Emergence Of Resistance Mechanisms Research Articles

Epidermal growth factor receptor antagonists in colorectal cancer: emerging strategies for precision therapy.

The global prevalence of colorectal cancer highlights the need to enhance treatment strategies for improved patient outcomes. The pivotal role of epidermal growth factor receptor (EGFR) signaling in regulating cellular processes for this disease pinpoints its value as a therapeutic target, despite the emergence of resistance mechanisms over time. This review discusses the clinical evidence supporting the use of EGFR inhibitors in molecularly-selected patients based on molecular characteristics (notably BRAF V600E and KRAS G12C) including combination approaches targeting different points in in the signaling pathway, as well as strategies such as EGFR inhibitor rechallenge. The role of HER2 inhibitors and emerging approaches such as bispecific antibodies are also reviewed. Recently, inhibitors targeting the KRAS G12C variant have emerged, albeit with modest monotherapy activity compared to other tumor types, emphasizing the influence of histologic origins on the EGFR signaling pathway. Integration of EGFR inhibitors into precision medicine has facilitated tailored therapies addressing resistance mechanisms. Patient selection for EGFR inhibitor rechallenge guided by ctDNA findings is crucial, with ongoing investigations exploring novel combinations to enhance EGFR blockade, highlighting the transformative potential of precision medicine in shaping the future of mCRC treatment toward personalized and targeted approaches.

Discovery of 9H-pyrimido[4,5-b]indole derivatives as dual RET/TRKA inhibitors

Aberrant RET kinase signaling is activated in numerous cancers including lung, thyroid, breast, pancreatic, and prostate. Recent approvals of selective RET inhibitors, pralsetinib and selpercatinib, has shifted the focus of RET kinase drug discovery programs towards the development of selective inhibitors. However, selective inhibitors invariably lose efficacy as the selective nature of the inhibitor places Darwinian-like pressure on the tumor to bypass treatment through the selection of novel oncogenic drivers. Further, selective inhibitors are restricted for use in tumors with specific genetic backgrounds that do not encompass diverse patient classes. Here we report the identification of a pyrimido indole RET inhibitor found to also have activity against TRK. This selective dual RET/TRK inhibitor can be utilized in tumors with both RET and TRK genetic backgrounds and can also provide blockade of NTRK-fusions that are selected for from RET inhibitor treatments. Efforts towards developing dual RET/TRK inhibitors can be beneficial in terms of encompassing more diverse patient classes while also achieving blockade against emerging resistance mechanisms.

Abstract CT203: Multi-omic analysis of serial biopsies to inform biomarkers of sensitivity to olaparib and durvalumab in patients with metastatic BRCA-wildtype triple negative breast cancer (mTNBC)

Abstract Background: The phase 2 Adaptive Multi-Drug Treatment of Evolving Cancers (AMTEC) trial (NCT03801369) evaluated the efficacy of the combination of the PARP inhibitor Olaparib (Ola) and the PD-L1 inhibitor Durvalumab (Durva) in participants with BRCA-wildtype mTNBC. The combination was found to be effective with a median progression free survival (mPFS) of 5.5 months (AACR 2022). Here we report on updated biomarker analyses from paired biopsies (Bx) pre- and on-Ola therapy from 14 AMTEC participants. Methods: AMTEC participants undergo a pre-Ola Bx (Bx1), then one (28-day) cycle of Ola monotherapy with a repeat on-Ola Bx (Bx2) before adding Durva to Ola. Multi-omic profiling of DNA, RNA, and protein signals in Bx1 and Bx2 using Deep candidate gene sequencing, RNAseq, Nanostring Digital Spatial Profiling (DSP), and multiplex immunohistochemistry (mIHC) was correlated with clinical outcomes to identify predictors of Ola + Durva sensitivity, and adaptive resistance to PARPi therapy. Results: We identified over 30 biomarkers, with the optimal predictive value arising from Bx2. Key potential biomarkers were confirmed in an independent test set. Markers of good prognosis: • Basal Immune Activated (BLIA)/Basal Immune Suppressed (BLIS subtype) - A BLIA subtype on Bx2 was associated with a mPFS of 7.36 months compared to 1.71 months for BLIS/Luminal androgen receptor subtypes (p=0.001) • Multi-omic immune composite - A positive immune consensus signature (RNA, DSP, mIHC) on Bx2 was associated with a mPFS of 7.36 months compared to 2.29 months (p=0.005) • B cell activation signature - We developed a novel B-cell RNA activation signature that was highly predictive of response in Bx2, with mPFS of 8.54 and 1.04 months in signature-positive and signature-negative tumors, respectively (p=0.0004) Markers of poor prognosis: • AKT/MAPK pathway - AKT pathway gene mutations in Bx2 with concomitant protein phosphorylation of pathway members correlated with a shorter mPFS of 2.4 months compared to 7.13 months for no pathway mutation/activation (p=0.03). MAPK pathway protein phosphorylation in Bx2 was a negative prognostic factor, with mPFS of 2.17 months (p=0.004) • Angiogenesis - An upregulated Hallmark of Cancer angiogenesis RNA signature on Bx2 was associated with a shorter mPFS of 1.94 months compared to 7.95 months with no angiogenesis upregulation (p=0.0009) The following markers did not show significant association with survival outcomes on AMTEC: • PD-L1 positivity by IHC (defined as >1% Tumor Proportion Score, 22C3 antibody) • Tumor mutational burden • HRD, as measured by Rad51 foci or by mutations in HR pathway members Conclusions: Findings highlight the value of paired Bxs to identify predictive biomarkers of PARPi + immune checkpoint inhibitor sensitivity. The striking predictive value of the BLIA/BLIS signature warrants evaluation in a larger trial. Emerging resistance mechanisms justify AMTEC trial expansion to include PARPi + MEKi or PARPi + AKTi in biomarker selected patients, which is now in progress. Citation Format: Zahi I. Mitri, Allison L. Creason, Jayne M. Stommel, Daniel Bottomly, Jeong Youn Lim, SMMART Clinical Trials Program, Christopher L. Corless, Shannon McWeeney, Gordon B. Mills. Multi-omic analysis of serial biopsies to inform biomarkers of sensitivity to olaparib and durvalumab in patients with metastatic BRCA-wildtype triple negative breast cancer (mTNBC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT203.

