Development Of Drug Resistance Research Articles

Binding kinetics of highly mutated HIV-1 subtype C protease inhibition by Lopinavir and Darunavir in the face of altered conformational dynamics.

Highly mutated HIV-1 protease (PR) compromises the efficacy of lopinavir (LPV) and darunavir (DRV) used to formulate salvage regimens in HIV/AIDS management. Here, we report the kinetics of inhibition of lopinavir (LPV) and darunavir (DRV) on highly mutated South African HIV-1 subtype C PR obtained from clinical isolates. The wild-type and mutant South African HIV-1 subtype C PR were cloned and purified. Enzyme inhibition assays and fluorescence spectroscopy were utilized to determine the binding kinetics of LPV and DRV with the wild-type and mutant HIV-1 PR variants. Like DRV, the results of this study show that LPV has a mixed-type inhibition mechanism, which indicates the possibility of a second binding site on HIV-1 PR. Both LPV and DRV poorly inhibited the highly mutated HIV-1 PR variants and had a markedly increased dissociation rate cons bound to the mutant variants compared to the wild type. The fast dissociation of these inhibitors translated into a short residence time of the inhibitor bound to the mutant HIV-1 PR variants. Fluorescent spectroscopy showed that the changes in the tertiary structure of the mutant HIV-1 PR variants were associated with a more open conformation. This open conformation was associated with altered conformational dynamics, which may have resulted in the loss of tight binding of LPV and DRV. This study's findings provide insight into the mechanism of resistance to LPV and DRV by highly mutated HIV-1 PR and provide information supporting the use of binding kinetics measurement in understanding HIV-1 PR inhibitor drug resistance evolution.

Fibroblast growth factor receptor four inhibitor FGF401 improves the efficacy of trastuzumab in FGFR4-overexpressing breast cancer cells.

Breast cancer is the most common cancer among women. Among them, human epidermal growth factor receptor-positive (HER2+) breast cancer is more malignant. Fortunately, many anti-HER2 drugs are currently used in clinical treatments to increase patient survival. However, some HER2+ patients (~15%) still develop drug resistance after receiving trastuzumab treatment, leading to treatment failure. Using CCLE and METABRIC database analyses, we found that fibroblast growth factor receptor 4 (FGFR4) mRNA was highly detected in tumors from HER2+ breast cancer patients (p < .001) and was associated with poorer survival in breast cancer patients. Through retrospective immunohistochemical staining analysis, we detected higher expression of FGFR4 protein in breast cancer tissues collected from patients who were resistant to trastuzumab therapy compared with breast cancer patients who responded to treatment. An FGFR4 inhibitor (FGF401) effectively inhibits tumor growth in trastuzumab-insensitive patient-derived xenograft (PDX) tumor-bearing mice. For molecular mechanism studies, we demonstrated that HER2/FGFR4 protein complexes were detected on the cell membrane of the tumor tissues in these trastuzumab-insensitive PDX tumor tissues. After trastuzumab treatment in these drug-resistant breast cancer cells, FGFR4 translocates and enters the nucleus. However, trastuzumab-induced nuclear translocation of FGFR4/HER2-intracellular domain protein complex in trastuzumab-resistant cancer cells is blocked by FGF401 treatment. We believe that FGFR4 overexpression and complex formation with HER2 can serve as molecular markers to assist clinicians in identifying trastuzumab-resistant tumors. Our results suggest that FGF401 combined with trastuzumab as adjuvant therapy for patients with trastuzumab-resistant breast cancer may be a potential new treatment strategy.

Screening of an Antibacterial and Coagulation Multifunctional Peptide and Its Application in Wound Healing.

