Abstract

Cancer vaccines based on tumor cell components have shown promising results in animal and clinical studies. The vaccine system contains abundant tumor antigen components, which can activate the immune system by antigens. However, their efficacy has been limited by the inability of antigens delivery, which are the core components of vaccines, further fail to be presented and activation of effective cells. Nanotechnology offers a novel platform to enhance the immunogenicity of tumor-associated antigens and deliver them to antigen-presenting cells (APCs) more efficiently. In addition, nanotreatment of tumor cells derivate active ingredients could also help improve the effectiveness of cancer vaccines. In this review, we summarize recent advances in the development of cancer vaccines by the combination of nanotechnology and tumor-based ingredients, including liposomes, polymeric nanoparticles, metallic nanoparticles, virus-like particles and tumor cells membrane, tumor lysate, and specific tumor antigens. These nanovaccines have been designed to increase antigen uptake, prolong antigen presentation, and modulate immune responses through codelivery of immunostimulatory agents. We also further discuss challenges and opportunities in the clinical translation of these nanovaccines.

Introduction

Cancer immunotherapy has revolutionized cancer treatment by harnessing the immune system to recognize and attack cancer cells [1–3]. Among various immunotherapeutic approaches, cancer vaccines have attracted significant attention due to their potential to induce long-lasting and specific immune responses against tumor cells [4–7]. By comparison with surgery, it hardly produce large wounds, nor does it cause higher toxicity in normal tissues and drug tolerance like chemotherapy in the treatment of tumor. However, the clinical efficacy of cancer vaccines based on tumor cell components, such as peptides and proteins, is limited by their poor immunogenicity and inefficient delivery to antigen presentation cells (APCs). Although, many cancer vaccines were applied with the immune-stimulatory agents, such as the programmed cell death protein 1 (PD-1), CpG oligonucleotide, etc. The clinical response rates remain low, especially in the hypoimmunogenic tumors. As we all know, pancreatic cancer is a more difficult type for immunotherapy as a low immunogenic tumor. Therefore, there are few effective cancer vaccines that can actively promote the pancreatic cancer treatment. Meanwhile, limited by the specific tumor antigens, the development of cancer vaccines was extremely difficult.

The field of cancer vaccines has seen significant advancements in recent years, with numerous studies focusing on different aspects of vaccine development and application [8–12]. These advances have been facilitated by the exploration of various types of cancer vaccines, including those targeting viral antigens, neoantigens, and other tumor-specific antigens [13–16]. One approach in cancer vaccine research involves targeting shared antigens from viral infections linked to certain cancer types, such as Epstein–Barr virus and HPV. Studies have shown promising results with vaccines targeting viral antigens like Epstein–Barr nuclear antigen 1 and latent membrane proteins, as well as HPV-related antigens, demonstrating favorable antitumor efficacy and the potential to establish durable T cell memory in rapidly progressing HPV-positive tumors [17]. Another significant area of research is neoantigen cancer vaccines. Neoantigens, derived from nonsynonymous cell variants, are only expressed in tumor cells and can bypass thymus-negative selection, leading to robust neoantigen-specific T cell responses [14]. Advances in genomic and transcriptional profiling have made identifying putative neoantigens possible, although challenges remain in predicting which neoantigens can effectively induce neoantigen-specific T cell responses [18–20]. Cancer vaccines are being developed to stimulate antitumor immunity with tumor antigens delivered in various forms, such as cells, peptides, viruses, and nucleic acids [21–24]. These vaccines aim to overcome tumor heterogeneity and reverse the immunosuppressive microenvironment. Challenges in the field include single antigen targeting, weak immunogenicity, off-target effects, and impaired immune response [25–27]. Current research and reviews provide insights into the principle of action, components, classification, delivery methods, and progress of cancer vaccines, offering a broader perspective for future vaccine design.