High-Voltage Toxin'Roll: Electrostatic Charge Repulsion as a Dynamic Venom Resistance Trait in Pythonid Snakes.

The evolutionary interplay between predator and prey has significantly shaped the development of snake venom, a critical adaptation for subduing prey. This arms race has spurred the diversification of the components of venom and the corresponding emergence of resistance mechanisms in the prey and predators of venomous snakes. Our study investigates the molecular basis of venom resistance in pythons, focusing on electrostatic charge repulsion as a defense against α-neurotoxins binding to the alpha-1 subunit of the postsynaptic nicotinic acetylcholine receptor. Through phylogenetic and bioactivity analyses of orthosteric site sequences from various python species, we explore the prevalence and evolution of amino acid substitutions that confer resistance by electrostatic repulsion, which initially evolved in response to predatory pressure by Naja (cobra) species (which occurs across Africa and Asia). The small African species Python regius retains the two resistance-conferring lysines (positions 189 and 191) of the ancestral Python genus, conferring resistance to sympatric Naja venoms. This differed from the giant African species Python sebae, which has secondarily lost one of these lysines, potentially due to its rapid growth out of the prey size range of sympatric Naja species. In contrast, the two Asian species Python brongersmai (small) and Python bivittatus (giant) share an identical orthosteric site, which exhibits the highest degree of resistance, attributed to three lysine residues in the orthosteric sites. One of these lysines (at orthosteric position 195) evolved in the last common ancestor of these two species, which may reflect an adaptive response to increased predation pressures from the sympatric α-neurotoxic snake-eating genus Ophiophagus (King Cobras) in Asia. All these terrestrial Python species, however, were less neurotoxin-susceptible than pythons in other genera which have evolved under different predatory pressure as: the Asian species Malayopython reticulatus which is arboreal as neonates and juveniles before rapidly reaching sizes as terrestrial adults too large for sympatric Ophiophagus species to consider as prey; and the terrestrial Australian species Aspidites melanocephalus which occupies a niche, devoid of selection pressure from α-neurotoxic predatory snakes. Our findings underline the importance of positive selection in the evolution of venom resistance and suggest a complex evolutionary history involving both conserved traits and secondary evolution. This study enhances our understanding of the molecular adaptations that enable pythons to survive in environments laden with venomous threats and offers insights into the ongoing co-evolution between venomous snakes and their prey.

Antimicrobial and antibiofilm effect of cannabinoids from Cannabis sativa against methicillin-resistant Staphylococcus aureus (MRSA) causing bovine mastitis.