As a new type of drug developed in the era of antibiotics, antimicrobial peptides (AMPs) have great potential in antibacterial, antiviral and antitumor aspects. Different from the principle of action of antibiotics, AMPs have a better effect on resisting drug-resistant bacteria. We synthesized a peptide library using solid-phase synthesis and successfully screened the antimicrobial peptide RKGAK. AMPs provide effective defense against both bacteria and fungi. Research has found that the lowest inhibitory concentrations of RKGAK against S. aureus, E. coli, and C. albicans are 75 μg/mL, 100 μg/mL, and 300 μg/mL. Not only that, this antibacterial peptide also has certain antiviral and coagulation effects. It is a multifunctional peptide that has great potential in many biomedical fields. It also has the advantages of being less likely to develop drug resistance, low hemolytic activity and low cytotoxicity. Loading the peptide into hydrogels has achieved good results in applications that promote wound healing.

Evaluating Antibiotic Resistance in the Bacterial Pathogens Isolated from Avian Species

The avian population plays a vital role as both a reservoir and sentinel for numerous pathogens. Antimicrobials are widely used for the treatment and control of infections in avian species. However, inappropriate selection, irrational use of these antibiotics may contribute to the development of drug resistance in population. This research evaluates the Antibiogram of Chlamydia psittaci, Klebsiella pneumonia, Escherichia coli, Staphylococcus epidermidis and Pasteurella multocida isolated from caged and free-living birds. Antimicrobial susceptibility was tested using the disk diffusion method on Mueller Hinton Agar, according to instructions by the Clinical and Laboratory Standards Institute. Chlamydia psittaci exhibited sensitivity to sulfamethoxazole (22mm), azithromycin (21mm), doxycycline (20mm) and gentamicin (15mm). In contrast, penicillin G, oxacillin, amoxicillin and chloramphenicol were ineffective. Escherichia coli shown azithromycin (20mm), doxycycline (16mm) and gentamicin (15mm) were most effective. Klebsiella pneumoniae demonstrated high sensitivity to sulfamethoxazole (20mm) and doxycycline (18mm), and showing complete resistance to oxacillin and rifampin. Pasteurella multocida exhibited the highest susceptibility to doxycycline (21mm), sulfamethoxazole (20mm), amoxicillin (20mm), chloramphenicol (19mm), norfloxacin (18mm) and gentamicin (16mm). Staphylococcus epidermidis demonstrated high sensitivity to norfloxacin (23mm), doxycycline (23mm), oxacillin (22mm), chloramphenicol (22mm), sulfamethoxazole (17mm) and gentamicin (15mm). The current study reveals sulfamethoxazole, doxycycline, and azithromycin were highly effective antibiotics against the bacterial pathogens infecting avian species. Penicillin G, amoxicillin, oxacillin, and ampicillin were resistant against the identified bacterial species.

Adaptive evolution of extensive drug resistance and persistence in epidemic ST11 KPC-producing Klebsiella pneumoniae during antimicrobial chemotherapy.

The global rise of carbapenem-resistant Klebsiella pneumoniae, including strains producing K. pneumoniae carbapenemase (KPC) types, poses a significant public health challenge due to their resistance to critical antibiotics. Treatment options for infections caused by KPC-producing K. pneumoniae (KPC-KP) are increasingly limited, particularly as these strains develop resistance to last-line antibiotics such as ceftazidime/avibactam and colistin. This study investigates the evolution of antibiotic resistance and persistence in a series of clonally related ST11 KPC-KP strains isolated from a single patient undergoing extended antimicrobial treatment. The patient, a 47-year-old male with a history of kidney transplantation, developed multiple KPC-KP lung infections during his hospital stay. Resistance to colistin and ceftazidime/avibactam emerged during treatment with these antibiotics. Key resistance mechanisms identified included the integration of ISKpn26 into mgrB gene, leading to mgrB inactivation and colistin resistance, and the emergence of novel blaKPC-2 variants (blaKPC-71 and blaKPC-179) that confer resistance to ceftazidime/avibactam. Despite the development of colistin resistance in a ceftazidime/avibactam-resistant KPC-KP strain following combination therapy, the patient's clinical condition significantly improved. Phenotypic assays showed that mgrB disruption in KPC-KP resulted in increased biofilm formation and higher susceptibility to phagocytosis. In mouse models, KPC-KP strains with mgrB disruption showed reduced virulence, increased lung colonization and persistence, and a lower inflammatory response, suggesting that mgrB disruption facilitates the transition from acute infection to colonization. This study highlights the complex interplay between antibiotic resistance and bacterial fitness, offering insights into why some patients experience clinical improvement despite severe drug resistance and incomplete bacterial clearance.

