Mechanisms Of Primary Resistance Research Articles

Putative mechanisms of primary resistance to EGFR-targeted therapies: A retrospective study

BackgroundsAdvanced lung adenocarcinoma (LUAD) patient with EGFR mutations often experience resistance to first-line epidermal growth factor tyrosine kinase inhibitors (EGFR-TKIs) therapy. Nonetheless, the mechanism and biomarkers of primary resistance remain unclear. Further exploration of independent prognostic factors will help clinicians identify patients who may not respond to EGFR-TKIs and select appropriate treatment strategies. MethodsA retrospective study involving 124 patients with stage IV LUAD harboring a common sensitizing EGFR mutation (exon 19 deletion or L858R mutation) who received EGFR-TKIs as first-line therapy was performed. All participants were tested by DNA-targeted sequencing in baseline samples, and there were 19 patients with progression-free survival (PFS) ≤ 3 months (cohort 1, C1, primary resistance), 22 patients with 3 < PFS < 8 months (cohort 2, C2, poor response) without known mutations associated with resistance, and 83 patients with PFS ≥ 8 months (cohort 3, C3, normal). ResultsThe most commonly mutated genes at baseline in patients prior to treatment within the entire study population. were TP53 (65 %), MYC (19 %), CDKN2A (12 %), MUC16 (12 %) and RBM10 (12 %). The baseline characteristics, except for the proportions of patients with EGFR L858R mutation and exon 19 deletion in C1 plus C2 compared to C3 (p = 0.036), were not significantly different among the cohorts. The frequencies of PIK3C2G, STK11, EPAS1, RARA and BTG2 variation were significantly higher in C1, the primary resistance group. Multivariate Cox analysis revealed that PIK3C2G (HR 15.70 95 % CI 3.24–76.05, p < 0.001), STK11 (HR 17.04, 95 % CI 3.68–78.92, p < 0.001), EPAS1 (HR 11.99, 95 % CI 2.57–56.03, p = 0.002), and BTG2 amplification (HR 9.53, 95 % CI 1.67–54.28, p = 0.011) were significantly associated with shorter PFS. ConclusionsThe genomic landscape varies significantly among patients with LUAD, which should be considered when making personalized treatment decisions. This information could provide insights into molecular changes and their effects on clinical treatment in diverse patients with LUAD harboring sensitizing EGFR mutations.

B-255 Identification of the protein expression of Androgen Receptor variants using Targeted proteomics in clinical Castration Resistant Prostate Cancer models

Abstract Background Castration Resistant Prostate Cancer (CRPC) is a treatment resistant form of prostate cancer (PCa). Currently, there is not a way to identify which patients will develop this resistance before full blown CRPC develops. Therefore, all PCa treatment approaches are similar yet this results in tumor regression in some cases and progression in others. Targeting the androgen receptor (AR) is still the main focus of current therapies even in CRPC. Emergence of AR splice variants (AR-Vs) after initial treatment is thought to be one of the primary mechanisms of resistance. AR-Vs lack the ligand binding domain rendering Androgen Deprivation Therapy (ADT) ineffective in tumors expressing these variants. Recent work has identified the DNA and RNA species of ARVs in CRPC but investigation into whether the protein is translated are unknown. One exception is the approval of an antibody test that detects a specific AR-v, AR-v7, from the blood of PC patients and predicts ADT response. However, there are instances where a patient may not express ARv7 and are still resistant to ADT. This may suggest that other AR-Vs, not currently detected at the protein level, are important predictors for ADT response. Methods The primary goal of this work is to expand the protein identification of known and unknown AR-Vs that may predict response to ADT using targeted liquid chromatography tandem mass spectrometry (LC-MS/MS). We developed the AR-V targeted LC-MS/MS assay by first designing AR splice variants from unique splice regions via RNA-guided sequences translated to amino acid sequences. The AR-variants full amino acid sequences were then in-silico trypsin digested using Expasy software tool that generated tryptic peptides. Blast analysis of the AR-V peptides was performed where no overlap with other human proteins was observed. These AR-V peptide sequences were commercially synthesized along with heavy isotopically analogs to be used as internal standards in the LC-MS/MS method. The method examined 9 peptides and included 2 to 3 MRM transitions per peptide and calibration curves were constructed using unlabeled peptides. Specimens examined consisted of a panel of PCa cell lines (n= 6), CRPC patient derived xenografts tumors (PDX) (n=4) and a negative control cell line (no androgen receptor expression), Cos-1, which served with a dual function and was used as matrix for peptide MRM optimization experiments. Linearity and sensitivity for each peptide included in the method was examined using calibration curves. Results The targeted MRM approached was successfully achieved for the quantification of 9 peptides from 8 proteins in 6 PCa cell lines and 4 PDX tumor specimens. Out of the 8 proteins evaluated, we observed 3 known targets (AR-exon1, AR-V7 and AR-V12) and 4 novel AR-Vs in 22Rv1 PCa cell lines. Additionally, CRPC PDX were assessed where 3 novel AR-Vs were detected. Conclusions These preliminary results using a LC-MS/MS platform show promising identification and quantification of novel AR-Vs that have not been measured before at the protein level. Subsequent analysis of larger cohorts will further elucidate AR-Vs role in PCa treatment resistance mechanisms and possibly assist in future treatment approaches.