Antimicrobial resistance (AMR) poses a serious threat to human, animal, and plant health on a global scale. Search and elimination techniques should be used to effectively counter the spread of methicillin-resistant Staphylococcus aureus (MRSA) infections. With only a few novel drugs in clinical development, the quest for plant-based alternatives to prevent the spread of antibiotic resistance among bacteria has accelerated. Treatment of MRSA infections is challenging owing to rapidly emerging resistance mechanisms coupled with their protective biofilms. In the present research, we examined the antibacterial properties of ten plant-derived ethanolic leaf extracts. The most effective ethanolic leaf extract against MRSA in decreasing order of zone of inhibition, Cannabis sativa L. > Syzygium cumini > Murraya koenigii > Eucalyptus sp. > while Aloe barbadensis, Azadirachta indica, had very little impact. Mangifera indica, Curcuma longa, Tinospora cordifolia, and Carica papaya did not exhibit inhibitory effects against MRSA; hence, Cannabis was selected for further experimental study. The minimal inhibitory concentration (MIC) of Cannabis sativa L. extract was 0.25mgml-1 with 86% mortality. At a sub-MIC dosage of 0.125mgml-1, the biofilm formation was reduced by 71%. The two major cannabinoids detected were cannabidiol and delta-9-tetrahydrocannabinol (Δ9-THC), which were majorly attributed to substantial inhibitory action against MRSA. The time-kill kinetics demonstrated a bactericidal action at 4 MIC over an 8-20-h time window with a 90% reduction in growth rate. The results from SEM, and light microscopy Giemsa staining revealed a reduction in cells in the treated group with increased AKP activity, indicating bacterial cell membrane breakdown. These findings suggested cannabinoids may be a promising alternative to antibiotic therapy for bovine biofilm-associated MRSA.

Silver Nanoparticles as Antimicrobial Agents: Mechanisms, Challenges, and Applications

Silver nanoparticles (AgNPs) have garnered considerable attention for their potent antimicrobial properties and broad spectrum of applications in various fields. This review delves into the multifaceted realm of AgNPs as antimicrobial agents, aiming to provide a comprehensive understanding of their mechanisms of action, challenges, and diverse applications. The article begins by tracing the historical significance of silver as an antimicrobial agents and transitions into the contemporary role of AgNPs in modern applications. The synthesis of AgNPs is explored, encompassing diverse methods such as chemical, physical, and biological approaches. Particular emphasis is placed on green synthesis methods, which not only yield nanoparticles with controlled properties but also align with principles of environmental sustainability and biocompatibility. The review subsequently dissects the intricate mechanisms underpinning AgNPs antimicrobial process. The interaction of AgNPs with microbial cell membranes, generation of reactive oxygen species (ROS), and disruption of vital microbial processes collectively contribute to their potent bactericidal and fungicidal activities. However, the incorporation of AgNPs into various applications is not devoid of challenges. This article examines potential toxicity concerns, addressing issues related to stability, aggregation, and controlled release of nanoparticles, as well as the emergence of resistance mechanisms in microorganisms. Delving into the realm of applications, the review unveils AgNPs’ significance in biomedical and clinical settings, where they are employed in wound healing, medical devices, and infection control. Moreover, the environmental implications of AgNPs are explored, including their use in water and air purification, as well as potential roles in food and agricultural sectors. Looking forward, the review discusses emerging trends and suggests future research directions. Combinational therapies, integration with advanced materials, and exploration of AgNPs’ potential in addressing global antimicrobial resistance are outlined as promising avenues. In conclusion, this comprehensive review underscores the vital role of silver nanoparticles as versatile and potent antimicrobial agents, shedding light on their mechanisms, challenges, and multifarious applications across diverse sectors.

Abstract 1875: Enhanced lung cancer treatment using AOH1996, a potent PCNA inhibitor

Abstract AOH1996 targets a cancer-associated form of PCNA (caPCNA) by altering the protein-protein interface between caPCNA and its many binding partners. AOH1996 does this by inserting into the PCNA-interacting protein-box (PIP-box) pocket that is, in part, defined by the interdomain connecting loop (IDCL), the site of interaction between PCNA and most of its many binding partners. As a result, AOH1996 inhibits PCNA functions such as DNA replication, DNA repair, and transcription-replication conflict (TRC) resolution leading to cell cycle arrest and apoptosis. These effects are cancer specific and AOH1996 has little effect on non-cancerous cells even at 6-fold the effective dose in cancer cells. AOH1996 was developed in our lab and is currently in Phase I human clinical trials. Six patients have been enrolled in the trial and have not experienced AOH1996-related toxicities as the doses have been escalated to levels found to be efficacious in preclinical animal studies. Resistance to EGFR tyrosine kinase inhibitors (TKIs) poses a significant challenge in the treatment of non-small cell lung cancer (NSCLC) patients harboring EGFR activating mutations. Initially, EGFR TKIs like gefitinib, erlotinib, afatinib and osimertinib deliver remarkable responses, inducing tumor regression and improving overall survival. However, the emergence of resistance mechanisms, such as secondary EGFR mutations and bypass signaling pathway activation, hampers the efficacy of these drugs over time. The functional relationship between PCNA and EGFR has implications for the treatment of lung cancer patients with activating EGFR mutations. Membrane-localized EGFR modulates the PCNA-centric process of DNA replication through signaling pathways such as the Ras-Raf-Mek-Erk, PI3K/AKT, and PLCγ1/PKC signaling pathways. Nucleus-localized EGFR enhances PCNA stability by phosphorylating PCNA on tyrosine 211 (Y211), which prevents its degradation. In addition, phosphorylation at Y211 by EGFR results in decreased association of PCNA with MutS, which results in reduced mismatch repair (MMR). In this study, we combine AOH1996 and osimertinib as an innovative approach to treating NSCLC cancers with activating EGFR mutations and find that the drug combination: 1.) enhances the killing of NSCLC cell lines with activating EGFR mutations and acquired resistance to TKIs; 2.) enhances the destabilization of PCNA on chromatin; and 3.) causes changes in the localization and colocalization of PCNA and EGFR. We conclude that the combination of AOH1996 and osimertinib holds much potential as an improved treatment regimen for lung cancer patients with activating EGFR mutations. Citation Format: Robert G. Lingeman, Robert J. Hickey, Linda H. Malkas. Enhanced lung cancer treatment using AOH1996, a potent PCNA inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1875.