Microwave Assisted Synthesis and Evaluation of Thiazole Derivatives as M. tuberculosis and Antimicrobial Activity

In last few decades, though significant progress has been made in the treatment and control strategies of tubercular infections by introducing new diagnostic and monitoring tools and combination therapy, it still continues to be severe problem. Thus with the aim of developing novel molecule with improved potency for treating Mycobacterium tuberculosis H37Rv strain infections and with decreased probability of developing drug resistance, herein we report the synthesis of thiazole derivatives, starting from ethyl acetoacetate, by microwave organic reaction enhancement method (MORE) and results of investigations of their antimycobacterial and antimicrobial activities. Many compounds have shown promising activity while others were inactive.

Effects of surfactin stress on gene expression and pathological changes in Spodoptera litura

Spodoptera litura (S. litura) is a polyphagous pest of the family Lepidoptera, which causes damage and yields losses to many crops. The long-term use of chemical pesticides for control not only seriously threatens environmental health, but also causes S. litura to develop drug resistance. Therefore, there is an urgent need to develop environmentally safe and friendly biogenic pesticides. However, the mechanism of action of the secondary metabolite (surfactin) of Bacillus Vélezensis (B. vélezensis) on lepidopteran pests (S. litura) has not been reported yet. We found that several metabolites and genes in S. litura were affected by surfactin exposure. The expressions of the metabolites (protoporphyrinogen (PPO), gluconolactone (GDL), and L-cysteate) were significantly down-regulated while glutamate and hydroxychloroquine were significantly up-regulated. The expression levels of genes related to drug metabolism and detoxification, include the glutathione s-transferase (GST) gene family and acetaldehyde dehydrogenase (ALDH), and apoptosis-inhibiting genes (seven in absentia homolog 1(SIAH1)) were significantly decreased. In addition, pathological changes occurred in intestinal wall cells, Malpighian tubule cells, and nerve cells of S. litura under surfactin stress. Conclusively, our results suggest that surfactin induces an increase in reactive oxygen species (ROS) and damages S. litura cells. Furthermore, based on the integrated analysis of transcriptomic and metabolomic data, it is hypothesized that surfactin may also trigger neurotoxicity and cardiotoxicity in S. litura while hindering the insect’s detoxification processes. This study lays a foundation for further exploration of surfactin as a potential biopesticide.

CREATING CO-CRYSTAL SALTS: Synthesis, Crystal Structure, Hirshfeld Surface Analysis and Quantum Chemical Calculations of Pyrimethamine and their Potential Anti- SARS-Cov-2 Activity

