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Advances in CRISPR-Cas9 for the Baculovirus Vector System: A systematic review
(2022-12) Sari-Ak, D; Alomari, O; Shomali, RA; Lim, J; Thimiri Govindaraj, Deepak B
The baculovirus expression vector systems (BEVS) have been widely used for the recombinant production of proteins in insect cells and with high insert capacity. However, baculovirus does not replicate in mammalian cells; thus, the BacMam system, a heterogenous expression system that can infect certain mammalian cells, was developed. Since then, the BacMam system has enabled transgene expression via mammalian-specific promoters in human cells, and later, the MultiBacMam system enabled multi-protein expression in mammalian cells. In this review, we will cover the continual development of the BEVS in combination with CRPISPR-Cas technologies to drive genome-editing in mammalian cells. Additionally, we highlight the use of CRISPR-Cas in glycoengineering to potentially produce a new class of glycoprotein medicines in insect cells. Moreover, we anticipate CRISPR-Cas9 to play a crucial role in the development of protein expression systems, gene therapy, and advancing genome engineering applications in the future.
AI and precision oncology in clinical cancer genomics: From prevention to targeted cancer therapies-an outcomes based patient care
(2022-05) Zodwa, Dlamini, Z; Skepu, Amanda; Kim, N; Mkhabele, M; Khanyile, R; Molefi, T; Mbatha, S; Setlai, B; Mulaudzi, T; Mabongo, M
Precision medicine is the personalization of medicine to suit a specific group of people or even an individual patient, based on genetic or molecular profiling. This can be done using genomic, transcriptomic, epigenomic or proteomic information. Personalized medicine holds great promise, especially in cancer therapy and control, where precision oncology would allow medical practitioners to use this information to optimize the treatment of a patient. Personalized oncology for groups of individuals would also allow for the use of population group specific diagnostic or prognostic biomarkers. Additionally, this information can be used to track the progress of the disease or monitor the response of the patient to treatment. This can be used to establish the molecular basis for drug resistance and allow the targeting of the genes or pathways responsible for drug resistance. Personalized medicine requires the use of large data sets, which must be processed and analysed in order to identify the particular molecular patterns that can inform the decisions required for personalized care. However, the analysis of these large data sets is difficult and time consuming. This is further compounded by the increasing size of these datasets due to technologies such as next generation sequencing (NGS). These difficulties can be met through the use of artificial intelligence (AI) and machine learning (ML). These computational tools use specific neural networks, learning methods, decision making tools and algorithms to construct and improve on models for the analysis of different types of large data sets. These tools can also be used to answer specific questions. Artificial intelligence can also be used to predict the effects of genetic changes on protein structure and therefore function. This review will discuss the current state of the application of AI to omics data, specifically genomic data, and how this is applied to the development of personalized or precision medicine on the treatment of cancer.
Application of drug repurposing-based precision medicine platform for leukaemia patient treatment
(2022-10) Kenmogne, Vanelle L; Nweke, EE; Takundwa, Mutsa M; Fru, PN; Thimiri Govindaraj, Deepak B
Drug resistance in leukaemia is a major problem that needs to be addressed. Precision medicine provides an avenue to reduce drug resistance through a personalised treatment plan. It has helped to better stratify patients based on their molecular profile and therefore improved the sensitivity of patients to a given therapeutic regimen. However, therapeutic options are still limited for patients who have already been subjected to many lines of chemotherapy. The process of designing and developing new drugs requires significant resources, including money and time. Drug repurposing has been explored as an alternative to identify effective drug(s) that could be used to target leukaemia and lessen the burden of drug resistance. The drug repurposing process usually includes preclinical studies with drug screening and clinical trials before approval. Although most of the repurposed drugs that have been identified are generally safe for leukaemia treatment, they seem not to be good candidates for monotherapy but could have value in combination with other drugs, especially for patients who have exhausted therapeutic options. In this review, we highlight precision medicine in leukaemia and the role of drug repurposing. Specifically, we discuss the several screening methods via chemoinformatic, in vitro, and ex vivo that have facilitated and accelerated the drug repurposing process.