In vitro activity of ceftazidime-avibactam and comparators against OXA-48-like Enterobacterales collected between 2016 and 2020.

Resistance to carbapenems among Enterobacterales is often due to the production of enzymes that are members of the oxacillinases (OXA)-48-like family. These organisms can also be resistant to other classes of drugs and are difficult to identify and treat. This study evaluated the activity of the drug ceftazidime-avibactam (CAZ-AVI) and other comparator agents against a global collection of Enterobacterales that produce OXA-48-like enzymes. CAZ-AVI was active against blaOXA-48-like Enterobacterales, and only colistin and tigecycline were similarly active among the comparator agents, highlighting the limited treatment options against these organisms. Continued surveillance of the distribution of these OXA 48-like producing Enterobacterales and monitoring of resistance patterns along with the implementation of antimicrobial stewardship measures to guide antibiotic use and appropriate treatment are necessary to avoid drug resistance among these organisms.

Narrative review on advancing breast cancer treatment: harnessing the power of PD-1/PD-L1 inhibitors for improved patient outcomes.

Cancer immunotherapy has significantly advanced the field of oncology, providing novel therapeutic strategies for various malignancies, including breast cancer. The programmed cell death protein 1/programmed cell death-ligand 1 (PD-1/PD-L1) pathway is pivotal in immune regulation, and its inhibitors have demonstrated therapeutic benefits in diverse tumors. This review aims to critically examine the role, clinical efficacy, safety, and future directions of PD-1/PD-L1 inhibitors in breast cancer treatment, with a focus on pembrolizumab, nivolumab, and tislelizumab, and to elucidate the challenges and prospects in this dynamic field. A comprehensive literature search was conducted, adhering to Narrative Review reporting checklist for transparent reporting. Data from selected studies were qualitatively analyzed to synthesize key findings related to the mechanisms of action, clinical applications, and challenges of PD-1/PD-L1 inhibitors in breast cancer. PD-1 inhibitors have shown remarkable efficacy in various malignancies, including advanced triple-negative breast cancer (TNBC), where they have been investigated both in combination with chemotherapy and as neoadjuvant/adjuvant treatment. The exploration of these inhibitors in other breast cancer subtypes, such as human epidermal growth factor receptor-positive and hormone receptor-positive breast cancer, is ongoing. The review highlights the challenges in patient selection, management of immune-related adverse events (irAEs), and the emergence of resistance mechanisms. It underscores the need for ongoing research focusing on identifying reliable predictive biomarkers, elucidating mechanisms of resistance, and optimizing treatment strategies. PD-1/PD-L1 inhibitors hold substantial promise in advancing breast cancer treatment. This review provides critical insights and emphasizes the clinical importance of continued scientific exploration to refine patient selection criteria, improve treatment outcomes, and expand the applications of immunotherapy in breast cancer. Further research is imperative to overcome the existing challenges and realize the full therapeutic potential of these inhibitors in breast cancer and other malignancies.

Pseudomonasaeruginosa antimicrobial susceptibility profiles, resistance mechanisms and international clonal lineages: update from ESGARS-ESCMID/ISARPAE Group

