Different Mechanisms Of Resistance Research Articles

Quantification of cerebrospinal fluid tumor DNA in lung cancer patients with suspected leptomeningeal carcinomatosis

Cerebrospinal fluid tumor-derived DNA (CSF-tDNA) analysis is a promising approach for monitoring the neoplastic processes of the central nervous system. We applied a lung cancer-specific sequencing panel (CAPP-Seq) to 81 CSF, blood, and tissue samples from 24 lung cancer patients who underwent lumbar puncture (LP) for suspected leptomeningeal disease (LMD). A subset of the cohort (N = 12) participated in a prospective trial of osimertinib for refractory LMD in which serial LPs were performed before and during treatment. CSF-tDNA variant allele fractions (VAFs) were significantly higher than plasma circulating tumor DNA (ctDNA) VAFs (median CSF-tDNA, 32.7%; median plasma ctDNA, 1.8%; P < 0.0001). Concentrations of tumor DNA in CSF and plasma were positively correlated (Spearman’s ρ, 0.45; P = 0.03). For LMD diagnosis, cytology was 81.8% sensitive and CSF-tDNA was 91.7% sensitive. CSF-tDNA was also strongly prognostic for overall survival (HR = 7.1; P = 0.02). Among patients with progression on targeted therapy, resistance mutations, such as EGFR T790M and MET amplification, were common in peripheral blood but were rare in time-matched CSF, indicating differences in resistance mechanisms based on the anatomic compartment. In the osimertinib cohort, patients with CNS progression had increased CSF-tDNA VAFs at follow-up LP. Post-osimertinib CSF-tDNA VAF was strongly prognostic for CNS progression (HR = 6.2, P = 0.009). Detection of CSF-tDNA in lung cancer patients with suspected LMD is feasible and may have clinical utility. CSF-tDNA improves the sensitivity of LMD diagnosis, enables improved prognostication, and drives therapeutic strategies that account for spatial heterogeneity in resistance mechanisms.

Resistance mechanisms of Saccharomyces cerevisiae against silver nanoparticles with different sizes and coatings

To investigate the underlying resistance mechanisms of Saccharomyces cerevisiae against Ag-NPs with different particle sizes and coatings, transcriptome sequencing (RNA-seq) technology was used to characterize the transcriptomes from S. cerevisiae exposed to 20-PVP-Ag, 100-PVP-Ag, 20-CIT-Ag and 100-CIT-Ag, respectively. The steroid biosynthesis was found as a general pathway for Ag-NPs stress responding, in which ERG6 and ERG3 were inhibited and ERG11, ERG25 and ERG5 were significantly up-regulated to resist the stress by supporting the later mutation and resistance and modulate drug efflux indirectly. The resistance mechanism of S. cerevisiae to 20-PVP-Ag seems different from that of 100-PVP-Ag, 20-CIT-Ag and 100-CIT-Ag. Under the 20-PVP-Ag, transmembrane transporter activity, transition metal ion homeostasis and oxidative phosphorylation pathway were main resistance pathways to enhance cell transport processes. While 100-PVP-Ag, 20-CIT-Ag and 100-CIT-Ag mainly impacted RNA binding, structural constituent of ribosome and ribosome pathway which can provide more energy to maintain the number and function of protein in cells. This study reveals the differences in resistance mechanisms of S. cerevisiae to Ag-NPs with different particle sizes and coatings, and explains several main regulatory mechanisms used to respond to silver stress. It will provide theoretical basis for the study of chemical risk assessment.

Impact of tumor microenvironment on efficacy of anti-CD19 CAR T cell therapy or chemotherapy and transplant in large B cell lymphoma

The phase 3 ZUMA-7 trial in second-line large B cell lymphoma demonstrated superiority of anti-CD19 CAR T cell therapy (axicabtagene ciloleucel (axi-cel)) over standard of care (SOC; salvage chemotherapy followed by hematopoietic transplantation) (NCT03391466). Here, we present a prespecified exploratory analysis examining the association between pretreatment tumor characteristics and the efficacy of axi-cel versus SOC. B cell gene expression signature (GES) and CD19 expression associated significantly with improved event-free survival for axi-cel (P = 0.0002 for B cell GES; P = 0.0165 for CD19 expression) but not SOC (P = 0.9374 for B cell GES; P = 0.5526 for CD19 expression). Axi-cel showed superior event-free survival over SOC irrespective of B cell GES and CD19 expression (P = 8.56 × 10–9 for B cell GES high; P = 0.0019 for B cell GES low; P = 3.85 × 10–9 for CD19 gene high; P = 0.0017 for CD19 gene low). Low CD19 expression in malignant cells correlated with a tumor GES consisting of immune-suppressive stromal and myeloid genes, highlighting the inter-relation between malignant cell features and immune contexture substantially impacting axi-cel outcomes. Tumor burden, lactate dehydrogenase and cell-of-origin impacted SOC more than axi-cel outcomes. T cell activation and B cell GES, which are associated with improved axi-cel outcome, decreased with increasing lines of therapy. These data highlight differences in resistance mechanisms to axi-cel and SOC and support earlier intervention with axi-cel.