Aptamer-Based Tumor-Targeted Diagnosis and Drug Delivery
(2022-07) Woldekidan, Haregewoin B; Woldesemayat, AA; Adam, G; Tafesse, Mesfin; Thimiri Govindaraj, Deepak B
Early cancer identification is crucial for providing patients with safe and timely therapy. Highly dependable and adaptive technologies will be required to detect the presence of biological markers for cancer at very low levels in the early stages of tumor formation. These techniques have been shown to be beneficial in encouraging patients to develop early intervention plans, which could lead to an increase in the overall survival rate of cancer patients. Targeted drug delivery (TDD) using aptamer is promising due to its favorable properties. Aptamer is suitable for superior TDD system candidates due to its desirable properties including a high binding affinity and specificity, a low immunogenicity, and a chemical composition that can be simply changed.Due to these properties, aptamer-based TDD application has limited drug side effect along with organ damages. The development of aptasensor has been promising in TDD for cancer cell treatment. There are biomarkers and expressed molecules during cancer cell development; however, only few are addressed in aptamer detection study of those molecules. Its great potential of attachment of binding to specific target molecule made aptamer a reliable recognition element. Because of their unique physical, chemical, and biological features, aptamers have a lot of potential in cancer precision medicine.In this review, we summarized aptamer technology and its application in cancer. This includes advantages properties of aptamer technology over other molecules were thoroughly discussed. In addition, we have also elaborated the application of aptamer as a direct therapeutic function and as a targeted drug delivery molecule (aptasensor) in cancer cells with several examples in preclinical and clinical trials.
Bioinformatics tools for gene and genome annotation analysis of microbes for synthetic biology and cancer biology applications
(Springer, 2021-08) Nweke, EE; Suleman, Essa; Du Plessis, M; Thimiri Govindaraj, Deepak B; Singh, V; Kumar, A
In this book chapter, we focus on application of genome annotation and analysis of microbes for synthetic biology and cancer biology research. We particularly emphasised on application of microbial genomics in synthetic biology and cancer biology. Finally, we delineated future perspective and potential route map for improving the microbial genome annotation and microbial genomics analysis. We infer that our future perspective strategies would assist in reshaping the genome annotation of microorganisms along with microbial genomics analysis. In addition, with better understanding on microbial genome annotation and microbial genomics analysis, we believe this would better enable synthetic biology and cancer biology applications.
Boar seminal plasma proteomic profiling for biomarker discovery
(2022-12) Mokwena, Pateswana W; Stoychev, Stoyan H; Buthelezi, Sindisiwe G; Naicker, Previn; Ramukhithi, FV; Lehloenya, KC
As an important technology in the swine industry, artificial insemination is of great significance in the preservation of elite breeds and the improvement of the utilisation rate of elite breeds, which makes the quality of preserved pig semen particularly critical. This abstract mainly evaluates a proteomic method to profile boar seminal plasma of different breeds and evaluate potential candidate biomarkers. It has considerable practical value, new technical methods, reasonable test steps, and important theoretical reference value for the test results. In this study, various sample preparation methods were compared (acetone precipitation [urea and SDS resuspension] vs non precipitated) to develop a standard workflow that can be used for large proteomics studies; the optimal method (non-precipitated) was then tested on a pilot study that was composed of seminal plasma from eight boars belonging to three different breeds (two Large White, three Kolbroek, and three Windsnyer). Briefly, proteins from seminal plasma were digested on an automated FlexDuo system using trypsin. The peptides were pefractionated into six fractions using the hydrophilic interaction liquid chromatography/strong anion exchange method. The fractionated peptides were then analysed on the Evosep One connected to a 6600 TripleTOF Sciex mass spectrometer. A data-independent acquisition method was used. Spectronaut software was used for protein identifications and differential protein analysis. A total of 1,314 peptides were identified across runs from 233 protein groups. The proteins identified covered a wide range of biological functions related to sperm function, such as AQN-1, AWN, and SOD. To mention a few candidate proteins of interest, superoxide dismutase, glutathione transferase, ß hexosaminidase, and the prostaglandin isomerase were diferentially expressed among breeds and also in individual boars within the same breed. The in-solution method used in this study is semi-automated and can be used for high throughput biomarker and validation studies.