Recent studies have shown that tumor antigens can be used for the design of cancer vaccines without the necessity to identify, parse, and reconstruct certain antigens [28–31]. A novel concept has been proposed that the whole antigen vaccine systems derived from tumor cells could be designed to conquer cancer treatments [32–34]. The tumor cells themselves are a huge antigen pool with countless tumor antigen epitopes, which can be used to activate antitumor immune response. Among them, more studies have focused on the application of tumor cell membrane as tumor antigens to stimulate APCs and further activate the antitumor immune response of T cells. In addition, the activation of T cells in immune response requires the involvement of costimulatory molecules. Some studies are based on this theoretical foundation, using genetic engineering to modify tumor cell membranes. By retaining antigen on the cell membrane and inducing the expression of costimulatory molecule CD80, T cells can be directly activated, thereby achieving antitumor immune response [35] (Figure 1). Of course, many studies combine tumor cell membranes that retain tumor antigens with other functional molecules to directly activate dendritic cells (DCs) for antigen presentation, leading to the activation of T cell immune response. These are two different approaches aimed at achieving the same goal [36–38]. A series of nanovaccines developed on this basis have been widely used in the treatment of different tumor cells and preclinical experiments.

Abstract

     Magnetic nanoparticles (MNPs) represent an advanced tool in the medical field because they can be modified according to biomedical approaches and guided by an external magnetic field in the human body. The first objective of this review is to exemplify some promising applications in the medical field, including smart drug-delivery systems, therapies against cancer cells, radiotherapy, improvements in diagnostics using magnetic resonance imaging (MRI), and tissue engineering. Complementarily, the second objective is to illustrate the mechanisms of action and theoretical foundations related to magnetoresponsive materials.

Introduction

     Magnetic nanoparticles (MNPs) are nanoscale particles (1–100 nm) that can be guided through an external magnetic field due to their superparamagnetic, ferrimagnetic, and ferromagnetic properties, which may provide features for biomedical applications. MNPs with superparamagnetic properties are of special interest because they exhibit strong magnetic interactions under an external magnetic field, which disappear once the external magnetic field is removed. This property allows for the design of ferrofluids, since MNPs can be stabilized in solutions because they do not present magnetic interactions when the external magnetic field is switched off; and allows for in vivo performance, as in (i) cell marking [1], (ii) drug systems guided by a magnetic field [2], (iii) image contrast agents [3], and (iv) as heat generators in hyperthermia treatments [4].

     MNPs generally contains two main parts: the core and the coating. The core presents a predominant quantum effect, which commonly incorporates magnetic elements such as Fe, Ni, or Co as well as their corresponding oxides, while the coating is responsible for stabilizing and protecting the core from the chemical effects of the medium [5]. The coating also plays an essential role since it provides specific properties and functions to the nucleus; for example, biocompatible natural polymers, such as chitosan or cellulose, work as cargo vectors of therapeutic agents to release in a controlled manner (Figure 1). Due to these characteristics, biomedical device synthesis has shown great interest in low toxic superparamagnetic MNPs, to avoid embolization or other secondary effects for the patient at the molecular or cellular level [6].

Conclusions

     MNPs, in combination with superconducting magnets, have begun to play an essential role in advances in areas such as MDDS, MRI, and hyperthermia, among other fields. Therefore, it is crucial to continue the synthesis of MNPs and de

Abstract

     The global occurrence of oral cancer has increased in recent years. Oral cancer diagnosed in the advanced stages results in morbidity and mortality. The use of technology may be beneficial for early detection and diagnosis, and thus help the clinician with better patient management. The advent of artificial intelligence (AI) has the potential to improve oral cancer screening. AI can precisely analyze an enormous dataset from various imaging modalities and provide assistance in the field of oncology. This review focused on the applications of artificial intelligence in the early diagnosis and prevention of oral cancer. A literature search was conducted in the PubMed and Scopus databases using the search terminology “oral cancer” and “artificial intelligence”. Further information regarding the topic was collected by scrutinizing the reference lists of selected articles. Based on the information obtained, this article reviews and discusses the applications and advantages of AI in oral cancer screening, early diagnosis, disease prediction, treatment planning, and prognosis. Limitations and the future scope of AI in oral cancer research are also highlighted.