Differences in metabolic transport and resistance mechanisms of Abemaciclib, Palbociclib, and Ribociclib.

Cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) play a crucial role in cancer treatment, particularly in breast cancer, and their mechanism of drug resistance is a topic of global interest in research. Hence, it is vital to comprehend the distinctions between various CDK4/6i, including their mechanisms of action and resistance mechanisms. This article aims to summarize the metabolic and transport variations as well as the differences in resistance among the three FDA-approved CDK4/6 inhibitors: Abemaciclib, Palbociclib, and Ribociclib. It also aims to discuss how these differences impact the effectiveness and safety of anticancer drugs. It was conducted in March 2023 to search PubMed, Embase, and Web of Science for literature related to this topic. Despite all being CDK4/6i, differences in their metabolism and transport were found, which are related to their chemical structure. Moreover, there are variations in preclinical pharmacology, pharmacokinetics, and clinical safety and efficacy of the different inhibitors. Genetic mutations, drug tolerance, and other factors may influence CDK4/6 resistance mechanisms. Currently, the resistance mechanisms differences of the three drugs remain largely unknown, and there are differences in the resistance mechanisms among them, necessitating further exploration and research.

Age-dependence of susceptibility to single and repeated deltamethrin exposure in pyrethroid-resistant Aedes aegypti strains

Monitoring insecticide resistance is crucial in disease-transmitting mosquitoes to allow assessment of viable candidate insecticides to use for control and to provide indication of changes in resistance. Insecticide resistance bioassays are typically performed on young female mosquitoes, yet disease is transmitted by older females, which may also have encountered insecticide multiple times during their adult life. If insecticide mortality rates increase with age directly, or indirectly via cumulative toxicity from repeated exposure, the strategy of testing young mosquitoes as the least susceptible cohort would be supported. We tested three hypotheses via examination of how age and cumulative exposure impact mortality rates to the pyrethroid deltamethrin in strains of Aedes aegypti from Jeddah, Saudi Arabia and the Cayman Islands, which show differences in resistance mechanisms. Females of different ages (5, 7, 10 and 14 days-old) were exposed using WHO tube assays to either a single dose of insecticide, or in a second experiment females (initially 5 days-old) were exposed daily over 10 days. Age only increased mortality in the Jeddah strain at 14 days-old and had no impact on the Cayman strain. This is consistent with greater impact linked to metabolic resistance in the Jeddah strain, though results from qPCR of four candidate genes, failed to provide evidence for a candidate underpinning an age-dependent change in resistance. With repeated exposure, mortality rates of surviving females decreased to very low levels, suggesting that surviving older cohorts of females may exhibit substantially lower susceptibility than young females in single exposure assays. Our results indicate that testing young females with a single insecticide exposure should capture minimum susceptibility for the majority of the population, but a small fraction of older females may prove particularly unresponsive to pyrethroid-based control measures.

Resistance mechanisms of epidermal growth factor receptor tyrosine kinase inhibitors in non-small cell lung cancer patients: A meta-analysis.

BackgroundDifferences in the resistance mechanisms of epidermal growth factor receptor tyrosine kinase inhibitors in patients with non‐small cell lung cancer (NSCLC) harboring epidermal growth factor receptor mutations are unknown. This meta‐analysis aimed to clarify the differences in resistance mechanisms after treatment with various epidermal growth factor receptor tyrosine kinase inhibitors.MethodsWe systematically searched PubMed, Cochrane, and Web of Science on July 29, 2020, for relevant studies on acquired resistance mechanisms against epidermal growth factor receptor tyrosine kinase inhibitors. The primary outcome measure was differences in the resistance mechanism between individual or generations of epidermal growth factor receptor tyrosine kinase inhibitors.ResultsIn total, 33 trials involving 2418 individuals were included and analyzed. T790M was significantly less frequent after afatinib treatment (40.2%, 95% confidence interval [CI]: 31.7%–48.7%) than after gefitinib and erlotinib treatments (52.5%, 95% CI: 48.7%–56.3%, p = 0.005). There were no significant differences between Asian and non‐Asian patients in the incidence of T790M after gefitinib, erlotinib, and afatinib treatments. Regarding epidermal growth factor receptor pathway‐independent resistant mechanisms, the incidences of small cell lung cancer transformation (osimertinib: 7.9%, 95% CI: 3.6%–12.2%, others: 2.3%, 95% CI: 0.8%–3.8%) and Kirsten rat sarcoma (KRAS) viral oncogene homolog mutation (osimertinib: 4.6%, 95% CI: 1.5%–7.7%, others: 0.2%, 95% CI: 0.0%–1.7%) were significantly higher following osimertinib treatment than with others.ConclusionsSignificant differences in the incidence of resistance mechanisms among epidermal growth factor receptor tyrosine kinase inhibitors exist, which should be taken into consideration when choosing the treatment strategy.