Candida parapsilosis cell wall proteome characterization and effectiveness against hematogenously disseminated candidiasis in a murine model
(2023-03) Gong, X; Srivastava, V; Naicker, Previn; Khan, A; Ahmad, A
Candida parapsilosis poses huge treatment challenges in the clinical settings of South Africa, and often causes infections among immunocompromised patients and underweight neonates. Cell wall proteins have been known to play vital roles in fungal pathogenesis, as these are the first points of contact toward environments, the host, and the immune system. This study characterized the cell wall immunodominant proteins of pathogenic yeast C. parapsilosis and evaluated their protective effects in mice, which could add value in vaccine development against the rising C. parapsilosis infections. Among different clinical strains, the most pathogenic and multidrug-resistant C. parapsilosis isolate was selected based on their susceptibility towards antifungal drugs, proteinase, and phospholipase secretions. Cell wall antigens were prepared by ß-mercaptoethanol/ammonium bicarbonate extraction from selected C. parapsilosis strains. Antigenic proteins were identified using LC–MS/MS, where 933 proteins were found, with 34 being immunodominant. The protective effect of the cell wall immunodominant proteins was observed by immunizing BALB/c mice with cell wall protein extracts. After the immunization and booster, the BALC/c mice were challenged with a lethal dose of C. parapsilosis. In vivo results demonstrated increased survival rates and lower fungal burden in vital organs in the immunized mice compared to the unimmunized mice, thereby confirming the immunogenic property of cell wall-associated proteins of C. parapsilosis. Therefore, these results advocated the potential of these cell wall proteins to act as biomarkers for the development of diagnostic assays and/or vaccines against infections caused by C. parapsilosis.
Candida parapsilosis cell wall proteome characterization and effectiveness against hematogenously disseminated candidiasis in a murine model
(2023-03) Gong, X; Srivastava, V; Naicker, Previn; Khan, A; Ahmad, A
Candida parapsilosis poses huge treatment challenges in the clinical settings of South Africa, and often causes infections among immunocompromised patients and underweight neonates. Cell wall proteins have been known to play vital roles in fungal pathogenesis, as these are the first points of contact toward environments, the host, and the immune system. This study characterized the cell wall immunodominant proteins of pathogenic yeast C. parapsilosis and evaluated their protective effects in mice, which could add value in vaccine development against the rising C. parapsilosis infections. Among different clinical strains, the most pathogenic and multidrug-resistant C. parapsilosis isolate was selected based on their susceptibility towards antifungal drugs, proteinase, and phospholipase secretions. Cell wall antigens were prepared by ß-mercaptoethanol/ammonium bicarbonate extraction from selected C. parapsilosis strains. Antigenic proteins were identified using LC–MS/MS, where 933 proteins were found, with 34 being immunodominant. The protective effect of the cell wall immunodominant proteins was observed by immunizing BALB/c mice with cell wall protein extracts. After the immunization and booster, the BALC/c mice were challenged with a lethal dose of C. parapsilosis. In vivo results demonstrated increased survival rates and lower fungal burden in vital organs in the immunized mice compared to the unimmunized mice, thereby confirming the immunogenic property of cell wall-associated proteins of C. parapsilosis. Therefore, these results advocated the potential of these cell wall proteins to act as biomarkers for the development of diagnostic assays and/or vaccines against infections caused by C. parapsilosis.
Comparison of the antiviral activity of the microbicide candidate griffithsin and its tandemers derivatives against different modes of HIV-1 transmission
(2020-05) Alexander, Kabamba; Malatji, Kanyane; Mulaudzi, Takalani
Tandemers 2MG, 2MG3, 3MG and 4MG are derivatives of the potent anti-HIV-1 microbicide candidate griffithsin (GRFT). We compared these compounds anti-HIV-1 activity to GRFT using the viruses CAP206.08 and CAAN5342.A2 that have decreased sensitivity to this lectin. The 2MG and 2MG3 tandemers had similar activity to GRFT against cell-free and cell-associated viruses, while 3MG and 4MG were significantly more potent. Furthermore, the restoration of the 234N or 295N glycan in these viruses, known to increase sensitivity to GRFT, also increased sensitivity to 2MG and 2MG3, and not to 3MG and 4MG. In addition, GRFT resistant viruses generated in-vitro were equally resistant to 2MG and 2MG3 while they had considerably low resistance to 3MG and 4MG. Lastly, all five compounds showed increased inhibitory activity in seminal and vaginal simulants although the effect was more pronounced in the former. These data support further studies of tandemers as potential microbicides.