Mechanisms of primary resistance to immune checkpoint inhibitors in NSCLC

Mechanisms of primary resistance to immune checkpoint inhibitors in NSCLC

Abstract IA-05: Targeting KRAS for pancreatic cancer treatment

Abstract The approval of clinically effective inhibitors for KRAS G12C mutant non-small cell lung cancers in 2021 and 2022 marks a significant milestone in anti-KRAS oncogene drug discovery. This success has fueled intense efforts to develop inhibitors targeting additional KRAS mutations. However, primary and treatment-associated acquired resistance limit the depth and duration of efficacy of KRAS G12C selective and likely other KRAS inhibitors. Thus, a major challenge moving forward will be the elucidation of mechanisms of resistance to then guide development of effective combination anti-KRAS therapies. Recent studies have begun to identify mechanisms of acquired resistance, where most involve alterations in components in the RAS signaling network that then cause reactivation of KRAS effector signaling that bypass drug activity. Our studies have established the critical role of reactivation of the RAF-MEK-ERK mitogen activated protein kinase cascade, and the ERK target MYC, in driving resistance in KRAS-mutant pancreatic cancer. To fully understand how aberrant ERK and MYC activation overcomes KRAS inhibitor response, we have established a comprehensive molecular portrait of ERK- and MYC-dependent activities. We have also identified YAP/TAZ-TEAD activation as a mechanism of acquired resistance and have characterized KEAP1 loss as a mechanism of primary resistance. Together, these studies have identified drug combination strategies that enhance KRAS inhibitor activity. Progress in these and other studies will be presented. Citation Format: Channing J. Der. Targeting KRAS for pancreatic cancer treatment [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr IA-05.

Substrate dependence of transport coupling and phenotype of a small multidrug resistance transporter in Pseudomonas aeruginosa.

Small multidrug resistance (SMR) transporters are a class of efflux pumps found in many pathogens, although their contributions to antibiotic resistance are not fully understood. We hypothesize that these transporters may confer not only resistance but also susceptibility, by dissipating the proton-motive force. This means to use an SMR transporter as a target; it merely needs to be present (as opposed to being the primary resistance mechanism). Here, we test this hypothesis with an SMR transporter found in Pseudomonas aeruginosa and find that it can perform both antiport (conferring resistance) and substrate-gated proton leak. Proton leak is detrimental to growth in Escherichia coli but not P. aeruginosa, suggesting that P. aeruginosa responds differently to or can altogether prevent ∆pH dissipation.