ScopePseudomonas aeruginosa, a ubiquitous opportunistic pathogen considered one of the paradigms of antimicrobial resistance, is among the main causes of hospital-acquired and chronic infections associated with significant morbidity and mortality. This growing threat results from the extraordinary capacity of P. aeruginosa to develop antimicrobial resistance through chromosomal mutations, the increasing prevalence of transferable resistance determinants (such as the carbapenemases and the extended-spectrum β-lactamases), and the global expansion of epidemic lineages. The general objective of this initiative is to provide a comprehensive update of P. aeruginosa resistance mechanisms, especially for the extensively drug-resistant (XDR)/difficult-to-treat resistance (DTR) international high-risk epidemic lineages, and how the recently approved β-lactams and β-lactam/β-lactamase inhibitor combinations may affect resistance mechanisms and the definition of susceptibility profiles. MethodsTo address this challenge, the European Study Group for Antimicrobial Resistance Surveillance (ESGARS) from the European Society of Clinical Microbiology and Infectious Diseases launched the ‘Improving Surveillance of Antibiotic-Resistant Pseudomonas aeruginosa in Europe (ISARPAE)’ initiative in 2022, supported by the Joint programming initiative on antimicrobial resistance network call and included a panel of over 40 researchers from 18 European Countries. Thus, a ESGARS-ISARPAE position paper was designed and the final version agreed after four rounds of revision and discussion by all panel members. Questions addressed in the position paperTo provide an update on (a) the emerging resistance mechanisms to classical and novel anti-pseudomonal agents, with a particular focus on β-lactams, (b) the susceptibility profiles associated with the most relevant β-lactam resistance mechanisms, (c) the impact of the novel agents and resistance mechanisms on the definitions of resistance profiles, and (d) the globally expanding XDR/DTR high-risk lineages and their association with transferable resistance mechanisms. ImplicationThe evidence presented herein can be used for coordinated epidemiological surveillance and decision making at the European and global level.

High-level nitrofurantoin resistance in a clinical isolate of Klebsiella pneumoniae: a comparative genomics and metabolomics analysis.

Nitrofurantoin is a commonly used chemotherapeutic agent in the treatment of uncomplicated urinary tract infections caused by the problematic multidrug resistant Gram-negative pathogen Klebsiella pneumoniae. The present study aims to elucidate the mechanism of nitrofurantoin action and high-level resistance in K. pneumoniae using whole-genome sequencing (WGS), qPCR analysis, mutation structural modeling and untargeted metabolomic analysis. WGS profiling of evolved highly resistant mutants (nitrofurantoin minimum inhibitory concentrations > 256 mg/L) revealed modified expression of several genes related to membrane transport (porin ompK36 and efflux pump regulator oqxR) and nitroreductase activity (ribC and nfsB, involved in nitrofurantoin reduction). Untargeted metabolomics analysis of total metabolites extracted at 1 and 4 h post-nitrofurantoin treatment revealed that exposure to the drug caused a delayed effect on the metabolome which was most pronounced after 4 h. Pathway enrichment analysis illustrated that several complex interrelated metabolic pathways related to nitrofurantoin bacterial killing (aminoacyl-tRNA biosynthesis, purine metabolism, central carbohydrate metabolism, and pantothenate and CoA biosynthesis) and the development of nitrofurantoin resistance (riboflavin metabolism) were significantly perturbed. This study highlights for the first time the key role of efflux pump regulator oqxR in nitrofurantoin resistance and reveals global metabolome perturbations in response to nitrofurantoin, in K. pneumoniae.IMPORTANCEA quest for novel antibiotics and revitalizing older ones (such as nitrofurantoin) for treatment of difficult-to-treat Gram-negative bacterial infections has become increasingly popular. The precise antibacterial activity of nitrofurantoin is still not fully understood. Furthermore, although the prevalence of nitrofurantoin resistance remains low currently, the drug's fast-growing consumption worldwide highlights the need to comprehend the emerging resistance mechanisms. Here, we used multidisciplinary techniques to discern the exact mechanism of nitrofurantoin action and high-level resistance in Klebsiella pneumoniae, a common cause of urinary tract infections for which nitrofurantoin is the recommended treatment. We found that the expression of multiple genes related to membrane transport (including active efflux and passive diffusion of drug molecules) and nitroreductase activity was modified in nitrofurantoin-resistant strains, including oqxR, the transcriptional regulator of the oqxAB efflux pump. Furthermore, complex interconnected metabolic pathways that potentially govern the nitrofurantoin-killing mechanisms (e.g., aminoacyl-tRNA biosynthesis) and nitrofurantoin resistance (riboflavin metabolism) were significantly inhibited following nitrofurantoin treatment. Our study could help inform the improvement of nitrofuran derivatives, the development of new pharmacophores, or drug combinations to support the resurgence of nitrofurantoin in the management of multidrug resistant K. pneumouniae infection.