Although the COVID-19 pandemic is over but the clouds of next pandemic can't be ignored and therefore, search for new effective and more safe drug candidates against SARS-CoV-2 is the need of time. The presented study investigates the potential for designing new least toxic but more effective drug candidates with enhanced therapeutic properties against SARS-CoV-2. Pyrimethamine, which was once a cornerstone in malaria treatment, has lost its effectiveness due to the development of drug resistance. Despite this, significant interest remains in modifying and repurposing the drug within medicinal chemistry. The presented study has demonstrated the synthesis of three novel organic co-crystal salts derived from pyrimethamine and the coformers homophthalic acid, chlorosalicylic acid, and 4-methoxy cinnamic acid via reflux method to obtain co-crystal salt I (PYM: HOM), II (PYM: CSAL), and III (PYM: MCIN), in a 1:1 stoichiometric ratio. Single-crystal X-ray diffraction, vibrational spectroscopy, and DSC/TGA analysis were used for the structural characterization and confirmation. Moreover, quantum chemical calculations were carried out to evaluate the structural and electronic properties of the binary complex of the co-crystals and crystal supercells. The interaction energies of the binary complexes were also computed to assess the relative stability of the crystal salts while comparing them with TGA/DTG data. As the COVID-19 pandemic has infected millions of people with mortality exceeding >1 million. There is an urgent need to find therapeutic agents that can help with virus eradication to prevent severe disease and deaths. These co-crystal salts were evaluated for their potential to inhibit SARS-CoV-2 propagation. Co-crystal salt II was observed as the most significant inhibitor at all the treatment points such as prophylactic, entry, and therapeutic (post-entry). Furthermore, pyrimethamine and coformers were also found to be effective at different targets as well. The identification of co-crystal salt II, comprising pyrimethamine and the coformers, shows promising antiviral activity against SARS-CoV-2, potentially laying the groundwork for the development of new drug candidates.

The putative error prone polymerase REV1 mediates DNA damage and drug resistance in Candida albicans

Antimicrobial-induced DNA damage, and subsequent repair via upregulation of DNA repair factors, including error-prone translesion polymerases, can lead to the increased accumulation of mutations in the microbial genome, and ultimately increased risk of acquired mutations associated with antimicrobial resistance. While this phenotype is well described in bacterial species, it is less thoroughly investigated amongst microbial fungi. Here, we monitor DNA damage induced by antifungal agents in the fungal pathogen Candida albicans, and find that commonly used antifungal drugs are able to induce DNA damage, leading to the upregulation of transcripts encoding predicted error-prone polymerases and related factors. We focus on REV1, encoding a putative error-prone polymerase, and find that while deleting this gene in C. albicans leads to increased sensitivity to DNA damage, it also unexpectedly renders cells more likely to incur mutations and evolve resistance to antifungal agents. We further find that deletion of REV1 leads to a significant depletion in the uncharacterized protein Shm1, which itself plays a role in fungal mutagenesis. Together, this work lends new insight into previously uncharacterized factors with important roles in the DNA damage response, mutagenesis, and the evolution of antifungal drug resistance.

Photoacoustic polydopamine-indocyanine green (PDA-ICG) nanoprobe for detection of senescent cells

Cellular senescence is considered an important tumour suppression mechanism in response to damage and oncogenic stress in early lesions. However, when senescent cells are not immune-cleared and persist in the tumour microenvironment, they can drive a variety of tumour-promoting activities, including cancer initiation, progression, and metastasis. Additionally, there is compelling evidence demonstrating a direct connection between chemo(radio)therapy-induced senescence and the development of drug resistance and cancer recurrence. Therefore, detection of senescent cells in tissues holds great promise for predicting cancer occurrence earlier, assessing tumour progression, aiding patient stratification and prognosis, and informing about the efficacy of potential senotherapies. However, effective detection of senescent cells is limited by lack of biomarkers and readout strategies suitable for in vivo clinical imaging. To this end, a nanoprobe composed of biocompatible polydopamine (PDA) nanoparticle doped with FDA-approved indocyanine green (ICG) dye, namely PDA-ICG, was designed as a contrast agent for senescence detection using photoacoustic imaging (PAI). In an in vitro model of chemotherapy-induced senescence, PDA-ICG nanoprobe showed an elevated uptake in senescent cells relative to cancer cells. In addition to its improved photostability, 2.5-fold enhancement in photoacoustic signal relative to ICG was observed. Collectively, the results indicate that the PDA-ICG nanoprobe has the potential to be used as a contrast agent for senescence detection of chemotherapy-induced senescence using PAI.