Introduction

     The global occurrence of Oral cancer (OC) has increased in recent years, with Oral squamous cell carcinomas (OSCCs) counting for more than 90% of these cancers.[1] OSCCs are also the sixth most common malignancy in the world. In 2012, The World Health Organization reported 529 000 new cases of OC and 300 000 deaths due to OC each year.[2] Oral cancer diagnosed in the advanced stage results in morbidity and mortality. A crucial factor in providing successful treatment is the early detection of cancerous lesions. Inaccessible lesions and the late detection of cancers are associated with low survival, increased symptoms, and a higher treatment cost.[3] Early diagnosis can increase the survival rate to 75–90%.1,4

Early detection includes the diagnosis of oral potentially malignant disorders and regular follow-ups. Oral potentially malignant disorders (OPMDs) have been defined as “any oral mucosal abnormality that is associated with a statistically increased risk of developing oral cancer.”[5] OPMDs include oral leukoplakia, proliferative verrucous leukoplakia, erythroplakia, oral lichen planus, oral submucous fibrosis, palatal lesions in reverse smokers, oral lupus erythematosus, actinic keratosis, and dyskeratosis congenita. Newly included lesions in the recent classification are oral lichenoid lesion and oral chronic graft-versus-host disease.[5]

Conclusion

     In recent years, the use of AI for the diagnosis and prognosis of diseases has evolved. Previous studies have proved that ML produces accurate results for OC detection. It assists clinicians in diagnostic processes and minimizes inadvertent errors. However, previous studies based on deep learning (neural network) provided more accuracy with the early detection of OC as compared to machine learning. AI presents the opportunity to develop new techniques combined with traditional approaches to improve the accuracy of detection of OC and OPMDs, as well as to predict the course of the precancerous/cancerous lesions from retrospective data. Future research could consider the innovation of data fusion algorithms combining various modalities, such as clinical, radiological, histological, and molecular assessments, to support the early diagnosis and outcome estimation of the disease.

Abstract

1. Introduction

2. Fundamentals of nanotechnology

3. The role of nanotechnology in achieving food security

4. Nanotubes in food and agriculture

5. Nanosensors in food and agriculture

6. The challenges of nanotechnology in food and agriculture system

7. Conclusions

References

Abstract

     Ensuring food security in developing countries is highly challenging due to low productivity of the agriculture sector, degradation of natural resources, high post farming losses, less or no value addition, and high population growth. Researchers are striving to adopt newer technologies to enhance supply to narrow the food demand gap. Nanotechnology is one of the promising technologies that could improve agricultural productivity via nano fertilizers, use of efficient herbicides and pesticides, soil feature regulation, wastewater management, and pathogen detection. It is equally beneficial for industrial food processing with enhanced food production with excellent market value, elevated nutritional and sensing property, improved safety, and better antimicrobial protection. Nanotechnology can also reduce post-farming losses by increasing the shelf life with the aid of nanoparticles. However, further investigation is required to solve the safety and health risks associated with the technology.

Introduction

     The foremost universal challenge on our planet is the question of establishing food security for a rapidly increasing population in the world. Predictions show that food demand is likely to rise from 59 to 98% for the world population reaching 9 billion by 2050 (Duro et al., 2020). Despite an increase of the world population particularly in developing countries, the global food supply interrupted by the expenditure of bio-resources for production of energy, manufacturing chemicals, high post farming loss, less value addition, inefficient distribution and marketing systems, and other factors (Barrett, 2021; Sekhon, 2014). Farmers throughout the world will focus on using new innovations and technologies for enhancing the production of crops through intensive and extensive agriculture. The current efforts further boosted through the use of nano-modified stimulants and precision farming. Agricultural efficiency, soil improvement, secure water use, distribution of food in stores, and its quality are basic factors of securing food that may be improved via advances in nanotechnology research (Ashraf et al., 2021; Sastry et al., 2011).