N-alkylisatin-based microtubule destabilizers bind to the colchicine site on tubulin and retain efficacy in drug resistant acute lymphoblastic leukemia cell lines with less in vitro neurotoxicity

BackgroundDrug resistance and chemotherapy-induced peripheral neuropathy continue to be significant problems in the successful treatment of acute lymphoblastic leukemia (ALL). 5,7-Dibromo-N-alkylisatins, a class of potent microtubule destabilizers, are a promising alternative to traditionally used antimitotics with previous demonstrated efficacy against solid tumours in vivo and ability to overcome P-glycoprotein (P-gp) mediated drug resistance in lymphoma and sarcoma cell lines in vitro. In this study, three di-brominated N-alkylisatins were assessed for their ability to retain potency in vincristine (VCR) and 2-methoxyestradiol (2ME2) resistant ALL cell lines. For the first time, in vitro neurotoxicity was also investigated in order to establish their suitability as candidate drugs for future use in ALL treatment.MethodsVincristine resistant (CEM-VCR R) and 2-methoxyestradiol resistant (CEM/2ME2-28.8R) ALL cell lines were used to investigate the ability of N-alkylisatins to overcome chemoresistance. Interaction of N-alkylisatins with tubulin at the the colchicine-binding site was studied by competitive assay using the fluorescent colchicine analogue MTC. Human neuroblastoma SH-SY5Y cells differentiated into a morphological and functional dopaminergic-like neurotransmitter phenotype were used for neurotoxicity and neurofunctional assays. Two-way ANOVA followed by a Tukey’s post hoc test or a two-tailed paired t test was used to determine statistical significance.ResultsCEM-VCR R and CEM/2ME2-28.8R cells displayed resistance indices of > 100 to VCR and 2-ME2, respectively. CEM-VCR R cells additionally displayed a multi-drug resistant phenotype with significant cross resistance to vinblastine, 2ME2, colchicine and paclitaxel consistent with P-gp overexpression. Despite differences in resistance mechanisms observed between the two cell lines, the N-alkylisatins displayed bioequivalent dose-dependent cytotoxicity to that of the parental control cell line. The N-alkylisatins proved to be significantly less neurotoxic towards differentiated SH-SY5Y cells than VCR and vinblastine, evidenced by increased neurite length and number of neurite branch points. Neuronal cells treated with 5,7-dibromo-N-(p-hydroxymethylbenzyl)isatin showed significantly higher voltage-gated sodium channel function than those treated with Vinca alkaloids, strongly supportive of continued action potential firing.ConclusionsThe N-alkylisatins are able to retain cytotoxicity towards ALL cell lines with functionally distinct drug resistance mechanisms and show potential for reduced neurotoxicity. As such they pose as promising candidates for future implementation into anticancer regimes for ALL. Further in vivo studies are therefore warranted.

CRISPR-Based Functional Genomics Studies Reveal Distinct and Overlapping Genes Mediating Resistance to Different Classes of Heterobifunctional Degraders of Oncoproteins: Implications for Novel Therapeutics across Diverse Neoplasias