Covalent binding of human two-domain CD4 to an HIV-1 subtype C SOSIP.664 trimer modulates its structural dynamics
(2022-07) Tumba, NL; Naicker, Previn; Stoychev, Stoyan H; Killick, MA; Owen, GR; Papathanasopoulos, MA
The human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) mediates host cell infection by binding to the cellular receptor CD4. Recombinant Env bound to CD4 has been explored for its potential as an HIV vaccine immunogen as receptor binding exposes otherwise shielded, conserved functional sites. Previous preclinical studies showed an interchain disulphide linkage facilitated between Env and 2dCD4S60C generates an immunogenic complex that elicits potent, broadly neutralizing antibodies (bNAbs) against clinically relevant HIV-1. This study investigated conformational dynamics of 2dCD4WT and 2dCD4S60C bound to an HIV-1C SOSIP.664 Env trimer using hydrogen-deuterium exchange mass spectrometry. The Env:2dCD4S60C complex maintains key contact residues required for MHCII and Env/gp120 binding and the residues encompassing Ibalizumab's epitope. Important residues remaining anchored, with an increased flexibility in surrounding regions, evidenced by the higher exchange seen in flanking residues compared to Env:2dCD4WT. While changes in Env:2dCD4S60C dynamics in domain 1 were moderate, domain 2 exhibited greater variation. Lack of stability-inducing H-bonds in these allosteric sites suggest the improved immunogenicity of Env:2dCD4S60C result from exposed CD4 residues providing diverse/novel antigenic targets for the development of potent, broadly neutralizing Ibalizumab-like antibodies.
CSIR Synthetic Biology and Precision Medicine Centre Biofoundry Program: Development for first of its kind Biofoundry lab in Africa
(2023-03) Thimiri Govindaraj, Deepak B
Background: Global bio-foundry Alliance (GBA) has been established between countries including the UK, US, Japan, Singapore, China, Australia, Denmark, and Canada through 16 research institutions. Global bio-foundry Alliance plays the key role in the synthetic biology drive towards a new global bioeconomy that is accelerated by advanced technology innovation. Establishment of Biofoundry program in South Africa and in Africa will plan key scientific and the strategic role in promoting synthetic biology and precision medicine program in Africa. This would further enable bioeconomy and industrial development towards SME program. At our CSIR Synthetic Biology and Precision medicine Centre, we are currently establishing biofoundry lab that will implement various synthetic biology and precision medicine projects in South Africa. Methods: We are currently establishing two research components in the CSIR Synthetic Biology and Precision Medicine Centre Bio-foundry program which includes industrial synthetic biology and functional precision medicine program. We implement Biofoundry biodesign and biological engineering Design-Build-Test-Learn (DBTL) cycle into our industrial synthetic biology and functional precision medicine program. In our Industrial synthetic biology program, we are working on a) ValitaCHO: Development of superior CHO cell line system for hyper-burst protein expression system using directed evolution and synthetic biology approaches; b) Lactochassis: Designer microbes for industrial synthetic biology platform applications; In our Cancer Precision Medicine program: we are working on drug repurposing based drug sensitivity screening platform for B-cell malignancies and ovarian cancer treatment for South African patient cohort. Results : We are currently at the Design phase of the Design-Build-Test-Learn (DBTL) cycle in our industrial synthetic biology and functional precision medicine program. We have so far have progressed in generation of the preliminary data on ValitaCHO cell-line chemstress fingerprinting profiling. We are currently designing the directed evolution approach for generation superior CHO cell line. In the Lactochassis project, we are currently designing the computational biology based genome mapping for Lactochassis. In our precision medicine platform, we are currently progressing in design and build phase of platform where we have currently procured 770 cancer drugs for drug repurposing platform which can be applied for blood and ovarian cancer cohort. Conclusion: Using Bio-design DBTL cycle, we aim to implement our industrial synthetic biology and cancer precision medicine platform at CSIR Synthetic Biology and Precision Medicine Centre. These platforms will enable establishment of one of the first Biofoundry labs in Africa. Funding: This work is funded by ICGEB Early Career Grant, NRF, MRC and Parliamentary Grant.
