Pharmaceuticals from Microbes

Biomacromolecules produced by microorganisms have been employed in healthcare ever since ancient times as part of fermented products or natural remedies, but from the discovery of penicillin in 1928 by Alexander Fleming, it is impossible to conceive medicine without microbial products. In addition to antibiotics, microorganisms produce secondary metabolites currently employed as anti-inflammatory, immunosuppressant, and antitumoral drugs, among others. As with any other well-established drugs, undesirable side effects may occur with these compounds due to excessive systemic drug concentrations, and their pharmacological activity can be lost by the development of resistance in the target cells. Besides, many microbial drugs have intrinsic physicochemical properties that limit their application in healthcare such as low aqueous solubility, low bioavailability, acute toxicity, and fast systemic and pre-systemic degradation.

Here we review the critical aspects of innovative strategies for microbial products of high interest for academia and healthcare industry. In order to improve some of the current drug limitations, researchers have explored multiple advanced formulation approaches based on disruptive technologies. By means of new biomaterials and nanotechnology, it is possible to maximize the possibilities for functionalization and interfacing with the biological environment, a characteristic that leads to unique properties as drug delivery carriers. These approaches have resulted in improved pharmacological effects and pharmaceutical characteristics as compared to classical formulations, representing the dawn of a new era in microbial healthcare products.

Vaccines save millions of lives each year from various life-threatening infectious diseases, and there are more than 20 vaccines currently licensed for human use worldwide. Moreover, in recent decades immunotherapy has become the mainstream therapy, which highlights the tremendous potential of immune response mediators, including vaccines for prevention and treatment of various forms of cancer. However, despite the tremendous advances in microbiology and immunology, there are several vaccine preventable diseases which still lack effective vaccines. Classically, weakened forms (attenuated) of pathogenic microbes were used as vaccines. Although the attenuated microbes induce effective immune response, a significant risk of reversion to pathogenic forms remains. While in the twenty-first century, with the advent of genetic engineering, microbes can be tailored with desired properties.

In this review, I have focused on the use of genetically modified bacteria for the delivery of vaccine antigens. More specifically, the live-attenuated bacteria, derived from pathogenic bacteria, possess many features that make them highly suitable vectors for the delivery of vaccine antigens. Bacteria can theoretically express any heterologous gene or can deliver mammalian expression vectors harboring vaccine antigens (DNA vaccines). These properties of live-attenuated microbes are being harnessed to make vaccines against several infectious and noninfectious diseases. In this regard, I have described the desired features of live-attenuated bacterial vectors and the mechanisms of immune responses manifested by live-attenuated bacterial vectors. Interestingly anaerobic bacteria are naturally attracted to tumors, which make them suitable vehicles to deliver tumor-associated antigens thus I have discussed important studies investigating the role of bacterial vectors in immunotherapy. Finally, I have provided important discussion on novel approaches for improvement and tailoring of live-attenuated bacterial vectors for the generation of desired immune responses.

Infectious diseases caused by pathogenic microorganisms are one of the leading causes of mortality worldwide. For many of these diseases, prophylactic and therapeutic treatments are available in the form of vaccines and drugs. Novel discoveries in the pathophysiology and immunology of these diseases have led to the identification of contributing factors to the progress of these diseases. Our immune system puts forth a strong defense against these infections, but the microbes develop strategies to evade the immune system and survive inside the host. There is an ongoing hunt to look for potent therapeutic agents against these harmful bugs. These therapeutic drugs also need to be delivered effectively for long-lasting protection. This has led to the enhanced emphasis on the type of a suitable delivery system that can carry these agents inside the human body in its bioactive form. Hence, to achieve newer ways to deal with infection, we need better delivery systems as powerful tools for infection control and treatment.

Biodegradable and biocompatible polymeric particles such as poly(lactic acid) and poly(lactic-co-glycolic acid) have emerged as one of the efficient delivery systems for many life-saving drugs. These polymers offer several advantages such as targeted delivery, sustained release, and maintenance of bioactivity; it also leads to dose sparing by reducing the exposure of bioactive molecules in the circulation. The polymeric particles are being extensively studied in several applications as delivery systems due to their ability to exhibit a broad range of desirable properties. The present review focuses on the polymer-based particulate delivery system as a plausible solution to circumvent the shortcomings of conventional therapeutic and prophylactic systems, and it discusses some of the methods for their preparation as well as mechanisms of action against infection. Here we also review the cellular interaction of nanoparticles because this interaction influences the effectiveness of the particles. The present review aims at different preparation methods of poly-lactide/poly-lactide-co-glycolide-based particles, their properties as carriers of bioactive molecules, and applications of polymeric particle-based bioactive delivery systems against microbes with an emphasis on recent findings. This review sheds light on the latest applications of particle-based delivery systems attempting to provide an updated study about the field.