     Newer technology that will increase the production and reduce food wastage is important to maintain sustainable living standards of the nation and improve food security. Nanotechnology can provide a path for producing foods with outstanding quality in highly improved workable form along with induction of nutrients bioavailability. Many research investigations are focusing on increasing the application of nanotechnology for the production of crop and food processing (Abobatta, 2018; Axelos and Van De Voorde, 2017; Dasgupta et al., 2015; Peters et al., 2016). Increment in articles, intellectual property, and patents in nano-agriculture-based food with fresh research tendencies in the processing of food, nutraceutical distribution, packing, quality control, and serviceable food is a highly expanding field in nanotechnology research (Dasgupta et al., 2015).

Conclusions

     Nanotechnology is a newly emerging, but highly expanding technology in many fields related to human activities and benefits worldwide. Its fascinating phenomena have been witnessed through several research findings that the nanoparticles and nanostructure improve various properties due to small size, larger surface area, and highly catalytic nature. Nanotechnology is crucial in achieving food security, especially in the agriculture sector. It can improve crop production by effective microbial, pest, and weed control with high economic value, security, and safety. It also plays a key role in food processing, food modification, stability, sensing, prolonged shelf life, minimized food losses, and provide safe food. Nanotechnology also minimizes post-harvest losses with better stability, safety, and packaging materials. Nanoparticles like Ag, Au, Zn, TiO2, ZnO, SiO2, and MgO commonly used in food processing, may also induce health risks due to easy penetration through the cell for adverse reactions within various humans, and animal organs as well as parts of plants. Such risks due to nanoparticles or nanocomposite could be controlled in future research via using greener synthesis and searching for easy and cheaper protocols for degradation and removal of existing nanomaterial from the sites of attack.

Abstract

Keywords

1. Introduction

2. MFNPs-based magnetoresponsive effects for biomedical applications

3. Conclusion and perspectives

Declaration of competing interest

Acknowledgments

References

Abstract

One of the most attractive features of magnetic ferrite nanoparticles (MFNPs) in biomedical application is that they can mediate external magnetic field to produce local magnetic field, magnetic thermal and magnetic force effects. These generated effects can later be utilized in the diagnosis and treatment of various diseases. The application performance is mainly determined by the nano-magnetism of MFNPs. Therefore, by modulating the magnetic properties, the improved magnetic resonance (MR) signals, magnetothermal, and magnetomechanical effects of the MFNPs can be achieved. In this review, we summarize the strategies used in the engineering of MFNPs to enhance MR imaging sensitivity and magnetic thermal conversion efficiencies. We will also discuss the detailed magnetoresponsive mechanism arising from the critical magnetic properties of MFNPs. Furthermore, we will highlight the recent progresses of the engineered MFNPs in biomedical applications, with emphasis in MR signal amplification, magnetothermal, and magnetomechanical response in biomedical applications.

1. Introduction

Since ancient Greek time, lodestone, i.e., bulk magnetite (Fe3O4), has been used in the treatment of common ailments. With the advancement in nano-biotechnology, the applications of such magnetic ferrite nanoparticles (MFNPs) have evolved, which extends to the modern biomedicine and clinical study [[1], [2], [3], [4], [5], [6], [7]]. As biological tissues are transparent to magnetic fields, the most attractive advantage of MFNPs in biomedical applications is that they can complete various functions triggered by invisible, tissue-penetrating magnetic field [8,9]. For instance, natural magnetotactic bacteria build iron oxide nanoparticles (IONPs) in a chain configuration to generate a permanent dipole in their cells so as to navigate toward their favorable habitats by sensing the Earth’s magnetic field [10]. These MFNPs possess good biocompatibility, which have already been used as imaging and therapeutic agents in preclinical and clinical settings [2,3,11]. As a result of the excellent magnetoresponsive properties of MFNPs, numerous exciting in vivo biomedical applications are created. These include imaging for non-invasive diagnosis of diseases [12], treatment of diseases with hyperthermia [13], magnetically driven nanorobots [14], magnetic activation of ion channel for cell signaling [15], manipulation of cell’s function and fate [16,17], and so on. Despite the good establishment of these concepts, there is a significant challenge in obtaining effective MFNPs that can achieve full functionality via the efficient conversion of the external energy of magnetic field. As such, due to their poor conversion ability, this greatly hinders their transition from bench to bed.

