Live biotherapeutics in cancer therapy.

Candida albicans is a semi-ubiquitous pathobiont that is known to significantly impact human health and wellbeing, causing a significant financial strain on the medical system. Due to increasing antifungal resistance, there is a growing need for novel fungal therapeutics to treat diseases caused by this fungus. The development and use of Live Biotherapeutic Products (LBPs) is an innovative and novel approach to potentially treating Candidiasis and other comorbidities associated with C. albicans infection. To evaluate their anti-pathogenic efficacy, it is necessary to understand the underlying mechanisms involved, via the use of biomimetic cell models. In this study, six LBPs were chosen to investigate their competitive inhibitory effect against C. albicans using a co-culture of Caco-2 cells and mucous-secreting HT29-MTX cells to mimic human gut epithelium. The LBP strains were supplied by Servatus Biopharmaceuticals and identified as SVT 01D1, SVT 04P1, SVT 05P2, SVT 06B1, SVT 07R1 and SVT 08Z1. Five out of the six LBPs showed a significant reduction in the adhesion of C. albicans and all six LBPs reduced C. albicans invasion in the co-culture cells to varying degrees. There was no significant difference between co-inoculation of C. albicans with the LBPs or pre-inoculation of LBPs before the addition of C. albicans. The potential of these LBPs as novel anti-fungal therapeutics for the treatment of C. albicans diseases can be further documented in clinical trials.

The use of live biotherapeutic products (LBPs), including single strains of beneficial probiotic bacteria or consortiums, is gaining traction as a viable option to treat inflammatory-mediated diseases like inflammatory bowel disease (IBD). However, LBPs’ persistence in the intestine is heterogeneous since many beneficial bacteria lack mechanisms to tolerate the inflammation and the oxidative stress associated with IBD. We rationalized that optimizing LBPs with enhanced colonization and persistence in the inflamed intestine would help beneficial bacteria increase their bioavailability and sustain their beneficial responses. Our lab developed two bioengineered LBPs (SBT001/BioPersist and SBT002/BioColoniz) modified to enhance colonization or persistence in the inflamed intestine. In this study, we examined colon-derived metabolites via ultra-high performance liquid chromatography-mass spectrometry in colitic mice treated with either BioPersist or BioColoniz as compared to their unmodified parent strains (Escherichia coli Nissle 1917 [EcN] and Lactobacillus reuteri, respectively) or to each other. BioPersist administration resulted in lowered concentrations of inflammatory prostaglandins, decreased stress hormones such as adrenaline and corticosterone, increased serotonin, and decreased bile acid in comparison to EcN. In comparison to BioColoniz, BioPersist increased serotonin and antioxidant production, limited bile acid accumulation, and enhanced tissue restoration via activated purine and pyrimidine metabolism. These data generated several novel hypotheses for the beneficial roles that LBPs may play during colitis.

Live biotherapeutic product (LBP), a type of biological product, holds promise for the prevention or treatment of metabolic disease and pathogenic infection. Probiotics are live microorganisms that improve the intestinal microbial balance and beneficially affect the health of the host when ingested in sufficient numbers. These biological products possess the advantages of inhibition of pathogens, degradation of toxins, and modulation of immunity. The application of LBP and probiotic delivery systems has attracted great interest to researchers. The initial used technologies for LBP and probiotic encapsulation are traditional capsules and microcapsules. However, the stability and targeted delivery capability require further improved. The specific sensitive materials can greatly improve the delivery efficiency of LBPs and probiotics. The specific sensitive delivery systems show advantages over traditional ones due to their better properties of biocompatibility, biodegradability, innocuousness, and stability. Moreover, some new technologies, including layer-by-layer encapsulation, polyelectrolyte complexation, and electrohydrodynamic technology, show great potential in LBP and probiotic delivery. In this review, novel delivery systems and new technologies of LBPs and probiotics were presented, and the challenges and prospects were explored in specific sensitive materials for LBP and probiotic delivery.