Genetic diversity of Plasmodium falciparum and Plasmodium vivax field isolates from the Nowshera district of Pakistan

BackgroundThe genetic diversity of malaria parasites contributes to their ability to adapt to environmental changes, develop drug resistance and circumvent the host immune system. This study aimed to analyse the genetic diversity of the Pfmsp1 and Pfmsp2 genes in Plasmodium falciparum and the Pvmsp-3α gene in Plasmodium vivax isolates from District Nowshera in Pakistan.MethodsBlood samples from 124 consenting patients with uncomplicated malaria presenting to different hospitals from the Nowshera district were collected between March and August 2019, representing 28 P. falciparum and 96 P. vivax isolates. The genomic DNA extracted from the isolates were subjected to nested PCR and allele-specific analysis. Pvmsp-3α amplified fragments were further treated with restriction fragment length polymorphism (RFLP)-based Hha1 restriction enzyme.ResultsOf the analyzed P. falciparum, 21 distinct alleles were detected, including 14 alleles for Pfmsp-1 and 7 alleles for Pfmsp-2. The sub-allelic families MAD20 (50%) of Pfmsp-1and FC27 (75%) of Pfmsp-2 were predominant. The multiplicity of infection (MOI) was calculated as 1.4 and 1.2 for Pfmsp-1 and Pfmsp-2, respectively, with an overall mean MOI of 1.34. In P. vivax, 4 allelic variants, Pvmsp-3α types A, B, C and D, were detected, while RFLP digestion of amplicons, detected 9 sub-allelic variants (A1-A4, B1, B2, C1, C2 and D1) at the Pvmsp-3α locus.ConclusionThis first ever report of molecular characterization of P. falciparum and P. vivax genotypes from District Nowshera, Pakistan reveals moderate to high allelic diversity in parasite population from District Nowshera, Pakistan.

Evaluation of molecular inclusion of azole antifungals by β-cyclodextrin using computational molecular approach

Fungal infection is an emerging global concern highlighting diseases like pneumonia, invasive candidiasis, chronic pulmonary aspergillosis, and meningitis. But major concern is development of antifungal drug resistance, necessitating novel pharmacological solutions. Azoles, particularly fluconazole and related compounds, are emphasized for their systemic efficacy via inhibition of CYP51, altering fungal cell membrane sterols. So, in this work we theoretically investigated the effect of complexation of azoles with β-cyclodextrin (β-CD), focusing on improving solubility and bioavailability. Using molecular docking, the interaction between azoles and β-CD is analyzed, predicting stability and binding energies. Docking studies elucidate specific binding modes and hydrogen bonding interactions, while PM3 calculations provide insights into electronic structures and properties such as HOMO–LUMO energy gaps, chemical potential, hardness, and electrophilicity. Our findings indicate that fluconazole-β-CD complexes, particularly FCZ-βCD2, exhibit stability and promising development potential due to low energy and favorable binding characteristics. The antifungal spectrum of all the inclusion complexes was predicted using the PASS web server which proved that that the complexes are more effective against fungal pathogens than bacterial pathogens. This preliminary theoretical study highlights β-CD’s potential as a carrier to enhance the therapeutic efficacy of antifungal drugs, addressing key challenges in treating fungal infections through improved solubility and bioavailability.

Understanding the Development of Compensatory Pathways in a Mutant Malaria Parasite Harbouring Hypomorphic Allele of Plant-Like Kinases.

One of the mechanisms for subverting the effect of drugs bythe malaria parasite is through rewiring of its transcriptome. The effect is more pronounced for target genes belonging to the multigene family. Plasmodium falciparum protein kinases belonging to the CDPK family are essential for blood stage development. As such, CDPKs are considered good targets for the development of anti-malarial compounds. The chemical genetics approach has been historically used to elucidate the function of protein kinases in higher eukaryotes. It requires the substitution of gatekeeper residue for another amino acid with a different side chain through genetic manipulation. Amino acid substitution at the gatekeeper position modulates the activity of a protein kinase and changes its susceptibility to a specific class of compounds known as bumped kinase inhibitors (BKIs) that help in the functional identification of the target gene. Here, we have exploited the chemical genetics approach to understand compensatory mechanisms evolved by a mutant parasite harboring a hypomorphic allele of cdpk1. Overall, our approach helps in identifying compensatory pathways that may be simultaneously targeted to prevent the development of drug resistance against individual kinases.