1- Introduction

2- pH-responsive polymeric nanocarriers

3- Temperature-responsive polymeric nanocarriers

4- Redox potential-responsive polymeric nanocarriers

5- Enzyme-responsive polymeric nanocarriers

6- Light-responsive polymeric nanocarriers

7- Dual-responsive polymeric nanocarriers

8- Future perspectives

9- Conclusions

References

Abstract

In the last decade, considerable attention has been devoted to the use of biodegradable polymeric materials as potential drug delivery carriers. However, bioavailability and drug release at the disease site remain uncontrollable even with the use of polymeric nanocarriers. To address this issue, successful methodologies have been developed to synthesize polymeric nanocarriers incorporated with regions exhibiting a response to stimuli such as redox potential, temperature, pH, and light. The resultant stimuli-responsive polymeric nanocarriers have shown tremendous promise in drug delivery applications, owing to their ability to enhance the bioavailability of drugs at the disease site. In such systems, drug release is controlled in response to specific stimuli, either exogenous or endogenous. This review reports recent advances in the design of stimuli-responsive nanocarriers for drug delivery in cancer therapy. In particular, the synthetic methodologies investigated to date to introduce different types of stimuli-responsive elements within the biomaterials are described. The sufficient understanding of these stimuli-responsive nanocarriers will allow the development of a better drug delivery system that will allow us to solve the challenges encountered in targeted cancer therapy.

Conclusions

In this review, it was focused on the most recent advances in the development of stimuli-responsive nanocarriers for drug delivery in cancer therapy. These smart drug delivery systems respond to the distinct changes in cancer cells, such as changes in pH gradient and elevated secretion of certain enzymes, rather than the conditions in normal cells. Targeting tumors with stimuliresponsive nanocarriers could not only increase the therapeutic benefit and minimize associated toxicity, but could also enhance the curative effect by specifically releasing the anti-cancer drug in a powerful precise mode, both temporally and spatially. The different approaches adopted to incorporate stimuli responsive elements within the polymeric nanocarriers were discussed, with a particular focus on polymeric materials with stimuliresponsive mechanisms in response to pH, redox potential, enzymes, temperature, and light. Despite the tremendous progress that has been made regarding the engineering of new stimuli-responsive materials, several challenges still remain to be addressed with respect to nanomedicine applications. For example, their biodegradability and biocompatibility profiles should be critically investigated prior to utilization in human clinical trials. Many of these systems have only been reported as an in vitro proof-of-concept and follow-up work in vivo preclinical models has been described for only a few. However, the translation of these stimuli-responsive delivery systems from the bench to the bedside might be facilitated, to some extent, if the regulatory requirements for human clinical trials are considered in light of the key features that render a biopolymer suitable for biomedical application, such as biocompatibility, biodegradability, high drug loading capability, programmable release, excellent in vivo stability, non-cytotoxicity, and ability to support effective targeting. Given the ongoing developments in the field of bionanotechnology along with the wide knowledge accumulated over recent years, it is convinced that the collaborative efforts of chemists, biologists, and medicinal and pharmaceutical scientists will revolutionize the design of responsive polymeric materials for cancer therapy to significantly improve both the quality and duration of the lives of patients with cancer.

