implications of large-scale VLP production are discussed in the context of process control, monitorization and optimization. The main up- and down-stream technical challenges are identified and discussed accordingly. Successful VLP-based vaccine blockbusters are briefly presented concomitantly with the latest results from clinical trials and the recent developments in chimeric VLP-based technology for either therapeutic or prophylactic vaccination.

669 citations

Journal ArticleDOI
TL;DR: N nanoscale size materials such as virus-like particles, liposomes, ISCOMs, polymeric, and non-degradable nanospheres have received attention as potential delivery vehicles for vaccine antigens which can both stabilize vaccine antIGens and act as adjuvants, thereby modulating the immune response to the antigen.
Abstract: Vaccination has had a major impact on the control of infectious diseases. However, there are still many infectious diseases for which the development of an effective vaccine has been elusive. In many cases the failure to devise vaccines is a consequence of the inability of vaccine candidates to evoke appropriate immune responses. This is especially true where cellular immunity is required for protective immunity and this problem is compounded by the move toward devising sub-unit vaccines. Over the past decade nanoscale size (<1000 nm) materials such as virus-like particles, liposomes, ISCOMs, polymeric, and non-degradable nanospheres have received attention as potential delivery vehicles for vaccine antigens which can both stabilize vaccine antigens and act as adjuvants. Importantly, some of these nanoparticles (NPs) are able to enter antigen-presenting cells by different pathways, thereby modulating the immune response to the antigen. This may be critical for the induction of protective Th1-type immune responses to intracellular pathogens. Their properties also make them suitable for the delivery of antigens at mucosal surfaces and for intradermal administration. In this review we compare the utilities of different NP systems for the delivery of sub-unit vaccines and evaluate the potential of these delivery systems for the development of new vaccines against a range of pathogens.

441 citations

Journal ArticleDOI
TL;DR: The main principles in the cloning of viral structural genes, the relevant host systems and the purification procedures that have been developed are discussed and the methods that are used to characterize the structural integrity, stability, and components of newly synthesized VLPs are analyzed.
Abstract: Over the last three decades, virus-like particles (VLPs) have evolved to become a widely accepted technology, especially in the field of vaccinology. In fact, some VLP-based vaccines are currently used as commercial medical products, and other VLP-based products are at different stages of clinical study. Several remarkable advantages have been achieved in the development of VLPs as gene therapy tools and new nanomaterials. The analysis of published data reveals that at least 110 VLPs have been constructed from viruses belonging to 35 different families. This review therefore discusses the main principles in the cloning of viral structural genes, the relevant host systems and the purification procedures that have been developed. In addition, the methods that are used to characterize the structural integrity, stability, and components, including the encapsidated nucleic acids, of newly synthesized VLPs are analyzed. Moreover, some of the modifications that are required to construct VLP-based carriers of viral origin with defined properties are discussed, and examples are provided.

349 citations

Journal ArticleDOI
07 Jul 2016-Nature
TL;DR: The computational design of a 25-nanometre icosahedral nanocage that self-assembles from trimeric protein building blocks is described, and such robust and customizable nanocages should have considerable utility in targeted drug delivery, vaccine design and synthetic biology.
Abstract: The dodecahedron [corrected] is the largest of the Platonic solids, and icosahedral protein structures are widely used in biological systems for packaging and transport. There has been considerable interest in repurposing such structures for applications ranging from targeted delivery to multivalent immunogen presentation. The ability to design proteins that self-assemble into precisely specified, highly ordered icosahedral structures would open the door to a new generation of protein containers with properties custom-tailored to specific applications. Here we describe the computational design of a 25-nanometre icosahedral nanocage that self-assembles from trimeric protein building blocks. The designed protein was produced in Escherichia coli, and found by electron microscopy to assemble into a homogenous population of icosahedral particles nearly identical to the design model. The particles are stable in 6.7 molar guanidine hydrochloride at up to 80 degrees Celsius, and undergo extremely abrupt, but reversible, disassembly between 2 molar and 2.25 molar guanidinium thiocyanate. The dodecahedron [corrected] is robust to genetic fusions: one or two copies of green fluorescent protein (GFP) can be fused to each of the 60 subunits to create highly fluorescent ‘standard candles’ for use in light microscopy, and a designed protein pentamer can be placed in the centre of each of the 20 pentameric faces to modulate the size of the entrance/exit channels of the cage. Such robust and customizable nanocages should have considerable utility in targeted drug delivery, vaccine design and synthetic biology.

339 citations

Journal ArticleDOI
TL;DR: This review article focuses on the applications of nanocarrier-based vaccine formulations and the strategies used for the functionalization of nanoparticles to accomplish efficient delivery of vaccines in order to induce desired host immunity against infectious diseases.
Abstract: Due to emergence of new variants of pathogenic micro-organisms the treatment and immunization of infectious diseases have become a great challenge in the past few years. In the context of vaccine development remarkable efforts have been made to develop new vaccines and also to improve the efficacy of existing vaccines against specific diseases. To date, some vaccines are developed from protein subunits or killed pathogens, whilst several vaccines are based on live-attenuated organisms, which carry the risk of regaining their pathogenicity under certain immunocompromised conditions. To avoid this, the development of risk-free effective vaccines in conjunction with adequate delivery systems are considered as an imperative need to obtain desired humoral and cell-mediated immunity against infectious diseases. In the last several years, the use of nanoparticle-based vaccines has received a great attention to improve vaccine efficacy, immunization strategies, and targeted delivery to achieve desired immune responses at the cellular level. To improve vaccine efficacy, these nanocarriers should protect the antigens from premature proteolytic degradation, facilitate antigen uptake and processing by antigen presenting cells, control release, and should be safe for human use. Nanocarriers composed of lipids, proteins, metals or polymers have already been used to attain some of these attributes. In this context, several physico-chemical properties of nanoparticles play an important role in the determination of vaccine efficacy. This review article focuses on the applications of nanocarrier-based vaccine formulations and the strategies used for the functionalization of nanoparticles to accomplish efficient delivery of vaccines in order to induce desired host immunity against infectious diseases.

309 citations