Development of a multiplex HIV/TB diagnostic assay based on the microarray technology
(2023-09) Malatji, Kanyane; Singh, Advaita A; Thobakgale, C; Alexandre, Kabamba
Currently there are diagnostic tests available for human immunodeficiency virus (HIV) and tuberculosis (TB); however, they are still diagnosed separately, which can delay treatment in cases of co-infection. Here we report on a multiplex microarray technology for the detection of HIV and TB antibodies using p24 as well as TB CFP10, ESAT6 and pstS1 antigens on epoxy-silane slides. To test this technology for antigen-antibody interactions, immobilized antigens were exposed to human sera spiked with physiological concentrations of primary antibodies, followed by secondary antibodies conjugated to a fluorescent reporter. HIV and TB antibodies were captured with no cross-reactivity observed. The sensitivity of the slides was compared to that of high-binding plates. We found that the slides were more sensitive, with the detection limit being 0.000954 µg/mL compared to 4.637 µg/mL for the plates. Furthermore, stability studies revealed that the immobilized antigens could be stored dry for at least 90 days and remained stable across all pH and temperatures assessed, with pH 7.4 and 25 °C being optimal. The data collectively suggested that the HIV/TB multiplex detection technology we developed has the potential for use to diagnose HIV and TB co-infection, and thus can be developed further for the purpose.
Development of insect cell line using CRISPR technology
(2021-02) Nweke, EE; Thimiri Govindaraj, Deepak B
In this chapter, we delineated the methods of CRISPR technology that has been used for the development of engineered insect cell line. We elaborated on how CRISPR/Cas9 genome editing in Drosophila melanogaster, Bombyx mori, Spodoptera frugiperda (Sf9 and Sf21), and Mosquitoes enabled the use of model or non-model insect system in various biological and medical applications. Also, the application of synthetic baculovirus genome along with CRISPR/Cas9 vector system to enable genome editing of insect cell systems for treatment of communicable and non-communicable diseases.
Dose immunogenicity study of a plant-produced influenza virus-like particle vaccine in layer hens
(2022-06) Abolnik, C; Smith, Tanya; Wandrag, BDR; Murphy, M; Rautenbach, M; Olibile, O; O'Kennedy, Maretha M
Avian influenza poses one of the largest known threats to global poultry production and human health, but effective poultry vaccines can reduce infections rates, production losses and prevent mortalities, and reduce viral shed to limit further disease spread. The antigenic match between a vaccine and the circulating field influenza A viruses (IAV) is a critical determinant of vaccine efficacy. Here, an Agrobacterium tumefaciens-mediated transient tobacco plant (Nicotiana benthamiana) system was used to rapidly update an H6 influenza subtype virus-like particle (VLP) vaccine expressing the hemagglutininn (HA) protein of South African H6N2 IAVs circulating in 2020. Specific pathogen free White Leghorn layer hens vaccinated twice with =125 hemagglutinating unit (HAU) doses elicited protective antibody responses associated with prevention of viral shedding, i.e. hemaglutination inhibition (HI) mean geometric titres (GMTs) of =7 log2, for at least four months before dropping to approximately 5–6 log2 for at least another two months. A single vaccination with a 250 HAU dose induced significantly higher HI GMTs compared lower or higher doses, and was thus the optimal dose for chickens. Use of an adjuvant was essential, as the plant-produced H6 HA VLP alone did not induce protective antibody responses. Plant-produced IAV VLPs enable differentiation between vaccinated and infected animals (DIVA principle), and with sucrose density gradient-purified yields of 20,000 doses per kg of plant material, this highly efficacious, safe and economical technology holds enormous potential for improving poultry health in lower and middle-income countries.