1- A brief history of CRISPR

2- CRISPR/Cas9: biology, functionality, and applications

3- Therapeutic targets for ex vivo cell-based CRISPR/Cas9 genome editing

4- CRISPR therapy in clinical trials

5- Challenges and opportunities

References

Abstract

The recently developed CRISPR/Cas9 technology has revolutionized the genome engineering field. Since 2016, increasing number of studies regarding CRISPR therapeutics have entered clinical trials, most of which are focusing on the ex vivo genome editing. In this review, we highlight the ex vivo cell-based CRISPR/Cas9 genome editing for therapeutic applications. In these studies, CRISPR/Cas9 tools were used to edit cells in vitro and the successfully edited cells were considered as therapeutics, which can be introduced into patients to treat diseases. Considering a large number of previous reviews have been focused on the CRISPR/Cas9 delivery methods and materials, this review provides a different perspective, by mainly introducing the targeted conditions and design strategies for ex vivo CRISPR/Cas9 therapeutics. Brief descriptions of the history, functionality, and applications of CRISPR/Cas9 systems will be introduced first, followed by the design strategies and most significant results from previous research that used ex vivo CRISPR/Cas9 genome editing for the treatment of conditions or diseases. The last part of this review includes general information about the status of CRISPR/Cas9 therapeutics in clinical trials. We also discuss some of the challenges as well as the opportunities in this research area.

A brief history of CRISPR

The term CRISPR, or the clustered regularly interspaced palindromic repeats, was first used by Jansen et al., in 2002 to describe a novel family of repetitive DNA sequences presented in the genomes of prokaryotes [1]. These unusual repeated sequences were first detected in 1987 by Nakata et al. in Escherichia coli [2], and then recognized to be widespread in archaea and bacteria by Mojica et al., in 2000 [3]. After Jansen et al. identified the CRISPR-associated (Cas) genes in 2002 [1], Mojica et al. [4], Vergnaud et al. [5], and Bolotin et al. [6] revealed the exrtachromosomal and phage-associated origins of the spacers that separate the individual direct repeats, in 2005. Speculations about CRISPR arrays as immune memory and defense mechanism against virus envisions were then proposed [6,7], while the first experimental evidence for CRISPR/Cas system based adaptive immunity was reported by Horvath et al., in 2007 [8]. Following the series discoveries on the basic function and mechanism of CRISPR systems [9–12], in 2010 Moineau et al. demonstrated the Cas9 enzyme (formerly named Cas5, COG3513, Csn1 or Csx12), which is guided by spacer sequences, cleaves target DNA [13]. In 2012, Doudna, Charpentier et al. [14], and Siksnys et al. [15] proved that Streptococcus pyogenes and Streptococcus thermophiles Cas9 can be guided by CRSIPR RNAs (crRNAs) to cleave target DNA in vitro, through forming a double-strand break (DSB). In 2013, Zhang et al. [16], Church et al. [17], and Doudna et al. [18] independently demonstrated the applications of engineered CRISPR/Cas9 systems for genome editing in mammalian cells. This represented a giant leap forward in the genome engineering field and triggered a tremendous number of studies regarding the use of the CRISPR/Cas9 platform for eukaryotic gene editing in a wide range of species, including flies [19,20], zebrafish [21–24], frogs [25], mice [26–28], rats [26,29], pigs [30], and monkeys [31,32].

DNA methylation is an epigenetic factor, which plays important roles in embryo and many other diseases development. This factor determines gene expression, and when half of them have CpG islands, DNA methylation and its enzyme effectors have been under the vast studies. Whole genome DNA demethylation is a crucial step of embryogenesis and also cell fate determination in embryos. Therefore, demethylation agents were used as a tool for dedifferentiation and transdifferentiation. Although many of these efforts have been successful, but using this method gave us a vast spectral cell type which is confusing. In this article, we briefly reviewed DNA methylation, and its role in embryogenesis and gene expression. In addition to that, we introduce studies that used this action as a direct method in induction of stem cells and cell fate decision.

Introduction

It has been shown that, epigenetic factors determine transcriptome of cells. These factors determine cell functions and identities [1]. Cell fate determination is among the most significant aspects influenced by these factors [2]. DNA methylation occurring on cytosine in mammals is a major epigenetic factor, which its presence and absence may pose control over 50% of genes, especially developmental-related genes [3]. It has been hypothesized that; DNA demethylation is one of the most significant events in initiation of embryo development after fertilization, which has been named as Zygote Gene Activation (ZGA) [4]. This procedure induces expression of many stemness-related genes. Recently, many studies showed role of DNA demethylation and other epigenetic factors in changing cell fate [2]. It has been proved that, DNA demethylation could change fate of specific somatic cells to another somatic identity in a process named as transdifferentiation [5]. This phenomenon can reprogram somatic cells directly to stem cells [6]. DNA methylation enzyme effectors can establish new fates in differentiation process of stem cells as well [7]. Although it is believed that, disruption of epigenome or reactivation of developmental process in demethylation process could be a potential answer to these phenomena, but related findings have shown that, different roles of DNA demethylation are very confusing to determine. Furthermore, the basic concept and application of this field have been hindered because of its reproducibility and cell fate heterogeneity.