Advances in Artificially Designed Antibacterial Active Antimicrobial Peptides.

Antibacterial resistance has emerged as a significant global concern, necessitating the urgent development of new antibacterial drugs. Antimicrobial peptides (AMPs) are naturally occurring peptides found in various organisms. Coupled with a wide range of antibacterial activity, AMPs are less likely to develop drug resistance and can act as potential agents for treating bacterial infections. However, their characteristics, such as low activity, instability, and toxicity, hinder their clinical application. Consequently, researchers are inclined towards artificial design and optimization based on natural AMPs. This review discusses the research advancements in the field of artificial designing and optimization of various AMPs. Moreover, it highlights various strategies for designing such peptides, aiming to provide valuable insights for developing novel AMPs.

Gallic Acid Induces HeLa Cell Lines Apoptosis via the P53/Bax Signaling Pathway.

Cervical cancer is a type of cancer that originates from the endometrium and is more common in developed countries and its incidence is increasing day by day in developing countries. The most commonly prescribed chemotherapeutic drugs limit their use due to serious side effects and the development of drug resistance. For this reason, interest in new active ingredients obtained from natural products is increasing. This study aimed to reveal the apoptotic and antiproliferative effects of gallic acid and doxorubicin combination therapy against the HeLa cell line. We investigated the anti-cancer effects of doxorubicin and gallic acid in the human HeLa cervical cell line by using the MTT test, Nucblue staining for the identification of apoptotic cells due to nuclear condensation using fluorescent substance, and apoptotic markers P53 and Bax for the RT-PCR test. The highest cytotoxic effect obtained in the study, the highest increase in apoptotic induction, and a significant difference in P53/Bax levels were seen in the gallic acid/doxorubicin combination. Additionally, it was determined that gallic acid exhibited an effective cytotoxic effect on HeLa and HaCat cells within 48 and 72 h of application. The obtained findings show that the gallic acid/doxorubicin combination applied to HeLa cells may be an alternative treatment against both the cytotoxic effect size and the side effects of the chemotherapy agent.

Utilizing the T cell immune response to treat non-small cell lung cancer with CRISPR/Cas9 technology

Abstract. Non-small cell lung cancer (NSCLC) has the greatest incidence rate and fatality, among all the lung cancer types. Its treatment has been widely concerned by the research community. Currently, many inhibitors used in clinical practice face drawbacks such as increased drug resistance in cancer cells and failure, as well as relatively high cytotoxicity. They not only fail to have a long-term effect on the lesion may also cause certain damage to normal cells. Therefore, finding a treatment method that cancer cells cannot or are difficult to develop drug resistance, can have long-lasting effects, and has low cytotoxicity is currently the research direction of scholars. CRISPR/Cas9 technology, a widely used and convenient gene editing method, can modify the microenvironment of T cells or cancer cells as a therapy of NSCLC. This article aims to review the current mechanisms of using CRISPR/Cas9 technology to treat NSCLC looking forward to more treatment sites and methods in the future.

Abstract A009: Increasing the Sensitivity to sorafenib of Resistant Hepatocellular Carcinoma Cells with Bovine Lactoferrin through Ferroptosis