Recent developments in the understanding of the relationship between the microbiota and its host have provided evidence regarding the therapeutic potential of selected microorganisms to prevent or treat disease. According to Directive 2001/83/EC, in the European Union (EU), any product intended to prevent or treat disease is defined as a medicinal product and requires a marketing authorization by competent authorities prior to commercialization. Even if the pharmaceutical regulatory framework is harmonized at the EU level, obtaining marketing authorisations for medicinal products remains very challenging for Live Biotherapeutic Products (LBPs). Compared to other medicinal products currently on the market, safety assessment of LBPs represents a real challenge because of their specific characteristics and mode of action. Indeed, LBPs are not intended to reach the systemic circulation targeting distant organs, tissues, or receptors, but rather exert their effect through direct interactions with the complex native microbiota and/or the modulation of complex host-microbiota relation, indirectly leading to distant biological effects within the host. Hence, developers must rely on a thorough risk analysis, and pharmaceutical guidelines for other biological products should be taken into account in order to design relevant non-clinical and clinical development programmes. Here we aim at providing a roadmap for a risk analysis that takes into account the specificities of LBPs. We describe the different risks associated with these products and their interactions with the patient. Then, from that risk assessment, we propose solutions to design non-clinical programmes and First in Human (FIH) early clinical trials appropriate to assess LBP safety.

Introduction The FunHoMic project is a Marie Skłodowska-Curie Innovative Training Network comprising 13 PhD students, 8 academic partners and 3 industry partnersaimingto understand the interplay between fungi, hostsand microbiota to improve prevention and treatment of fungal infections. Importance About 2 billion people suffer fungal infections, which have a mortality rate close to that of malaria or breast cancer. Candida albicans has a high clinical and economic burden, making it of particular interest to the FunHoMic project. 70% of women experience at least one episode of vulvovaginal candidiasis (“thrush”) during their lifetime; 8% suffer recurring infections. C. albicans may live as a commensal but can cause symptoms when the fungus-host-microbiota equilibrium is disrupted. Infections by C. albicans have a significant clinical impact, with fatalities in severe cases. Many factors are associated with C. albicans infections; intensive care, neutropenic and diabetic patients are most at risk of systemic infection. Rising antifungal drug resistance has led to certain C. albicans infections having no treatment option. Aim The FunHoMic consortium combines projectson fungal pathogenesis, immunology, microbial ecology and’omics technologies to understand and exploit interactions between fungus, host and microbiota. Identification of novel bio markers on the fungal side such as genetic polymorphisms or on the host side such asmicrobiota profiles, metabolites and/or immune markers can lead to patient classification based on relative risk of infection. This could be the beginning of personalised management for fungal infections using preventive or therapeutic interventions like new antifungals, immune modulators or Live Biotherapeutic Products (LBPs). This project has received funding from the European Union’s Horizon 2020 research and innovation programme under the MarieSklodowska-Curie grant agreement No 812969.

HIV is a leading cause of morbidity and mortality in sub-Saharan Africa, where adolescent girls and young women are three times more likely to acquire HIV than their male counterparts. A key risk factor for HIV is bacterial vaginosis (BV), a condition characterised by the loss of beneficial Lactobacillus species and increased abundance of non-optimal, inflammatory bacteria. Although BV affects approximately 25% of women in sub-Saharan Africa, effective therapeutics are lacking. Live biotherapeutics containing optimal Lactobacillus spp. represent a promising strategy to improve BV treatment outcomes and reduce HIV infection risk. We isolated 181 vaginal Lactobacillus spp. from 25 BV-negative South African women and characterized 50 selected isolates. This led to the identification of live biotherapeutic candidates for African women with distinct genomes compared to isolates from other geographical regions. This study contributes to current knowledge of the characteristics that should be considered when screening novel isolates for this purpose.