Since 1990s Malaysia aspired to make biotechnology and bioeconomy as her engines of economic growth to utlise the abundance of natural resources and biodiversity. The public sector plays an integral role in developing the sector and various incentives are in place for the private sector to be actively involved and to forge collaboration with the public sector. The country launched its National Biotechnology Policy in 2005 and later launched its National Bioeconomy Programme in 2010 to become the first country in South East Asia and second in Asia after China to have such an initiative. Malaysia is also very proactive in its biosafety law and regulations and has most of the related legal instrument in place. A lot of success has been recorded since the inception of the National Biotechnology Policy in terms of job creation, contribution to GDP through biobusinesses and investment from foreign companies, but the sector is not spared from challenges too. Due to the nature of the discipline that is multidisciplinary and that requires huge amount of investment, expertise and political will, there are a lot of barriers before the country emerges as a bioeconomy player. This paper discusses the public policies, initiatives and funding mechanisms in place in Malaysia that drive its research, development and commercialisation in the area of biotechnology and bioeconomy. The authors also discuss the challenges faced in Malaysia in implementing the policies.

The government’s biotechnology agenda: a historical background

The biotechnology initiatives in Malaysia commenced in mid 1990s where the government committed significant amount of investment, time and energy to make the country an international biotechnology hub. The biotechnology agenda was developed to capitalise Malaysia’s rich natural resources and biodiversity and turn them into biobusiness and wealth. Historically, Malaysia has been actively pursuing agricultural research through a number of world recognised research institutes to promote higher yield and better quality of rubber, oil palm and agricultural commodities [1]. The country also boasts a strong manufacturing sector with the presence of multinational companies. These were seen as a pre-requisite to build the biotechnology sector. The modus operandi to make the country a biotechnology hub might have changed due to change in leadership in early 2000, but the commitments remained strong which is seeing various initiatives being rolled out by the government to strengthened biotechnology and bioeconomy to make these the pillars of economic engine. In Malaysia, government institutes and agencies continue to play a central role in accelerating the biotechnology and bioeconomy agenda.

The use of conventional extraction method particularly chromatography-based method involves complex scale-up, batch operation, low capability in process integration, laborious processing cycles, require high energy input and high cost (9). On the other hand, the removal of cell debris by filtration or centrifugation may be challenging when processing high biomass load due to heterogeneous distribution of particle size and high viscosity (10). On top of that, a higher number of operational steps would incur higher overall cost owing to the loss of some amount of target compound in each processing stage. In response to the increasing market demand of biotechnological products, a more versatile and economical bioseparation process which can offer higher process throughput, shorter processing time and scalability is necessary to cope with the current ratelimiting downstream processes (11). However, it should be noted that the selection of an appropriate separation method is crucial in the downstream processing. Otherwise, inappropriate extraction technique could negatively affect the biofunctionality of target molecules and would therefore limit their application to a great extent (12). Alternatively, ATPS exhibits application potential with great technical and economic advantages in downstream processing. Several discrete stages which involve separation, concentration into smaller volume and purification can be reduced and substituted by ATPS, allowing the overall recovery process become more energy-efficient and cost-effective (3). Compared to the conventional method, the use of ATPS for bioseparation offers many advantages (2). These include simple process operation, rapid separation, high selectivity, low energy consumption and low cost (3,8,13e16). ATPS has also been able to offer a biocompatible environment for the isolation of biotechnological products due to the presence of high water content in both phases (1,17). The extremely low interfacial tension of this system (between 0.0001 and 0.1 dyne/cm) creates high interfacial contact area of the dispersed phases, which in turn, enhances the efficiency of the mass transfer (1). Chemical cost is considered one of the dominant cost factors for large-scale bioseparation process. The use of inexpensive phase components in ATPS would make the whole downstream processing more economical. Problem of downstream pollution may also be avoided by recycling the ATPS phase components (2). Extraction using ATPS is relatively rapid and the processing capacity of this technological simple technique is quite high. Due to the simplicity and reliability of scaling-up approach, the extent of ATPS extraction to industrial scale application is feasible and practical (8,11,13).