Abstract Hepatocellular carcinoma (HCC) remains one of the most lethal cancers worldwide, with a high mortality rate largely due to late-stage diagnosis and the limited efficacy of current therapeutic options. Sorafenib is one of the first-line therapies for advanced HCC. However, its effectiveness is often compromised by the development of drug resistance, significantly limiting its long-term therapeutic potential. This limitation has driven extensive research into alternative therapeutic strategies that can target this resistance, thereby improving the clinical outcomes of patients. Bovine Lf (bLf), which is functionally similar to human Lf (hLf), is safe for human use and has demonstrated anticancer properties, including synergistic effects with chemotherapy. In our study, we explored the role of bovine lactoferrin (bLf) as a potential agent to increase the sensitivity of sorafenib-resistant HCC cells. We developed sorafenib-resistant HCC cell lines through repeated exposure to the drug, with IC50 calculations confirming their increased drug tolerance. These resistant cells were subsequently treated with a combination of sorafenib and bLf. This combination significantly reduced cell viability, with a sorafenib IC50 below the level of the parental sensitive cell line. RNA-seq data revealed significant changes in gene expression profiles, highlighting the differential expression of both mRNAs and long noncoding RNAs (lncRNAs) among resistant, nonresistant, and resistant bLf-treated cells. Further insights were gained through gene set enrichment analysis (GSEA), which identified ferroptosis as a critical pathway significantly enriched in resistant cells treated with bLf and sorafenib vs sorafenib treatment only. Among the differentially expressed lncRNAs, LINC02015 was the most upregulated in resistant cells, and the most downregulated in resistant cells treated with bLf and sorafenib. This lncRNA has emerged as a potentially influencing ferroptosis in other pathologies. Notably, LINC02015 was found to be correlated with HMOX1, a gene with established roles in oxidative stress and iron metabolism. Based on these findings, we conducted a more focused analysis of the ATF3/HMOX1/GPX4 axis, a signaling pathway previously implicated in the regulation of ferroptosis and cellular responses to oxidative stress. Our data suggest that this axis could play a role in modulating lactoferrin-induced ferroptosis in the context of sorafenib resistance. In conclusion, our study provides compelling evidence that bovine lactoferrin (bLf) has the potential to serve as a novel therapeutic agent for overcoming sorafenib resistance in hepatocellular carcinoma by inducing ferroptosis. By potentially targeting the LINC02015/ATF3/HMOX1/GPX4 axis, bLf not only enhances the efficacy of sorafenib but also opens new avenues for the development of combinatory treatment strategies aimed at improving outcomes for HCC patients. Citation Format: Hury V Hernandez-Galdamez, Mireya De la Garza, Karla Rubio, Ivonne Ramirez-Diaz, Josue Guzman-Linares, Theresa Azcarraga Acosta, Guillermo Barreto, stefan günther, Teresita del niño Jesus Flores-Tellez, Mario A Aguilar-Chaparro, Pablo Muriel de la torre, Magda Reyes-López, Saul Villa-Treviño, Carolina Piña-Vazquez. Increasing the Sensitivity to sorafenib of Resistant Hepatocellular Carcinoma Cells with Bovine Lactoferrin through Ferroptosis [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: RNAs as Drivers, Targets, and Therapeutics in Cancer; 2024 Nov 14-17; Bellevue, Washington. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(11_Suppl):Abstract nr A009.

Mediator kinase inhibitors suppress triple-negative breast cancer growth and extend tumor suppression by mTOR and AKT inhibitors

Triple-negative breast cancers (TNBC) are treated primarily by chemotherapy and lack clinically validated therapeutic targets. In particular, inhibitors of the PI3K/AKT/mTOR pathway, abnormally activated in many breast cancers, failed to achieve clinical efficacy in TNBC due to the development of adaptive drug resistance, which is largely driven by the transcriptomic plasticity of TNBC. Expression of CDK8/19 Mediator kinases that control transcriptional reprogramming correlates with relapse-free survival and treatment failure in breast cancer patients, including TNBC. We now investigated how CDK8/19 inhibitors affect the growth of TNBC tumors and their response to mTOR and AKT inhibitors. In contrast to the effects of most anticancer drugs, all the tested human TNBC models (including patient-derived xenografts) responded to CDK8/19 inhibitors in vivo even when they did not respond in vitro. Furthermore, CDK8/19 inhibition extended the host survival of established lung metastases in a murine TNBC model, where the primary tumors were not significantly affected. CDK8/19 inhibitors synergized with an mTORC1 inhibitor everolimus and a pan-AKT inhibitor capivasertib in vitro and strongly potentiated these drugs in long-term in vivo studies. Transcriptomic analysis of tumors that responded or became adapted to everolimus revealed that drug adaptation in vivo was associated with major transcriptional changes in both tumor and stromal cells. Combining everolimus with a CDK8/19 inhibitor counteracted many of these changes and induced combination-specific effects on the expression of multiple genes that affect tumor growth. These results warrant the exploration of CDK8/19 Mediator kinase inhibitors as a new type of drugs for TNBC therapy.