Heterobifunctional proteolysis-targeting chimeric compounds leverage the activity of E3 ligases (e.g. CRBN and VHL) to induce neopmorphic ubiquitination and proteasomal degradation of target oncoproteins, with potent preclinical activity against diverse neoplasias. Despite intense recent efforts to develop pharmacological “degraders” against many different oncoproteins, the mechanisms regulating tumor cell sensitivity to different classes of these “degraders” remain incompletely understood. To address this question in an unbiased manner, we performed genome-scale CRISPR/Cas9-based gene editing loss-of-function (LOF) studies in MM.1S multiple myeloma (MM) cells treated with CRBN-mediated degraders of BET bromodomain proteins (dBET6) or CDK9 (Thal-SNS-032); or with VHL-mediated degraders of BET bromodomain proteins (ARV-771 or MZ-1). We observed that MM cell resistance to any of these “degraders” does not involve genes with recurrent LOF in MM patients and association with high-risk MM (e.g. for TP53, PTEN, negative regulators of cell cycle, et.c.), suggesting that these degraders may exhibit activity against tumor cells with prognostically adverse genetic features.In tumor cells resistant to the CRBN-mediated degraders dBET6 and Thal-SNS-032, we observed significant enrichment of sgRNAs targeting CRBN itself or (to a lesser extent) other components or regulators of its cullin RING ligase (CRLCUL4A) complex, including members of the COP9 signalosome (COPS7A, COPS7B, COPS2, COPS3, COPS8, GPS1, etc.), DDB1, or the E2 ubiquitin conjugating enzyme UBE2G1. In tumor cells resistant to the VHL-mediated degraders MZ-1 and ARV-771, we observed pronounced enrichment of sgRNAs for CUL2, VHL itself, other members (e.g. RBX1, elongin B/C [TCEB1, TCEB2] of the CUL2 complex with VHL), as well as COP9 signalosome genes (COPS7B, COPS8) and UBE2R2. We also validated, using individual sgRNAs for several of these candidate genes that their CRISPR knockout can decrease tumor cell response to dBET6 and Thal-SNS-032 treatment (e.g. for CRBN, COPS7B, COPS2, or COPS8) or MZ-1 and ARV-771 (e.g. for VHL, COP7B and COPS8). Notably, the sgRNAs against COP9 signalosome genes conferred less pronounced decrease in sensitivity to VHL-, than CRBN-based, degraders, suggesting that COP9 signalosome loss has differential roles in the function of CUL4ACRBN vs. CUL2VHL and potentially other CRL complexes. Tumor cells isolated from our CRISPR knockout screens with confirmed resistance to a given degrader were then treated with other degraders operating through the same or different E3 ligase; and against the same or different oncoprotein: we observed cross-resistance between degraders operating through the same E3 ligase against different oncoproteins, but not for degraders targeting the same protein via different E3 ligase/CRLs: this result is consistent with our observation for substantial gene-level differences (despite pathway-level similarities) for resistance mechanisms for CRBN- vs. VHL-based degraders. In conclusion, our study systematically defined at genome-scale the resistance mechanisms of tumor cells against degraders which leverage the same E3 ligase against different targets; or target the same oncoprotein through different E3 ligases/CRL complexes. We observed that for multiple types of degraders, tumor cell resistance is primarily mediated by prevention of, rather than adaptation to, breakdown of the target oncoprotein. The observed pathway-level similarities and major individual gene-level differences in resistance mechanisms for CRBN- and VHL-mediated degraders likely reflects the different composition and regulation of the respective CRL complexes mediating the action of these classes of degraders Our observations suggest that preventing or delaying resistance to pharmacological degradation of oncoproteins may require concurrent or sequential/alternating use of degraders operating through different E3 ligases and ideally, different CRL complexes; while synthetic lethal strategies to prevent COP9 signalosome LOF may also be contemplated to counteract a common, but quantitatively less pronounced, potential mechanism of resistance for several different classes of degraders. Collectively, our study highlights important new directions in the development of new pharmacological degraders for blood cancers and other neoplasias. DisclosuresRichardson:Karyopharm: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Research Funding; Oncopeptides: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees. Licht:Celgene: Research Funding. Boise:Abbvie: Consultancy; AstraZeneca: Honoraria. Gray:C4 Therapeutics: Consultancy. Mitsiades:TEVA: Research Funding; Janssen/ Johnson & Johnson: Research Funding; EMD Serono: Research Funding; Takeda: Other: employment of a relative; Abbvie: Research Funding.

Multiple target site resistance to glyphosate in junglerice (Echinochloa colona) lines from California orchards.

In California specialty cropping systems such as vineyards and orchards, Echinochloa colona is present as a summer annual weed. It is able to germinate throughout the growing season whenever favorable conditions are present, and management relies heavily on glyphosate applications. Glyphosate-resistant (GR) E. colona biotypes are present in the state, but the levels of resistance observed suggest that there may be differences in mechanisms of resistance among populations. Echinochloa colona lines collected from different regions of California's Central Valley presented resistance levels ranging from 1.4 to 4.3-fold compared to susceptible lines. No differences in the absorption and translocation of [14 C]-glyphosate were observed among lines. Resistant lines accumulated eight-fold less shikimic acid after treatment with 435 and 870 g a.e. ha-1 glyphosate compared to the most susceptible line. Sequencing of a region of the EPSPS gene revealed three single nucleotide changes leading to amino acid substitutions at Proline 106, including Pro106Leu, Pro106Thr and Pro106Ser. These results indicate that an altered target site in EPSPS is contributing to resistance in these lines and resistance has evolved independently, multiple times in the Central Valley of California. Additional research is needed to further understand the genomic contributions of resistance loci in this polyploid weed species. © 2018 Society of Chemical Industry.

Plot based heritability estimates and categorization of cassava genotype response to cassava brown streak disease

Cassava brown streak disease (CBSD) caused by Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV) is a threat to food security in sub-Saharan Africa, where the disease persistently reduces overall root quality and quantity resulting in up to 100% yield losses. Complexities in CBSD symptom expression and the damage caused on leaves, stems and roots throughout the 12 months of cassava growth require that appropriate ways of categorizing genotype response and optimal stages of evaluation be identified. This study aimed at: 1) determining plot based heritability of CBSD based on symptom expression and 2) categorizing genotype resistance to CBSD based on symptom expression. Herein, 41 genotypes were evaluated for two years at Namulonge with an additional evaluation conducted across three locations. Evaluations were done at three, six, nine and twelve months after planting. Genotype responses to CBSD varied significantly. High broad sense heritability estimates of up to 0.81 (incidence) and 0.71 (severity) were obtained.Average disease severity scores had higher broad sense heritability estimates (0.53 and 0.65) than maximum disease severity scores (0.33 and 0.61) for root and foliar severities respectively. These findings are important in choosing an appropriate evaluation method for CBSD. Genotypes displayed differing CBSD responses in type, locality and severity of symptoms. This suggested that genotypes had differences in mechanisms of resistance that can be exploited in CBSD resistance breeding.