Efficacy of a plant-produced clade 2.3.4.4 H5 influenza virus-like particle vaccine in layer hens
(2022-06) Abolnik, C; O’Kennedy, Martha M; Murphy, MA; Wandrag, BDR
Outbreaks caused by Goose/Guangdong H5 highly pathogenic avian influenza (HPAI)-lineage viruses continue to occur in unprecedented numbers throughout Eurasia, the Middle East and Africa, causing billions of dollars in economic losses and the deaths or destruction of hundreds of millions of poultry, and pose a zoonotic threat. Here, a recombinant virus-like particle (VLP) displaying the hemagglutinin protein of a clade 2.3.4.4b H5N8 HPAI strain was produced in tobacco plants (Nicotiana benthamiana) and its immunogenicity with four commercial adjuvants was compared in layer hens. After two immunizations with 250 hemagglutinating unit doses, hens that received intramuscular injections of H5 VLPs formulated with Emulsigen D, Emulsigen P or Montanide ISA 71VG seroconverted with hemagglutination inhibition geometric mean titres (GMTs) of 7.3 log2 (± 1.17), 8 log2 (± 1.08) and 7.9 log2 (±1.07), respectively, but the GMT of hens inoculated by eye drop with VLPs plus Carbigen only reached 2.05 log2 (± 1.64). The H5 VLP plus Emulsigen-P vaccinated hens and a sham-vaccinated group were then challenged with a high dose of the homologous H5N8 HPAI virus. Vaccinated hens were completely protected and showed no clinical signs, whereas the sham-vaccinated birds all died within 3–4 days. The average oropharyngeal shedding in vaccinated hens was reduced by 3,487-fold and 472-fold on days 2 and 3 post challenge, respectively, whereas average cloacal shedding was reduced by 2,360,098-fold and 15,608-fold on days 2 and 3, respectively, compared to the sham-vaccinated controls. No virus was detected in the vaccinated hens after day 8 post challenge, and the plant-produced H5 VLP vaccine completely prevented H5N8 HPAI virus transmission to eggs. This highly efficacious, safe and non-toxic plant-produced H5 VLP vaccine with DIVA (differentiation of infected from vaccinated animals) capability could be rapidly produced with a yield of at least 85,000 doses per Kg of plant leaf material.
Efficacy of a plant-produced infectious bronchitis virus-like particle vaccine inspecific pathogen-free chickens
(2023-10) Sepotokele, KM; O’Kennedy, Martha M; Hayes, MC; Wandrag, DBR; Smith, P; Abolnik, C
Infectious bronchitis (IB) Gammacoronavirus causes a highly contagious respiratory disease in chickens that is listed by the World Organisation for Animal Health (WOAH). Its high mutation ability has resulted in numerous variants against which the commercially available live or recombinant vaccines singly offer limited protection. Agrobacterium-mediated transient expression in Nicotiana benthamiana (tobacco) plants was used here to produce a virus-like particle (VLP) vaccine expressing a modified full-length IBV spike (S) protein of a QX-like IB variant. In a challenge study with the homologous live IB QX-like virus, VLP-vaccinated birds produced S protein-specific antibodies comparable to those produced by live-vaccinated birds seroconverting with mean geometric titers of 6.8 and 7.2 log2, respectively. The VLP-vaccinated birds had reduced oropharyngeal and cloacal viral shedding compared to an unvaccinated challenged control and were more protected against tracheal ciliostasis than the live-vaccinated birds. While the results appeared similar, plant-produced IB VLPs are safer, more affordable, easier to produce and update to antigenically match any emerging IB variant, making them a more suitable alternative to IBV control than live-attenuated vaccines.
Establishment of ex vivo drug sensitivity screening platform for leukaemia and multiple myeloma using a South African patient cohort
(2023-10) Kenmogne, VL; Takalani, AMT; Nweke, EE; Takundwa, Mutsa M; Fabian, June; Maher, Heather; Du Toit, Justin; Philip-Cherian, V; Fru, PF; Govindaraj, Deepak BT
Our objective is to develop a functional precision medicine platform designed to directly identify tailored drug regimens that target individual patient cancer cells and give benefit to the same donors by supporting clinical decision-making. We demonstrate our ex vivo drug sensitivity screening platform for precision medicine using Leukaemia and Multiple Myeloma samples from a South African patient cohort as proof of concept.