Molecular Epidemiology and Horizontal Transfer Mechanism of optrA-Carrying Linezolid-Resistant Enterococcus faecalis.

The aim of this work was to provide a theoretical and scientific basis for the treatment, prevention, and control of clinical drug-resistant bacterial infections by studying the molecular epidemiology and horizontal transfer mechanism of optrA-carrying linezolid-resistant Enterococcus faecalis strains (LREfs) that were clinically isolated in a tertiary hospital in Kunming, China. Non-repetitive LREfs retained in a tertiary A hospital in Kunming, China. The strains were identified by Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The transferability and horizontal transfer mechanism of optrA gene were analyzed using polymerase chain reaction (PCR), whole-genome sequencing (WGS), and conjugation experiments. A total of 39 LREfs strains were collected, and all of them were multi-drug resistant. There were 30 LREfs strains (76.9%) carrying the optrA gene, The cfr, poxtA genes and mutations in the 23S rRNA gene were not detected. The conjugation experiments showed that only three of 10 randomly selected optrA-carrying LREfs were successfully conjugated with JH2-2. Further analysis of one successfully conjugated strain revealed that the optrA gene, located in the donor bacterium, formed the IS1216E-erm(A)-optrA-fexA-IS1216E transferable fragment under the mediation of the mobile genetic element (MGE) IS1216E, which was then transferred to the recipient bacterium via horizontal plasmid transfer. Carrying the optrA gene is the primary resistance mechanism of LREfs strains. The optrA gene could carry the erm(A) and fexA genes to co-transfer among E. faecalis. MGEs such as insertion sequence IS1216E play an important role in the horizontal transfer of the optrA gene.

First detection of kdr L1014F allele in Anopheles ziemanni and Anopheles pharoensis in Cameroon and distribution of the allele in members of the Anopheles gambiae complex

BackgroundKnockdown resistance (kdr) is one of the primary resistance mechanisms present in anopheline species. Although this mutation is largely spread across the Anopheles gambiae s.l. members, its prevalence in other species is still not well documented.MethodsThe present study investigated the distribution and allelic frequencies of kdr in An. gambiae s.l., An. pharoensis, and An. ziemanni samples collected in 2022 and 2023 in nine sites spread across five ecogeographical settings in Cameroon. Members of the An. gambiae complex were identified molecularly by polymerase chain reaction (PCR). kdr L1014F and L1014S alleles were screened by PCR and confirmed by sequencing.ResultsAn. gambiae (49.9%), An. coluzzii (36.5%), and An. arabiensis (13%) were identified, and the frequency of the kdr L1014F was high in both An. gambiae and An. coluzzii in all sites. The kdr L1014F allele was detected for the first time in 8 out of 14 An. ziemanni samples examined and in 5 out of 22 An. pharoensis samples examined. The kdr L1014S allele was scarce and found only in the heterozygote “RS” state in An. arabiensis and An. gambiae in Yangah and Santchou.ConclusionsThe present study sheds light on the rapid expansion of the kdr L1014F allele in malaria vectors in Cameroon and stresses the need for surveillance activities also targeting secondary malaria vectors to improve the control of malaria transmission.Graphical

PD-L1 induces autophagy and primary resistance to EGFR–TKIs in EGFR-mutant lung adenocarcinoma via the MAPK signaling pathway