Outcomes in patients with non-small-cell lung cancer and acquired Thr790Met mutation treated with osimertinib: a genomic study

Osimertinib is approved for the treatment of non-small-cell lung cancer in patients who develop the EGFR Thr790Met mutation after treatment with epidermal growth factor receptor (EGFR) tyrosine-kinase inhibitors (TKIs). We assessed outcomes in patients with non-small-cell lung cancer and the EGFR Thr790Met mutation who were treated with osimertinib, a third-generation EGFR TKI, after previous treatment failure with one or more other EGFR TKIs. Eligible patients had been enrolled at one centre in the AURA study, had shown resistance to a previous EGFR TKI, and had EGFR-activating mutations and acquired Thr790Met mutation detectable in tumour tissue or plasma. Patients took 20-240 mg osimertinib per day until disease progression or development of intolerable side-effects. Plasma samples were collected every 6 weeks and tumour tissue biopsy was done at study entry and was optional after disease progression. We tested samples for resistance mechanisms, including EGFR-activating, Thr790Met, and Cys797Ser mutations, and assessed associations with overall survival, progression-free survival, and survival after disease progression. Of 71 patients enrolled in AURA, 53 were eligible for this analysis. Median progression-free survival was 11·1 months (95% CI 8·4-13·9) and overall survival was 16·9 months (11·7-29·1). 47 patients had disease progression. Median overall survival after osimertinib progression was 5·4 months (95% CI 4·1-10·0). Plasma samples were available for 40 patients after disease progression. 12 (30%) of these had the Thr790Met mutation (four of whom also had Cys797Ser mutations). Patients without detectable EGFR-activating mutations in plasma before treatment had the best overall and post-progression survival (22·4 months, 95% CI 15·6-not reached, and 10·8 months, 7·2-not reached, respectively). Loss of the Thr790Met mutation but presence of EGFR-activating mutations in plasma were associated with the shortest progression-free survival (median 2·6 months, 95% CI 1·3-not reached). In 22 post-progression tumour samples, we found one squamous cell and two small-cell transformations. We detected Thr790Met in nine (50%) of 18 samples, Cys797Ser in two (17%) of 12, cMET amplification in five (50%) of ten, BRAF mutation in one (8%) of 13, and KRAS mutation in one (8%) of 13. Heterogeneous resistance mechanisms developed in patients receiving osimertinib. Differences in resistance mechanisms might dictate future development strategies for osimertinib in clinical trials. AstraZeneca, Taiwan Ministry of Science and Technology.

A Higher Proportion of the EGFR T790M Mutation May Contribute to the Better Survival of Patients with Exon 19 Deletions Compared with Those with L858R

IntroductionIncreasing evidence has demonstrated that exon 19 deletions (Del19) and L858R mutation in EGFR have different prognostic and predictive roles in NSCLC. We aimed to investigate whether these two mutations produced differences in mechanisms of resistance to EGFR tyrosine kinase inhibitors. MethodsConsecutive patients with advanced EGFR-mutant NSCLC who acquired resistance to EGFR tyrosine kinase inhibitors and underwent postprogression biopsy were enrolled. Mechanisms including T790M mutation, mesenchymal-epithelial transition proto-oncogene (MET) amplification, and histological transformation, as well as KRAS, phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha gene (PIK3CA) mutation, and anaplastic lymphoma receptor tyrosine kinase gene (ALK) fusion, were analyzed. ResultsThe prevalence of T790M mutation was significantly higher in the Del19 subgroup than that in L858R subgroup (50.4% versus 36.5%, p = 0.043). Apart from this, there was no difference in other mechanisms including MET amplification and histological transformation. The median overall survival (OS) of patients with T790M mutation was 36.0 months (95% confidence interval [CI]: 30.9–41.2), which was significantly longer than the 26.5 months (95% CI: 24.0–29.0) in MET-positive patients, 19.7 months (95% CI: 18.2–21.2) in patients with histological transformation, and 23.0 months (95% CI: 17.4–28.6) in the KRAS/PIK3CA/ALK-altered population (p = 0.021). The hazard ratios of the MET-amplification subgroup and subgroup with histological transformation were 1.809-fold and 2.370-fold higher than that in T790M-positive subgroup. The median OS times were months 33.3 (95% CI: 28.9–37.7) in the Del19 subgroup and 26.4 months (95% CI: 23.2–29.6) in the L858R subgroup (p = 0.028). However, in multivariable analysis adjusted for T790M genotype, the EGFR mutation subtype was no longer found to be significant. ConclusionsSignificant OS benefit was observed in patients with T790M mutation, suggesting that a larger proportion of T790M mutation might contribute to the better survival of patients with Del19.