Distinct evolutionary trajectories following loss of RNA interference in Cryptococcus neoformans

While increased mutation rates typically have negative consequences in multicellular organisms, hypermutation can be advantageous for microbes adapting to the environment. Previously, we identified two hypermutator Cryptococcus neoformans clinical isolates that rapidly develop drug resistance due to transposition of a retrotransposon, Cnl1. Cnl1-mediated hypermutation is caused by a nonsense mutation in a gene encoding an RNA interference (RNAi) component, ZNF3, combined with a tremendous transposon burden. To elucidate adaptive mechanisms following RNAi loss, two bioinformatic pipelines were developed to identify RNAi loss-of-function (LOF) mutations in a collection of 387 sequenced C. neoformans isolates. Remarkably, several RNAi-loss isolates were identified that are not hypermutators and have not accumulated transposons. To test whether these RNAi LOF mutations can cause hypermutation, the mutations were introduced into a nonhypermutator strain with a high transposon burden, which resulted in a hypermutator phenotype. To further investigate whether RNAi-loss isolates can become hypermutators, in vitro passaging was performed. Although no hypermutators were found in two C. neoformans RNAi-loss strains after short-term passage, hypermutation was observed in a passaged Cryptococcus deneoformans strain with an increased transposon burden. Consistent with a two-step evolution, when an RNAi-loss isolate was crossed with an isolate containing a high Cnl1 burden, F1 hypermutator progeny inheriting a high transposon burden were identified. In addition to Cnl1 transpositions, insertions of a gigantic DNA transposon KDZ1 (~11 kb) contributed to hypermutation in the progeny. Our results suggest that RNAi loss is relatively common (7/387, ~1.8%) and enables distinct evolutionary trajectories: hypermutation following transposon accumulation or survival without hypermutation.

Identifying how drug efflux mechanisms impact Acinetobacter baumannii evolutionary paths to ciprofloxacin resistance

Acinetobacter baumannii is a multi-drug resistant pathogen commonly found in clinical settings. This pathogen frequently uses efflux pumps to mitigate antibiotic treatment stress and eliminate the drug. When A. baumannii is exposed to antibiotics, it often develops mutations in the efflux pump regulator genes, causing an increase in efflux pump production. We hypothesize that efflux is a key pathway that leads to treatment failure in A. baumannii infections. The extent to which increasing drug efflux impacts other cellular functions remains unknown. To identify how efflux pump mutations impact growth, resistance, and evolvability, wildtype A. baumannii laboratory strain 17978UN, along with four mutants of this strain, each with a single nucleotide polymorphism (SNP) in an efflux pump regulator (adeL L341R, adeN I49N, adeR D23Y, and adeS R152S), were propagated in the presence of antibiotic and an efflux pump inhibitor to place selective pressure on the isolates. SNPs increase the production of efflux pumps; inhibiting efflux ability will determine if the effect of each SNP is nullified. All evolved populations demonstrated differences in fitness and antibiotic resistance in comparison to their respective ancestors; the extent of adaptation affecting each phenotype was highly dependent on the regulator that was mutated. Counterintuitively, efflux inhibitors also placed stress on wild-type A. baumannii which leads to antibiotic resistance. These results demonstrate that efflux regulator mutations can influence population adaptability and cause treatment failure. Understanding the role that different efflux systems play in treatment failure and drug resistance evolution will be instrumental in developing treatment strategies that hinder the development of antibiotic resistance.