Resistance to epidermal growth factor receptor (EGFR)–tyrosine kinase inhibitors (TKIs) is a significant cause of treatment failure and cancer recurrence in non-small cell lung cancer (NSCLC). Approximately 30% of patients with EGFR-activating mutations exhibit primary resistance to EGFR–TKIs. However, the potential mechanisms of primary resistance to EGFR–TKIs remain poorly understood. Recent studies have shown that increased expression of programmed death ligand-1 (PD-L1) is associated with EGFR–TKIs resistance. Therefore, the present study aimed to investigate the mechanism of PD-L1 in primary resistance to EGFR–TKIs in EGFR-mutant lung adenocarcinoma (LUAD) cells. We found that PD-L1 was associated with poor prognosis in patients with EGFR-mutant LUAD, while the combination of EGFR–TKIs with chemotherapy could improve its therapeutic efficacy. In vitro and in vivo experiments revealed that PD-L1 promoted the proliferation and autophagy and inhibited the apoptosis of LUAD cells. Mechanistic studies demonstrated that upregulation of PD-L1 was critical in inducing autophagy through the mitogen-activated protein kinase (MAPK) signaling pathway, which was beneficial for tumor progression and the development of gefitinib resistance. Furthermore, we found that gefitinib combined with pemetrexed could synergistically enhance antitumor efficacy in PD-L1-overexpression LUAD cells. Overall, our study demonstrated that PD-L1 contributed to primary resistance to EGFR–TKIs in EGFR-mutant LUAD cells, which may be mediated by inducing autophagy via the MAPK signaling pathway. These findings not only help improve the prognosis of patients with EGFR-mutant LUAD but also provide a reference for the research of other cancer types.

Phenotypic and Genotypic Characterization of AmpC Beta-Lactamase in Clinical Isolates of Pseudomonas aeruginosa Findings From a Tertiary Care Hospital.

Background and aim Pseudomonas aeruginosa is an opportunistic pathogen responsible for various healthcare-related infections, which are difficult to treat due to intrinsic and acquired resistance. This study aimed to investigate AmpC β-lactamase production using phenotypic and genotypic methods in Pseudomonas aeruginosa strains isolated from a tertiary care hospital in Karad, Maharashtra, India. Material and methods Over one year, a descriptive cross-sectional study was conducted at the Department of Microbiology, Krishna Institute Medical Sciences, Krishna Vishwa Vidyapeeth, Karad. Phenotypic detection of AmpC beta-lactamase was performed using the Cefoxitin-Cloxacillin Double-Disc Synergy Test method, and genotypic detection was conducted using conventional polymerase chain reaction (PCR) targeting the bla Pseudomonas-derived cephalosporinases (PDC) and bla cephamycinase (CMY) genes. Results Out of 205 clinical isolates of Pseudomonas aeruginosa, 110 (53.66%) showed AmpC production phenotypically, while 86 (41.95%) were positive genotypically. The blaPDC gene was detected in 36.10% of isolates, and the blaCMY gene in 10.73% ofisolates. Conclusions The study findings indicate that AmpC-β-lactamase stands out as the primary resistance mechanism in strains of Pseudomonas aeruginosaisolated from the hospital. PCR study concluded that blaPDC (36.10 %) was the leading gene responsible for AmpC synthesis among study isolates. Early detection of AmpC beta-lactamase production by employing phenotypic and genotypic methods is crucial for detecting antibiotic resistance. This dual approach enables healthcare professionals to decide on the most effective antibiotics and mitigate the development of resistance.

Chlorantraniliprole Resistance in Spodoptera frugiperda: Resistance Monitoring, Resistance Risk, and Resistance Mechanisms.

Spodoptera frugiperda is a significant global pest, and chlorantraniliprole (CAP) is extensively used in China for its control. Understanding CAP resistance in S. frugiperda is crucial for effective management of this pest. Field populations exhibited varying degrees of resistance to CAP (RR = 1.74-5.60-fold). After 10 generations of selection, the CAP-resistant strain developed over 10-fold resistance, with a realized heritability (h2) of 0.10. Genetic analysis reveals inheritance patterns as autosomal, incomplete recessive, and monofactorial. The CAP-resistant strain showed limited cross-resistance to lufenuron and tetrachlorantraniliprole, negative cross-resistance to spinetoram, and no observed cross-resistance to other insecticides. Biochemical analysis suggested that P450-mediated detoxification is the primary resistance mechanism, with 26 genes overexpressed in the CAP-resistant strain. Additionally, the knockdown of CYP4L13, CYP6B39, CYP6B40, and CYP4G74 significantly increased the sensitivity of the resistant larvae to CAP. These findings highlight the resistance risk of CAP in S. frugiperda and emphasize the crucial role of P450 enzymes in resistance.