MIC score, a new tool to compare bacterial susceptibility to antibiotics application to the comparison of susceptibility to different penems of clinical strains of Pseudomonas aeruginosa.

This study aimed to compare the susceptibility to carbapenems (imipenem, meropenem and doripenem) of clinical strains of Pseudomonas aeruginosa. It also studied whether susceptibility to imipenem or meropenem could predict, reliably, susceptibility to doripenem. Pseudomonal strains were collected from respiratory specimens, half of them from cystic fibrosis patients. MICs were determined according to European Committee on Antimicrobial Susceptibility Testing recommendations. Carbapenems were compared according to the susceptible, intermediate or resistant categories. A new approach also allowed comparing these carbapenems in a 'MIC score' taking into account the differences in breakpoints between drugs. One hundred thirty-nine strains were studied. They were found to be statistically more susceptible to meropenem than to the two other drugs. However, this difference was small: less than one dilution between the agents. This study also highlighted a significant correlation between susceptibility to penems taken in pairs. However, susceptibility to imipenem or meropenem did not reliably predict susceptibility to doripenem. Despite potential differences in resistance mechanisms, the Pseudomonas aeruginosa strains showed close susceptibility to three carbapenems. This was true for both cystic fibrosis patients and others. However, there were variations between strains. That justifies MICs to be determined for each of the three penems. This might be useful in case of elevated MICs and/or for potentially difficult-to-treat infections such as pneumonia in patients with cystic fibrosis patients.

ANGI-09COMPARISON OF PROGRESSION PATTERN (PP) AND ADVERSE EVENTS (AES) WITH USE OF BEVACIZUMAB (BEV) ACROSS MULTIPLE LINES IN THE TREATMENT IN GLIOBLASTOMA (GBM)

BACKGROUND: RTOG 0825 and AVAGlio studies showed mixed survival results when bev was added to first line GBM treatment. In North America, use of bev is only approved for recurrent disease. Others have explored the reasons for bev failure; two hypotheses included 1) bev promoting distant and diffuse PP as a resistance mechanism 2) AEs related to the use of bev. This study was undertaken to describe and compare PPs and AEs associated with bev in newly diagnosed and recurrent GBM. METHODS: Sixty-four GBM patients treated with bev at McGill University Hospitals between 2008-2014 were identified. 30/64 (46.9%) patients received bev 1st line (B1L) and 34/64 (53.1%) received bev in 2nd line and beyond (B2L+). The median length of treatment was 24.4 weeks (range 0-232.7). Retrospective data collection through chart review for PPs was categorized: local, distant, diffuse and multifocal progression (LoP, DsP, DfP, MfP) or multi-pattern progression (MpP). AEs were documented as per CTCAE version 4.0. RESULTS: B1L vs B2L+ showed following PP: LR 46.7% vs 26.5%, DsP 3.3% vs 2.9%, DfP 20% vs 47%, MfR 10% vs 8.8%, MpP 3.3% vs 11.8%. No difference between B1L vs B2L+ were observed in DsP (p = 0.3) and DfP (p = 0.4) PP. Grade 3/4 AEs in B1L vs B2L + : fatigue 33.3% vs 17.6%, hypertension 26.7% vs 5.9%, thrombocytopenia 26.7% vs 11.8%, neutropenia 26.7% vs 11.8%, anemia 23.3% vs 11.8%, leucopenia 20% vs 8.8%, deep vein thrombosis, 23.3% vs 5.9%, seizures 16.7% vs 8.8%, brain hemorrhage 6.7% vs <1% and delayed wound healing 6.7% vs 2.9%. More total grade 3/4 AEs occurred in B1L (p = 0.000519). CONCLUSION: In our cohort, no PP differences occurred between B1L vs B2L + , suggesting no difference in resistance mechanism. Moreover, similar AEs occurred in B1L and B2L with more grade 3/4 A/Es in B1L group, supporting use in B2L+.