Isothermal microcalorimetry (IMC) is a potent analytical method for the real-time assessment of microbial metabolic activity, which serves as an indicator of microbial viability. This approach is highly relevant to the fields of probiotics and Live Biotherapeutic Products (LBPs), offering insights into microbial viability and growth kinetics. One important characteristic of IMC is its ability to measure microbial metabolic activity separately from cellular enumeration. This is particularly useful in situations where continuous tracking of bacterial activity is challenging. The focus on metabolic activity significantly benefits both probiotic research and industrial microbiology applications. IMC's versatility in handling different media matrices allows for the implementation of viability assessments under conditions that mirror those found in various industrial environments or biological models. In our study, we provide a proof of concept for the application of IMC in determining viability and growth dynamics and their correlation with bacterial count in probiotic organisms. Our findings reinforce the potential of IMC as a key method for process enhancement and accurate strain characterization within the probiotic sector. This supports the broader objective of refining the systematic approach and methods used during the development process, thereby providing detailed insights into probiotics and LBPs.

Clostridioides difficile infection (CDI) is a major cause of healthcare- and antibiotic-associated diarrhea. While fecal microbiota transplantation (FMT) shows promise for recurrent CDI, its mechanisms and long-term safety are not fully understood. Live biotherapeutic products (LBPs) using predefined bacterial consortia offer an alternative option, but the rational design of LBPs remains challenging. Here, we employ a computational pipeline and three metagenomic datasets to identify microbial strains for LBPs targeting CDI. We constructed the CDI-related microbial genome catalog, comprising 3741 nonredundant metagenome-assembled genomes (nrMAGs), and identified multiple potential protective nrMAGs, including strains from Dorea formicigenerans, Oscillibacter welbionis, and Faecalibacterium prausnitzii. Importantly, some of these protective nrMAGs were found to play an important role in FMT success, and most top protective nrMAGs can be validated by various previous findings. Our results demonstrate a framework for selecting microbial strains targeting CDI, paving the way for the computational design of LBPs against other enteric infections.

Live biotherapeutics products (LBP) are a novel range of therapeutic options in medicine. In this review, authors discuss basic composition and mechanism of action of LBP, provide a comprehensive focused overview of published in vitro and in vivo studies on efficacy of LBP for prevention and treatment of infectious diseases such as viral (HIV, COVID-19), bacterial (C.difficile infection, bacterial vaginosis, multi-drug resistant organisms) and fungal (Candida) organisms. This review should be of interest to clinicians to understand the broad application of LBP in infectious diseases world beyond recurrent C.difficile infection and to researchers on unexplored prospects of LBP and the need for further investigation in this emerging field to improve its clinical application.

Engineered commensal Bacteroides thetaiotaomicron enables tumor-targeted delivery and dual-mode regulation in colitis-associated colorectal cancer.

The way biological systems are built and designed has been revolutionized by synthetic biology. Further enhancements like predictive modeling, optimization and systematic design of complex biological systems, is now possible due to integration of Artificial Intelligence into synthetic biology. This review shares insights on the role of AI in advancement of synthetic biology, including genome editing, metabolic pathway optimization and biological circuit design etc. AI-driven tools contribute to the increased efficiency and precision. Application of deep learning and machine learning has made it possible to make CRISPR-cas9, de novo protein design and gene circuit development more precise. However, there are still some persistent challenges, especially in curating high-quality biological datasets and bridging interdisciplinary gaps between computational and experimental scientists. Future perspectives focus on causal reasoning in AI models, integration of physics based algorithms, and promoting collaboration across disciplines to achieve breakthroughs in both synthetic biology and AI. By joining these fields, the transformative power of synthetic biology and AI can be unlocked and applied in the fields of medicine, biotechnology and environmental sustainability, pioneering a way for a new era of bioengineering.