The Role of the PI3K/Akt/mTOR Axis in Head and Neck Squamous Cell Carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is one of the most common malignancies globally, representing a significant public health problem with a poor prognosis. The development of efficient therapeutic strategies for HNSCC prevention and treatment is urgently needed. The PI3K/AKT/mTOR (PAM) signaling pathway is a highly conserved transduction network in eukaryotic cells that promotes cell survival, growth, and cycle progression. Dysfunction in components of this pathway, such as hyperactivity of PI3K, loss of PTEN function, and gain-of-function mutations in AKT, are well-known drivers of treatment resistance and disease progression in cancer. In this review, we discuss the major mutations and dysregulations in the PAM signaling pathway in HNSCC. We highlight the results of clinical trials involving inhibitors targeting the PAM signaling pathway as a strategy for treating HNSCC. Additionally, we examine the primary mechanisms of resistance to drugs targeting the PAM pathway and potential therapeutic strategies.

Liposomal delivery of organoselenium-cisplatin complex as a novel therapeutic approach for colon cancer therapy

Cisplatin is a widely-used chemotherapeutic agent for the treatment of various solid neoplasms including colon cancer. Cisplatin-induced DNA damage is restricted due to dose-related adverse reactions as well as primary resistance mechanisms. Therefore, it is imperative to utilize novel therapeutic approaches to circumvent cisplatin limitations and attenuate its normal tissues toxicity. In this study, we exploited a novel PEGylated liposomes with greater efficiency to treat colon cancer. For this, an organoselenium compound (diselanediylbis decanoic acid (DDA)) was synthesized, and liposomes composed of Egg PC or HSPC, as well as DOPE, mPEG2000-DSPE, cholesterol and DDA at varying molar ratios were prepared by using thin-film method. Cisplatin loading was performed through incubation with liposomes. Characterization of nanoliposomes indicated a favarable size range of 91–122 nm and negative zeta potential of −9 to −22 mv. The organoselenium compound significantly improved cisplatin loading efficiency within the liposomes (83.4 %). Results also revealed an efficient bioactivity of cisplatin liposome on C26 cells compared to the normal cells. Further, DDA bearing liposomes significantly improved drug residence time in circulation, reduced toxicity associated with the normal tissues, and enhanced drug accumulation within the oxidative tumor microenvironment. Collectively, results indicated that cisplatin encasement within liposomes by using this method could significantly improve the therapeutic efficacy in vivo, and merits further investigations.

Antibody-Drug Conjugates in Urothelial Cancer: From Scientific Rationale to Clinical Development.

Antibody-drug conjugates (ADCs) have been a significant advancement in cancer therapy, particularly for urothelial cancer (UC). These innovative treatments, originally developed for hematological malignancies, use target-specific monoclonal antibodies linked to potent cytotoxic agents. This rational drug design efficiently delivers cancer cell-killing agents to cells expressing specific surface proteins, which are abundant in UC owing to their high antigen expression. UC is an ideal candidate for ADC therapy, as it enhances on-target efficacy while mitigating systemic toxicity. In recent years, considerable progress has been made in understanding the biology and mechanisms of tumor progression in UC. However, despite the introduction of immune checkpoint inhibitors, advanced UC is characterized by rapid progression and poor survival rates. Targeted therapies that have been developed include the anti-nectin 4 ADC enfortumab vedotin and the fibroblast growth factor receptor inhibitor erdafitinib. Enfortumab vedotin has shown efficacy in prospective studies in patients with advanced UC, alone and in combination with pembrolizumab. The anti-Trop-2 ADC sacituzumab govitecan has also demonstrated effectiveness in single-armed studies. This review highlights the mechanism of action of ADCs, their application in mono- and combination therapies, primary mechanisms of resistance, and future perspectives for their clinical use in UC treatment. ADCs have proven to be an increasingly vital component of the therapeutic landscape for urothelial carcinoma, filling a gap in the treatment of this progressive disease.