DETERMINE THE EFFICACY OF CARBOPLATIN WITH VITAMIN C IN DALTON'S LYMPHOMA TUMOR IN MICE MODEL

Carboplatin, a plat inum - containing anticancer drug treatment cau sed a significant increase in the life span of ascites Dalton’s lymphoma tumor bearing mice. However , as compared to carboplatin alone, combination treatment with vitamin C plus carboplatin resulted in better therapeutic efficacy against Dalton's lymphoma tumor. To investigate difference in resistance mechanism to d i a m i n e ( cyclobutane - 1,1 - di carboxylato (2 - ) - O,O’) platinum (CBDCA) and changes in antioxidant levels in liver tissues in relation to different stages of Dalton’s lymphoma tumor growth and the fol lowing carboplatin treatment. The reduction in intracellular accumulation of CBDCA is thought to be an important factor indicating that CBDCA resistant in DLA cell are mutually cross - resistant. Glutathione level increased in liver of tumor bearing compared to normal mice. I ntracellular accumulation of platinum found to be higher in tissue, after carboplatin treatment the platinum accumulation were significantly decre a sed . Antioxidant defense systems such as superoxide dismutase , glutathione peroxidase gluta thione reductase activities and Glutathione – S - transferase activity w ere determined in tissues of carboplatin treatment at different stages of Dalton’s lymphoma tumor gr owth . Resistance mechanism based on reduction of intracellular drug accumulations and i ncreased activity of GST also depend the level of antioxidant enzymes , GST activities are playing significant role in development of resistance in the presences of tumor under various newly developed platinum derivative factor. Anticancer drugs kill suscep tible cells through induction of apoptosis. Alteration of apoptotic pathways in drug –

Parallel evolution and local differentiation in quinolone resistance in Pseudomonas aeruginosa

The emergence and spread of antibiotic resistance in pathogens is a major impediment to the control of microbial disease. Here, we review mechanisms of quinolone resistance in Pseudomonas aeruginosa, an important nosocomial pathogen and a major cause of morbidity in cystic fibrosis (CF) patients. In this quantitative literature review, we find that mutations in DNA gyrase A, the primary target of quinolones in Gram-negative bacteria, are the most common resistance mutations identified in clinical samples of all origins, in keeping with previous observations. However, the identities of non-gyrase resistance mutations vary systematically between samples isolated from CF patients and those isolated from acute infections. CF-derived strains tend to harbour mutations in the efflux pump regulator nfxB, while non-CF strains tend to bear mutations in the efflux regulator mexR or in parC, which encodes one of two subunits of DNA topoisomerase IV. We suggest that differences in resistance mechanisms between CF and non-CF strains result either from local adaptation to different sites of infection or from differences in mutational processes between different environments. We further discuss the therapeutic implications of local differentiation in resistance mechanisms to a common antibiotic.

Abstract A151: Differential mechanisms of acquired resistance to IGF-1R antibody therapy and a small molecule inhibitor, BMS-754807 in a human rhabdomyosarcoma Rh41 model

Abstract Background: Overexpression and/or overactivation of IGF-1R have been linked to a number of cancer types including, colon, breast, prostate, lung and sarcoma. These findings support IGF-1R as a promising therapeutic target. Agents targeting IGF-1R, including IGF-1R antibodies and small molecule inhibitors are currently in clinical development. A common phenomenon resulting in clinical failures is the development of resistance to anti-cancer therapies. In the present study, the goal is to compare the mechanisms of acquired resistance to mAb391, an IGF-1R-blocking antibody and to BMS-754807, a small molecule dual inhibitor targeting IGF-1R/IR, and to provide insights on biomarkers for early detection of resistance and differentiation of IGF-1R targeted therapies. Methods: The human Rhabdomyosarcoma cell line Rh41, which is sensitive to IGF-1R inhibition and responsive to both agents, was used to develop two acquired-resistant cell lines, Rh41-807R and Rh41-mAb391R. In addition, tumor xenograft models were developed from the resistant cell lines. Gene expression profiling was performed on the sensitive parental (Rh41), Rh41-807R, and Rh41-mAb391R cells and tumors to identify biomarkers involved in acquired resistance to BMS-754807 and/or mAb391. RNA expression profiles for selected biomarkers from both cells and tumors correlated well with protein expression patterns, as determined by Western blot analysis. Results: Differences in resistance mechanisms between Rh41-807 and Rh41-mAb391 resistant lines were observed and each utilizes different alternative growth signaling pathways as an escape mechanism. PDGFR was found to be overexpressed at the RNA level in both Rh41-807R cells (89-fold) and tumors (27-fold) and was constitutively activated; siRNA knockdown of PDGFR in Rh41-807R re-sensitized the cells to BMS-754807. Synergistic activities were observed when BMS-754807 was combined with agents that also inhibited PDGFR in the Rh41-807R models. AXL RNA expression was found to be elevated in Rh41-mAb391R (8-fold), but down-regulated in Rh41-807R cells (5-fold). In addition, BMS-754807 was active in Rh41- mAb391R cells and able to overcome resistance to the IGF-1R antibody treated cells. However, the converse was not true i.e., mAb391 did not overcome resistance in the Rh41-807R cell line. Conclusions: PDGFR overexpression and activation may drive resistance to BMS-754807, whereas differential resistance mechanisms are involved in resistance to the IGF-1R antibody. The activity of BMS-754807 in the Rh41- mAb391R acquired-resistant model suggests that treatment with BMS- 754807 may overcome resistance in patients who have failed treatment with IGF-1R antibodies. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A151.