The remarkable physical and chemical properties of pullulan, especially the biodegradability, biocompatibility, and nontoxicity, have been exploited in the past few decades and adapted in order to design more efficient drug delivery systems. This polysaccharide itself and its derivatives, which possess more reactive functional groups generated by functionalization of pullulan, were able to form conjugates or complexes with a variety of drugs, especially with hydrophobic drugs. By modulating the hydrophilic-hydrophobic balance in the support macromolecule structure and favoring various types of physical interactions between drug and carrier, the researchers attempted to optimize the charging and subsequent transport of drugs to target cells such as liver cell receptors or cancer cells.

In this chapter, beside the pullulan-based systems with antibacterial, antifungal, antitumor, antioxidant, anti-inflammatory, immunomodulatory, antilipidemic, or antiglycemic properties, other pharmaceutical formulations potentially useful to treat heart or bone diseases were reviewed. All studies highlighted the versatility of pullulan derivatives to form micelles, films, hydrogels, microparticles, and nanoparticles. Also, the results from in vivo and in vitro tests of cytotoxicity and the profiles of drug release from these carriers were encouraging such that the usage of pullulan polysaccharides for the future medical applications remains an open field.

Development of microbial cell factories via application of synthetic biology, protein engineering for metabolic engineering has revolutionized the maximum use of microbial consortium for biosynthesis and structural alteration of valuable flavonoids. From a single enzyme expression to complex metabolic pathway, it has been possible to manipulate strains of Escherichia coli, Saccharomyces cerevisiae, Streptomyces, and Bacillus for target-based modification of compounds to industrial level in laboratory. Biotransformation, a biotechnological approach, can be applied to structurally modify and generate library of natural products such as flavonoid derivatives.

This chapter highlights the significance of engineered new molecules and biotransformation approaches used to generate flavonoids by the use of microbial platforms. Basically, E. coli has been engineered by expressing secondary metabolites post modifying enzymes, glycosyltransferases, O-methyl transferases, and prenyltransferases, in particular to generate the natural and nonnatural flavonol derivatives. Indigenously present cytoplasmic cofactors, coenzymes, and donor substrates are utilized by such enzymes for target-based chemical modifications. Engineering the central carbon flux pathway to enhance the flow of carbon toward target donor substrates and cofactors such as nucleotide diphosphate (NDP)-sugars, S-adenosyl methionine, dimethylallyl pyrophosphate, and other cofactors which enhanced the cytoplasmic pool while maximizing the biotransformation efficiency for level up production are discussed. Moreover, heterologous expression of different pathway genes from different organisms and engineering of glycosyltransferases and O-methyl transferases into bacterial host does help to generate nonnatural flavonol glycosides.

The invention of genetic engineering tools has given birth to a new type of pharmaceuticals known as biopharmaceuticals. These are the drug molecules that have therapeutic effects and are synthesised in biological cell systems. Drug like recombinant insulin is a prominent prototype example of biopharmaceutical which is commonly available in the market at cheap prices for diabetic patients, globally. The production of these therapeutic molecules differs from chemically synthesised low molecular weight drugs. Upstream and downstream processes altogether comprise the production process of biopharmaceuticals. The downstream processing costs 70% of the total production cost of a particular biopharmaceutical, largely contributed by expensive chromatographic techniques such as affinity, hydrophobic interaction, ion exchange and size exclusion. Although chromatography is a reliable and conventional approach to carry out single step purification of biopharmaceuticals, the columns are run in a series to increase the purification fold. This makes the process tedious, and problems like diffusional spreading and resolution are also observed with chromatography procedures. The concern is important as we aim to bring various biopharmaceuticals into market that can treat innumerable diseases at a cheap price.

The current chapter emphasises the process and technology related to the upstream process and the three chronological steps – initial recovery, purification and polishing – involved in downstream processing of biopharmaceuticals. The chapter encompasses the hurdles encountered in the downstream processing in particular with chromatography process that makes high-quality production of biopharmaceuticals an expensive affair thus making it difficult to reach the public. New technologies designed to offer faster and cheaper purification such as aqueous two-phase extraction system, and nano-magnetic-based antibodies separation system have been discussed further. Moreover, we have reviewed and emphasised the requirement of using combination of physical, mathematical, biological and computational approaches, which can help to design efficient production and purification systems for the ample, cheap and continuous market supply of this new category of drugs.

Doxorubicin and daunorubicin are notable members of the type II polyketide synthase family and clinically important cancer chemotherapeutic agents and are produced by a mutant strain S. peucetius ATCC 27952. They belong to the anthracycline-type antitumor drugs. Doxorubicin remains one of the most widely used antitumor drugs for the treatment of various cancers because of its broad spectrum of activity. As a result, numerous works have been carried to unravel the biosynthetic pathway and the underlying regulatory mechanisms to gain insight into the mechanisms of the genes involved. Consenquently, there is a need to develop an overproducing strain at the industrial scale, to produce doxorubicin as an anticancer drug. Therefore a significant amount of progress has been made in unraveling the bottlenecks in the pathway, manipulating the biosynthesis, improving production, and generating novel derivatives by engineering S. peucetius strain.