Discovery of 2-phenylquinazolines as potent Mycobacterium avium efflux pump inhibitors able to synergize with clarithromycin against clinical isolate.

Nontuberculous mycobacteria (NTM), which include the Mycobacterium avium complex, are classified as difficult-to-treat pathogens due to their ability to quickly develop drug resistance against the most common antibiotics used to treat NTM infections. The overexpression of efflux pumps (EPs) was demonstrated to be a key mechanism of clarithromycin (CLA) resistance in NTM. Therefore, in this work, 24 compounds from an in-house library, characterized by chemical diversity, were tested as potential NTM EP inhibitors (EPIs) against Mycobacterium smegmatis mc2155 and M. avium clinical isolates. Based on the acquired results, 12 novel analogs of the best derivatives 1b and 7b were designed and synthesized to improve the NTM EP inhibition activity. Among the second set of compounds, 13b emerged as the most potent NTM EPI. At a concentration of 4 µg/mL, it reduced the CLA minimum inhibitory concentration by 16-fold against the clinical isolate M. avium 2373 overexpressing EPs as primary mechanism of CLA resistance.

Inhibition of non-small cell lung cancer metastasis by knocking down APE1 through regulating myeloid-derived suppressor cells-induced immune disorders.

Non-small cell lung cancer (NSCLC) represents a highly immunogenic malignancy. Immunologic tolerance facilitated by myeloid-derived suppressor cells (MDSCs) is implicated in primary or secondary resistance mechanisms in NSCLC. The potential role of APE1 in regulating NSCLC metastasis by targeting MDSCs remains uncertain. This study utilized a plasmid, Plxpsp-mGM-CSF, to induce elevated granulocyte-macrophage colony-stimulating factor (GM-CSF) expression in A549 cells. Tumor transplantation experiments involved A549, A549+GM-CSF, and A549+GM-CSF-siAPE1 cell lines. Evaluation encompassed MDSCs, Treg cells, IgG, CD3, and CD8 levels. Notably, lung cancer tissues and cells displayed markedly reduced APE1 expression. siAPE1 transfection significantly curtailed tumor growth compared to the A549+GM-CSF group. APE1 knockdown orchestrated immune system modulation in lung tumor mice, characterized by diminished MDSCs but augmented Treg cells, IgG, CD3, and CD8. Additionally, APE1 knockdown led to reduced levels of pro-MDSC cytokines (HGF, CCL5, IL-6, CCL12) and a concurrent upregulation of the anti-MDSC cytokine IL-1ra. Furthermore, APE1 knockdown impeded cell viability in both A549 and H1650 cells. Transplantation of A549-GM-CSF amplified MDSC levels, fostering accelerated tumor growth, while mitigating MDSC levels through APE1 knockdown hindered tumor progression and alleviated inflammatory infiltration in lung cancer tissues. Strategies targeting the APE1/MDSC axis offer a promising approach for lung cancer prevention and treatment, presenting novel insights for NSCLC management.

Modified Carbapenem Inactivation Method and Ethylenediaminetetraacetic Acid (EDTA)-Carbapenem Inactivation Method for Detection of Carbapenemase-Producing Enterobacterales and Pseudomonas aeruginosa.