Influence of parameters on the partitioning behavior of ATPS

The selective distribution of the compounds to be separated between the two phases serves as the basis of partitioning in ATPS. In ATPS, the partition profile of the solutes depends on different physicochemical interactions between the biomaterial and the phase forming chemicals (18). Interactions such as van der Waals’ forces, hydrogen bond, electrostatic interactions, steric effects, hydrophobicity, biospecific affinity interactions as well as conformational effects between the phase components and the substances contribute to the partitioning of the particular substance (1). To achieve an effective bioseparation process, the partitioning behaviour of biomolecule in ATPS can be influenced by changing some of the dominating factors such as molecular weights and size of polymers, type and composition of phase component, type of ions in the system, the addition of neutral salts like NaCl, tie line length (TLL), system temperature and pH (3,5,13).

Actinomycetes are excellent sources for novel bioactive compounds, which serve as potential drug candidates for antibiotics development. While industrial efforts to find and develop novel antimicrobials have been severely reduced during the past two decades, the increasing threat of multidrug-resistant pathogens and the development of new technologies to find and produce such compounds have again attracted interest in this field. Based on improvements in whole-genome sequencing, novel methods have been developed to identify the secondary metabolite biosynthetic gene clusters by genome mining, to clone them, and to express them in heterologous hosts in much higher throughput than before. These technologies now enable metabolic engineering approaches to optimize production yields and to directly manipulate the pathways to generate modified products.

Actinomycetes as sources for novel drugs

For more than 70 years, actinomycetes (order Actinomycetales) have been recognized as important sources for bioactive natural compounds. From the roughly 18 000 known bioactive bacterial compounds, more than 10 000 were described from bacteria of the actinomycete genus Streptomyces [1]. Many well-known antibiotics, such as tetracycline, erythromycin, vancomycin, and streptomycin, originate from the secondary metabolism of actinomycetes. Beyond antibiotics, other medically useful natural products that were isolated from this group of bacteria include the immunosuppressant rapamycin, the anticancer agents doxorubicin and bleomycin, the anthelmintic avermectin, and the antifungal compound nystatin.

 Food-producing animals (e.g., cattle, chickens, pigs, and turkeys) are the major reservoirs for many foodborne pathogens such as Campylobacter species, non-Typhi 30 serotypes of Salmonella enterica, Shiga toxin-producing strains of Escherichia coli, and Listeria monocytogenes. The zoonotic potential of foodborne pathogens and their ability to produce toxins causing diseases or even death are sufficient to recognize the seriousness of 33the situation. Foodborne pathogens cause millions of cases of sporadic illness and chronic 34 complications, as well as large and challenging outbreaks in many countries and between countries. The magnitude of this problem is demonstrated by the significant proportion of the 1.5 billion diarrheal episodes in children less than 3 years of age that are caused by enteropathogenic microorganisms, which results in more than 3 million deaths per year (EFSA-ECDC, 2016). Surveys estimate that in the United States alone, bacterial enteric pathogens cause 9.4 million episodes of foodborne illness in humans, 55,961 hospitalizations, and 1,351 deaths each year (Scallan et al., 2011). However, it is estimated that the reported incidence of food-borne disease represents less than 1% to 10% of the real incidence (Scallan et al., 2011). The importance of food-producing animals as carriers of pathogenic bacteria is real; for example, beef is reported to be the vector of 7% of the 1.7 million cases of foodborne disease that was recorded during 1996 to 2000 in England and Wales (Anderson et al., 2009).