In the 4 th century BC Ge Hong, recognized the utility of fecally originated products "yellow soup" in managing diarrheal disease [1] In Europe, the first recorded use of Fecal Microbiome Transplantation (FMT) took place in veterinary medicine. The procedure was carried out by Fabricius Acquapendente (1537-1619), an Italian anatomist and surgeon, who transferred gastrointestinal content from a healthy animal to a sick one.Transfaunation', became a widespread treatment for animals and described its use for restoring normal rumination in cattle and in the last century the process has been used in veterinary medicine as transfaunation [2]. It is in the past decade that interest has reignited the restoration of the microbiome as a means of treating C. difficile infection (CDI) recurrences. In this supplement various aspects of this management process are discussed from a variety of perspectives. Prior to the approvals of these 2 LBPs there were significant challenges posed by regulators in terms of regulations and analytical frameworks. These are elegantly highlighted by the Microbiome Therapeutics Innovation (MITG) and Dadan Barberio (10.3389/frmbi.2024.1441290). It was clear that existing regulatory frameworks for governing LBPs had notable gaps. These gaps included microbial identification, potency and bioburdens. The MITG lead collaborative approaches in engaging experts in this field to hold discussions with the FDA. These meetings discussed catalyzing improvements in LBP analytics and refreshing the regulatory landscape. It was clear that a multi-faceted approach to continued development LBPs is essential.Van Dijk et al discussed the challenges faced with expanding the evolution of LBP development and ensure product safety and effectiveness. Previous recent experiences of non-regulated fecal materials for transplantation were associated with infections leading to some deaths notably in immunocompromised patients. Thus, moving from traditional methods to developing safe products for use in fecal transplantation to live microbial products (LMP).. LMPs are sub-divided into specific subcategories and the product characteristics lead to the development of appropriate guidelines. LMP are in the GMPcompliant manufacturing guidance. However, for injectable LMPs the agencies do not have regulatory guidelines as sterility is required. Clearly this is inappropriate for LMPs. Safetyrelated quality analysis is often inappropriate. However, recent FDA guidance has detailed 29 pathogens which must be absent from currently approved LBPs. This list is updated as necessary. Van Overall Live Biotherapeutic Products hold significant promise as therapeutic or preventative options to an array of human diseases and conditions. The development of these products is subject to evolving regulatory including manufacturing processes and safety needs. As these become more defined LBPs follow on the work of Ge Hong and his "yellow soup".

Treatment of bacterial infections with broad-spectrum antibiotics is a strategy severely limited by the decreased ability of the perturbed resident microbiota to control expansion of antibiotic-resistant pathogens. Live biotherapeutic products (LBPs) could provide an alternative to antibiotics in infection control by restoring gut colonization resistance and controlling expansion of resistant strains, an important therapeutic need not being addressed with existing anti-infective drug modalities. We review opportunities and challenges in developing LBPs for multidrug-resistant organisms colonization and infection control, with a focus on commercial fecal microbiota transplantation-like products and defined bacterial consortia, and spanning considerations related to availability of models for rational drug candidate selection and dose regimen selection, good manufacturing practice, intellectual property, and commercial viability.

A healthy vaginal microbiome (VMB) is dominated by Lactobacillus spp. and provides the first line of defense against invading pathogens. Vaginal dysbiosis, characterized by the loss of Lactobacillus dominance and increase of microbial diversity, has been linked to an increased risk of adverse genital tract diseases, including bacterial vaginosis, aerobic vaginitis, vulvovaginal candidiasis, sexually transmitted infections, and pregnancy complications such as preterm birth. Currently, antibiotics and antifungals are recommended first-line treatments with high cure rates, but they also can lead to high recurrence and resistance development. As an alternative, lactobacilli have been utilized to restore the vaginal microbiota. In this review article, we discuss vaginal dysbiosis in various gynecological infections and potential interventions based on Live Biotherapeutic Products (LBPs) with a focus on those that use intravaginal treatment modalities to modulate the VMB. Based on these, we provide insights on key factors to consider in designing phenotypic and genotypic screens for selecting bacterial strains for use as vaginally administered microbiome-directed therapeutics. Lastly, to highlight current progress within this field, we provide an overview of LBPs currently being developed with published clinical trial completion for recurrent BV, VVC, and UTI. We also discuss regulatory challenges in the drug development process to harmonize future research efforts in VMB therapy.

Engineering probiotics for therapeutic applications: recent examples and translational outlook