Anti-PD-1/PD-L1 therapy for colorectal cancer: Clinical implications and future considerations

Colorectal cancer (CRC) is the third most prevalent cancer in the world. The PD-1/PD-L1 pathway plays a crucial role in modulating immune response to cancer, and PD-L1 expression has been observed in tumor and immune cells within the tumor microenvironment of CRC. Thus, immunotherapy drugs, specifically checkpoint inhibitors, have been developed to target the PD-1/PD-L1 signaling pathway, thereby inhibiting the interaction between PD-1 and PD-L1 and restoring T-cell function in cancer cells. However, the emergence of resistance mechanisms can reduce the efficacy of these treatments. To counter this, monoclonal antibodies (mAbs) have been used to improve the efficacy of CRC treatments. mAbs such as nivolumab and pembrolizumab are currently approved for CRC treatment. These antibodies impede immune checkpoint receptors, including PD-1/PD-L1, and their combination therapy shows promise in the treatment of advanced CRC. This review presents a concise overview of the use of the PD-1/PD-L1 blockade as a therapeutic strategy for CRC using monoclonal antibodies and combination therapies. Additionally, this article outlines the function of PD-1/PD-L1 as an immune response suppressor in the CRC microenvironment as well as the potential advantages of administering inflammatory agents for CRC treatment. Finally, this review analyzes the outcomes of clinical trials to examine the challenges of anti-PD-1/PD-L1 therapeutic resistance.

Noncoding RNAs as an emerging resistance mechanism to immunotherapies in cancer: basic evidence and therapeutic implications.

The increasing knowledge in the field of oncoimmunology has led to extensive research into tumor immune landscape and a plethora of clinical immunotherapy trials in cancer patients. Immunotherapy has become a clinically beneficial alternative to traditional treatments by enhancing the power of the host immune system against cancer. However, it only works for a minority of cancers. Drug resistance continues to be a major obstacle to the success of immunotherapy in cancer. A fundamental understanding of the detailed mechanisms underlying immunotherapy resistance in cancer patients will provide new potential directions for further investigations of cancer treatment. Noncoding RNAs (ncRNAs) are tightly linked with cancer initiation and development due to their critical roles in gene expression and epigenetic modulation. The clear appreciation of the role of ncRNAs in tumor immunity has opened new frontiers in cancer research and therapy. Furthermore, ncRNAs are increasingly acknowledged as a key factor influencing immunotherapeutic treatment outcomes. Here, we review the available evidence on the roles of ncRNAs in immunotherapy resistance, with an emphasis on the associated mechanisms behind ncRNA-mediated immune resistance. The clinical implications of immune-related ncRNAs are also discussed, shedding light on the potential ncRNA-based therapies to overcome the resistance to immunotherapy.

Development of lipid nanoparticles with nystatin for an antifungal action

Fungal diseases currently affect about a quarter of the population worldwide. Fungi infections caused by Candida albicans have been described as a significant concern to public health. The spectrum of clinical diseases caused by this fungi species range between vulvovaginal candidiasis, oral candidiasis, candidemia and mucositis. The emergence of resistance mechanisms towards antifungal therapy greatly hampers successful management of illness and patient outcome. Nystatin, an antifungal drug, is categorized as a class IV of Biopharmaceutical Classification System, presenting low aqueous solubility and low intestinal permeability. Nowadays, the emerging platform of nanotechnology and lipid nanoparticles, notably solid lipid nanoparticles (SLN), has been subject to growing attention over recent past, owing to the promising properties of vectorization among a substantial variety of pharmaceutical drugs. Due to its hydrophobic proprieties, nystatin was encapsulated in SLN. Thus aiming to understand the relationship between the use of nanosystems and the improvement of the therapeutic effect. The aim of this work was to formulate SLN with nystatin by different methods (high speed homogenization and ultrasonication) with optimization of several parameters and formulation of 2 gels (one of them containing nanoparticles).
 Initially, 3 lipids were used: Compritol® 888 ATO, cetyl palmitate and Precirol® ATO 5 and, after the study of several parameters (size, encapsulation efficiency (EE) and polymorphic behaviour of the lipids), Precirol® ATO 5 was chosen as the lipid with the most satisfactory results. The results of the present work showed that the assay method of nystatin was linear, specific and presented repeatability. The average diameter of empty nanoparticles (NPs) and with drug (Precirol-NYS NPs) was, respectively, 306 nm and 260 nm and an EE of 67.8%. Regarding stability, SLN with drug proved to be more stable than SLN without drug. The polymer used for formulation of gels was the polymer commonly known by the trade name Carbopol® 940. The yield of 0.5% Carbopol® gel preparations and 0.5% Carbopol® gel + 10% Precirol-NYS NPs were 87.2% and 91.39%, respectively.