Establishment of ex vivo drug sensitivity screening platform for leukaemia and multiple myeloma using a South African patient cohort
(2023-10) Kenmogne, VL; Takalani, AMT; Nweke, EE; Takundwa, Mutsa M; Fabian, June; Maher, Heather; Du Toit, Justin; Philip-Cherian, V; Fru, PF; Govindaraj, Deepak BT
Our objective is to develop a functional precision medicine platform designed to directly identify tailored drug regimens that target individual patient cancer cells and give benefit to the same donors by supporting clinical decision-making. We demonstrate our ex vivo drug sensitivity screening platform for precision medicine using Leukaemia and Multiple Myeloma samples from a South African patient cohort as proof of concept.
Ex vivo drug sensitivity screening in multiple myeloma identifies drug combinations that act synergistically
(2022-03) Giliberto, M; Thimiri Govindaraj, Deepak B; Cremaschi, A; Skånland, SS; Gade, A; Tjønnfjord, GE; Schjesvold, F; Munthe, LA; Taskén, K
The management of multiple myeloma (MM) is challenging: An assortment of available drug combinations adds complexity to treatment selection, and treatment resistance frequently develops. Given the heterogeneous nature of MM, personalized testing tools are required to identify drug sensitivities. To identify drug sensitivities in MM cells, we established a drug testing pipeline to examine ex vivo drug responses. MM cells from 44 patients were screened against 30 clinically relevant single agents and 44 double- and triple-drug combinations. We observed variability in responses across samples. The presence of gain(1q21) was associated with low sensitivity to venetoclax, and decreased ex vivo responses to dexamethasone reflected the drug resistance observed in patients. Less heterogeneity and higher efficacy was detected with many combinations compared to the corresponding single agents. We identified new synergistic effects of melflufen plus panobinostat using low concentrations (0.1-10 nm and 8 nm, respectively). In agreement with clinical studies, clinically approved combinations, such as triple combination of selinexor plus bortezomib plus dexamethasone, acted synergistically, and synergies required low drug concentrations (0.1 nm bortezomib, 10 nm selinexor and 4 nm dexamethasone). In summary, our drug screening provided results within a clinically actionable 5-day time frame and identified synergistic drug efficacies in patient-derived MM cells that may aid future therapy choices.
In vitro study on efficacy of PHELA, an African traditional drug against SARS-CoV-2
(2022-06) Matsabisa, MG; Alexandre, Kabamba B; Ibeji, CU; Tripathy, S; Erukainure, OL; Malatji, Kanyane; Chauke, Sipho H; Okole, Blessed N; Chabalal, HB
In 2019, coronavirus has made the third apparition in the form of SARS-CoV-2, a novel strain of coronavirus that is extremely pathogenic and it uses the same receptor as SARS-CoV, the angiotensin converting enzyme 2 (ACE2). However, more than 182 vaccine candidates have been announced; and 12 vaccines have been approved for use, although, even vaccinated individuals are still vulnerable to infection. In this study, we investigated PHELA, recognized as an herbal combination of four exotic African medicinal plants namely; Clerodendrum glabrum E. Mey. Lamiaceae, Gladiolus dalenii van Geel, Rotheca myricoides (Hochst.) Steane & Mabb, and Senna occidentalis (L.) Link; as a candidate therapy for COVID-19. In vitro testing found that PHELA inhibited> 90% of SARS-CoV-2 and SARS CoV infection at concentration levels of 0.005 mg/ml to 0.03 mg/ml and close to 100% of MERS-CoV infection at 0.1 mg/ml to 0.6 mg/ml. The in vitro average IC50 of PHELA on SARS-COV-2, SARS-CoV and MERS-COV were ~ 0.01 mg/ml. Secondly in silico docking studies of compounds identifed in PHELA showed very strong binding energy interactions with the SARS-COV-2 proteins. Compound 5 showed the highest afnity for SARS-COV-2 protein compared to other compounds with the binding energy of - 6.8 kcal mol-1. Our data showed that PHELA has potential and could be developed as a COVID-19 therapeutic.

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