Genome-Wide Analysis of Aromatase Inhibitor-Resistant, Tamoxifen-Resistant, and Long-Term Estrogen-Deprived Cells Reveals a Role for Estrogen Receptor

Acquired resistance to either tamoxifen or aromatase inhibitors (AI) develops after prolonged treatment in a majority of hormone-responsive breast cancers. In an attempt to further elucidate mechanisms of acquired resistance to AIs, MCF-7aro cells resistant to letrozole (T+LET R), anastrozole (T+ANA R), and exemestane (T+EXE R), as well as long-term estrogen deprived (LTEDaro) and tamoxifen-resistant (T+TAM R) lines were generated. This is the first complete panel of endocrine therapy-resistant cell lines, which were generated as multiple independent biological replicates for unbiased genome-wide analysis using affymetrix microarrays. Although similarities are apparent, microarray results clearly show gene signatures unique to AI-resistance were inherently different from LTEDaro and T+TAM R gene expression profiles. Based on hierarchical clustering, unique estrogen-responsive gene signatures vary depending on cell line, with some genes up-regulated in all lines versus other genes up-regulated only in the AI-resistant lines. Characterization of these resistant lines showed that LTEDaro, T+LET R, and T+ANA R cells contained a constitutively active estrogen receptor (ER)alpha that does not require estrogen for activation. This ligand-independent activation of ER was not observed in the parental cells, as well as T+EXE R and T+TAM R cells. Further characterization of these resistant lines was performed using cell cycle analysis, immunofluorescence experiments to visualize ER subcellular localization, as well as cross-resistance studies to determine second-line inhibitor response. Using this well-defined model system, our studies provide important information regarding differences in resistance mechanisms to AIs, TAM, and LTEDaro, which are critical in overcoming resistance when treating hormone-responsive breast cancers.

Characteristics associated with Woolly Apple Aphid Eriosoma lanigerum, resistance of three apple rootstocks

AbstractThe resistance characteristics of the apple resistance genes (Er1, Er2, and Er3) to the woolly apple aphid, Eriosoma lanigerum (Hausmann) (Homoptera: Aphididae) were studied according to the performance measured on apple cultivars containing these resistance genes. The resistance characteristics of Northern Spy (Er1), Robusta 5 (Er2), and Aotea (Er3) were compared to the susceptible cultivar Royal Gala, by measuring the aphid settlement, development, and survival rates correlated with electronically monitored probing behaviour. Er1 and Er2 had a higher level of resistance with a significantly shorter period of phloem feeding, suggesting that the resistance factors were present in the phloem tissue. Phenological measurements indicated that the aphids showed poor settlement, development, and survival on Er2. Er1 also showed low settlement and survival, although not as low as Er2. Aphid performance and feeding on Aotea (Er3) were similar to Royal Gala, suggesting that some woolly apple aphids in New Zealand may have recently overcome Er3 resistance. The differences in resistance mechanisms of Er1, Er2, and Er3 are discussed in relation to the strategy of pyramiding these genes to give a durable resistance to woolly apple aphid.

Comparison of idarubicin and daunorubicin regarding intracellular uptake, induction of apoptosis, and resistance

Anthracycline antibiotics are widely used as anticancer agents. Idarubicin (4-demethoxydaunorubicin; Ida), a semisynthetic derivative of daunorubicin (Dnr) is more potent than the parent compound in vitro and in vivo. The equitoxic dose of Ida in patients is about one-fourth of that of Dnr. We compared these drugs regarding cytotoxicity, apoptosis induction, and resistance mechanisms in human leukaemic cell lines. Cytotoxicity was studied by means of the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide assay and drug-induced apoptosis by means of the Annexin V–fluorescein isothiocyanate method at similar intracellular concentrations (extracellular concentrations of 0.35 μM for Ida and 1 μM for Dnr). Ida was at least twice as potent as Dnr in MOLT-4, HL60, CEM, and K562 cell lines. It took 8 h for Ida to induce approximately 20% apoptosis, but at least 22 h for Dnr to reach 20% apoptosis at identical intracellular concentration. Ida induces a faster and higher apoptosis rate compared with Dnr. The human chronic myelogenous leukaemia cell line (K562) was selected for resistance to Dnr and Ida with and without verapamil (Ver). Continuous incubation with Dnr, but not with Ida, led to an increased mdr1 gene expression as assessed by real-time PCR. The development of mdr1 gene expression in Dnr-resistant cells could be reversed by the presence of Ver. Ver also reversed the cytotoxicity to Dnr, but not to Ida, in K562/Dnr cells. The results show that Ida is more effective than Dnr in inducing apoptosis and that there are differences in resistance mechanisms between the drugs.