Ganetespib (STA-9090) augments sorafenib efficacy via necroptosis induction in hepatocellular carcinoma: Implications from preclinical data for a novel therapeutic approach

Sorafenib, a multikinase inhibitor, is a first-line treatment for advanced hepatocellular carcinoma, but its long-term effectiveness is limited by the emergence of resistance mechanisms. One such mechanism is the reduction of microvessel density and intratumoral hypoxia caused by prolonged sorafenib treatment. Our research has demonstrated that HSP90 plays a critical role in conferring resistance to sorafenib in HepG2 cells under hypoxic conditions and N-Nitrosodiethylamine-exposed mice as well. This occurs through the inhibition of necroptosis on the one hand and the stabilization of HIF-1α on the other hand. To augment the effects of sorafenib, we investigated the use of ganetespib, an HSP90 inhibitor. We found that ganetespib activated necroptosis and destabilized HIF-1α under hypoxia, thus enhancing the effectiveness of sorafenib. Additionally, we discovered that LAMP2 aids in the degradation of MLKL, which is the mediator of necroptosis, through the chaperone-mediated autophagy pathway. Interestingly, we observed a significant negative correlation between LAMP2 and MLKL. These effects resulted in a reduction in the number of surface nodules and liver index, indicating a regression in tumor production rates in mice with HCC. Furthermore, AFP levels decreased. Combining ganetespib with sorafenib showed a synergistic cytotoxic effect and resulted in the accumulation of p62 and inhibition of macroautophagy. These findings suggest that the combined therapy of ganetespib and sorafenib may offer a promising approach for the treatment of hepatocellular carcinoma by activating necroptosis, inhibiting macroautophagy, and exhibiting a potential antiangiogenic effect. Overall, continued research is critical to establish the full therapeutic potential of this combination therapy.

Carbapenem-Resistant and ESBL-Producing Enterobacterales Emerging in Central Texas.

Carbapenem-resistant Enterobacterales (CRE) are subject to intense global monitoring in an attempt to maintain awareness of prevalent and emerging resistance mechanisms and to inform treatment and infection prevention strategies. CRE and extended-spectrum beta-lactamase (ESBL)-producing Enterobacterales are not usually examined collectively in regards to their shared pool of resistance determinants. Here, we genetically and phenotypically assess clinical isolates of CRE and extended-spectrum beta-lactamase (ESBL)-producing Enterobacterales in the growing region of Central Texas, where CRE are emergent and occurrence of non-carbapenemase-producing-CRE (non-CP-CRE) infections is increasing. CRE (n=16) and ESBL-producing Enterobacterales (n=116) isolates were acquired from a regional hospital in Central Texas between December 2018 and January 2020. Isolates were assessed genetically and phenotypically using antibiotic susceptibility testing, targeted PCR, and whole genome sequencing. CRE infections are increasing in incidence in Central Texas, and Klebsiella pneumoniae is causing the majority of these infections. Moreover, K. pneumoniae sequence type (ST) 307 is commonly found among both non-CP-CRE and EBSL-producing strains. Isolates carry similar plasmids harboring the gene for the ESBL CTX-M-15 and belong to the global lineage, rather than the Texas lineage, of ST307. Antibiotic resistance profiles, sequence data, and clinical records suggest that porin mutations may promote the transition of ST307 isolates from ESBL-producing to non-CP-CRE. In addition to antibiotic resistance mechanisms, several CRE isolates harbor active colicinogenic plasmids, which might influence the competitiveness of these bacteria during patient colonization. K. pneumoniae of the global ST307 lineage is circulating in Central Texas and is responsible for both non-CP CRE and ESBL-producing Enterobacterales infections. Enhanced surveillance is needed to understand the possible routes for the emergence of non-CP-CRE from EBSL-producing strains.

Truncal Dynamics May Trump: Serial ctDNA to Predict Early Therapeutic Response.

Promising utility of using serial ctDNA in metastatic colorectal cancer to both refine patient selection, reduce toxicity due to chemotherapy, and to evaluate emerging resistance mechanisms may lead the way to novel therapeutic strategies. However, important questions remain in validating its use as a predictive biomarker of treatment response. See related article by Vidal et al., p. 379.

Evaluation of Dilution Susceptibility Testing Methods for Aztreonam in Combination with Avibactam against Enterobacterales.

As multidrug and pan-resistance among Enterobacterales continue to increase, there is an urgent need for more therapeutic options to treat these infections. New β-lactam and β-lactam inhibitor (BLI) combinations have a broad spectrum of activity, but those currently approved do not provide coverage against isolates harboring metallo-β-lactamases (MBL). Aztreonam (ATM) and avibactam (AVI) in combination (ATM/AVI; AVI at 4 μg/mL fixed concentration) provides a similarly broad range of activity while maintaining activity against MBL-producing isolates. The in vitro susceptibility testing of ATM/AVI by standard methods was evaluated during development. This study investigated the impact of nonstandard testing conditions on the activity of ATM/AVI as observed during broth microdilution testing as well as the equivalency between agar dilution and broth microdilution MIC values when testing a diverse panel of Enterobacterales (N = 201). Nonstandard test conditions evaluated included inoculum density, atmosphere of incubation, media pH, varied medium cation concentrations, incubation time, varied serum concentrations, testing in pooled urine instead of media, addition of blood to the media, and the presence of surfactant. Generally, apart from low pH and high inoculum density, nonstandard testing parameters did not affect ATM/AVI broth microdilution MIC values. Correlation of MIC values obtained by agar dilution and broth microdilution resulted in an essential agreement of 97.0% for all tested Enterobacterales. Variation of standard testing conditions had little impact on broth microdilution MIC values for ATM/AVI. The correlation between broth microdilution and agar dilution MICs suggests both methods are reliable for determination of ATM/AVI MIC values. IMPORTANCE Increasing antibiotic resistance and emergence of pan-resistant isolates threaten the ability to control infections and to provide many other medical interventions such as surgery and chemotherapy, among others. New therapies are required to control emerging resistance mechanisms, including the increase in metallo-β-lactamases. Some new antibiotic combinations provide coverage against highly resistant isolates but are unable to target organisms that produce metallo-β-lactamases. Aztreonam in combination with avibactam provides a broad spectrum of activity against highly resistant isolates that also targets metallo-β-lactamase-producing organisms. An important part of drug development is the ability for clinical labs to determine the susceptibility of isolates to the antimicrobial. This manuscript investigates the in vitro susceptibility testing of aztreonam/avibactam with nonstandard testing conditions and a correlation study between broth microdilution and agar dilution against clinical isolates encoding a variety of resistance mechanisms. Overall, aztreonam/avibactam was generally unaffected by changes in testing conditions and showed strong agar/broth correlation.

Transcriptional profiling of matched patient biopsies clarifies molecular determinants of enzalutamide-induced lineage plasticity

The androgen receptor (AR) signaling inhibitor enzalutamide (enza) is one of the principal treatments for metastatic castration-resistant prostate cancer (CRPC). Several emergent enza clinical resistance mechanisms have been described, including lineage plasticity in which the tumors manifest reduced dependency on the AR. To improve our understanding of enza resistance, herein we analyze the transcriptomes of matched biopsies from men with metastatic CRPC obtained prior to treatment and at progression (n = 21). RNA-sequencing analysis demonstrates that enza does not induce marked, sustained changes in the tumor transcriptome in most patients. However, three patients’ progression biopsies show evidence of lineage plasticity. The transcription factor E2F1 and pathways linked to tumor stemness are highly activated in baseline biopsies from patients whose tumors undergo lineage plasticity. We find a gene signature enriched in these baseline biopsies that is strongly associated with poor survival in independent patient cohorts and with risk of castration-induced lineage plasticity in patient-derived xenograft models, suggesting that tumors harboring this gene expression program may be at particular risk for resistance mediated by lineage plasticity and poor outcomes.

Acquisition of resistance to ceftazidime-avibactam during infection treatment in Pseudomonas aeruginosa through D179Y mutation in one of two blaKPC-2 gene copies without losing carbapenem resistance.

Ceftazidime/Avibactam (CAZ/AVI) is frequently used to treat KPC-producing Pseudomonas aeruginosa (KPC-PA) and Enterobacterales. CAZ/AVI resistance is driven by several mechanisms. In P. aeruginosa this mainly occurs through alteration of AmpC, porins, and/or efflux pump overexpression, whereas in Enterobacterales it frequently occurs through D179Y substitution in the active site of KPC enzyme. This aminoacid change abolishes AVI binding to the KPC active site, hence inhibition is impaired. However, this substitution also decreases KPC-mediated resistance to carbapenems (“see-saw” effect). The goal of this work was to characterize the in vivo acquisition of CAZ/AVI resistance through D179Y substitution in a KPC-PA isolated from a hospitalized patient after CAZ/AVI treatment. Two KPC-PA isolates were obtained. The first isolate, PA-1, was obtained before CAZ/AVI treatment and was susceptible to CAZ/AVI. The second isolate, PA-2, was obtained after CAZ/AVI treatment and exhibited high-level CAZ/AVI resistance. Characterization of isolates PA-1 and PA-2 was performed through short and long-read whole genome sequencing analysis. The hybrid assembly showed that PA-1 and PA-2A had a single plasmid of 54,030 bp, named pPA-1 and pPA-2 respectively. Each plasmid harbored two copies of the bla KPC-containing Tn4401b transposon. However, while pPA-1 carried two copies of bla KPC-2, pPA-2 had one copy of bla KPC-2 and one copy of bla KPC-33, the allele with the D179Y substitution. Interestingly, isolate PA-2 did not exhibit the “see-saw” effect. The bla KPC-33 allele was detected only through hybrid assembly using a long-read-first approach. The present work describes a KPC-PA isolate harboring a plasmid-borne CAZ/AVI resistance mechanism based on two copies of bla KPC-2-Tn4401b and D179Y mutation in one of them, that is not associated with loss of resistance to carbapenems. These findings highlight the usefulness of a fine-tuned combined analysis of short and long-read data to detect similar emerging resistance mechanisms.