The rising incidence of carbapenem resistance in Enterobacterales and Pseudomonas aeruginosa is a concern. Since carbapenemase production is the primary resistance mechanism, detecting and identifying the genes responsible for it is crucial to effectively monitor its spread. This study aims to detect positivity for the modified carbapenem inactivation method (mCIM) and ethylenediaminetetraacetic acid (EDTA)-carbapenem inactivation method (eCIM) for the detection of carbapenemase-producing Enterobacterales and Pseudomonas aeruginosa. Methods:A cross-sectional study was carried out at a tertiary care hospital, including 250 clinical isolates of Enterobacterales and Pseudomonas aeruginosa. These isolates exhibited resistance to at least one of the carbapenems as determined by the VITEK AST 2 System (bioMérieux, USA). The isolates were subjected to mCIM testing, and those that tested positive were further tested using eCIM. The results were interpreted in accordance with the guidelines provided by the Clinical and Laboratory Standards Institute (CLSI) 2023. Out of the total 250 carbapenem-resistant Enterobacterales and Pseudomonas aeruginosa isolates, 151 (60.4%) were Klebsiella pneumonia, 44 (17.6%) were Escherichia coli, 10 (4.0%) were Enterobacter cloacae, 6 (2.4%) were Providencia spp., 4 (1.6%) were Serratia marcescens, 4 (1.6%) were Proteus mirabilis and 31 (12.4%) were Pseudomonas aeruginosa. Positivity for the mCIM was observed in 96% (240 out of 250) of the isolates. Of the mCIM-positive isolates, 234 (97.5%) also tested positive for eCIM, indicating metallo-β-Lactamase (MLB) production. A statistically significant association was found between both mCIM and eCIM positivity and the degree of resistance to carbapenem (p<0.05). Conclusion: This study shows that the inexpensive method, a combination of mCIM and eCIM assists in differentiating between serine carbapenemase producers and MLB producers, thereby guiding the selection of appropriate therapy and useful in infection control in resource-limited settings.

Efficacy and safety of anthracycline combined with alkylating agent for treatment of advanced leiomyosarcoma.

e23558 Background: Leiomyosarcoma(LMS) accounts for 12-14% of all soft tissue sarcomas(STSs) and this STS subtype has high metastatic potential. We evaluated the efficacy and safety of epirubicin combined with temozolomide(EPI-TMZ) for treatment of advanced LMS. Methods: Oral TMZ(200 or 300 mg) was administered each day on days 1-5, and intravenous EPI(60 mg/m2) was administered on days 1-4, with even distribution over these 96h within a 21‐day cycle. After 8 cycles of intensive chemotherapy, monodrug maintenance therapy consisted of TMZ alone(dosing as in the primary therapy, most choices, stop taking medication after 1 year control), and two-drug maintenance therapy consisted of TMZ with thalidomide(only 3 patients). All patients-initiated EPI-TMZ treatment between January 2018 and January 2024. Results: We examined 31 patients who received EPI-TMZ. There were 25 females and 6 males, with average age 50.5 years old. All were unresectable patients derived from abdominal retroperitoneum, uterus, inferior vena cava, mesentery, small intestine, stomach, thighs, elbows, wrists, pancreas, nasal cavity, meantime with distant metastasis. Average Ki67 is above 40%. This was a first-line treatment in 16 patients, a second- or third-line in 13, and a fourth-line in 2. At the time of data cutoff (January 31, 2024), the median PFS was 12.6 months, 1 patient had cCR, 9 PR and 20 SD. The ORR was 32.3%(10/31) and DCR was 96.8%(30/31). The only patient who progressed after two cycles of medication had PMS2 deficiency, and studies have reported that it may have a primary resistance mechanism. Even the shortest disease control period is also 5 months, indicating this regime has significant clinical benefit. Some patients have undergone tumor reduction surgery to improve quality of life after achieving significant tumor relief. The current longest PFS is 42 months up to now and under continuous tumor control. Previous retrospective studies indicated the best mPFS was 9.2 months and the best ORR was 30.9%. 2023 ESMO declared prospective doxorubicin with trabectedin obtained mPFS of 12.19 months. The most common treatment-related adverse effects were leukopenia, neutropenia, thrombocytopenia, anemia, nausea, vomiting, fatigue, and oral mucositis. Two patients had severe febrile neutropenia, but there were no treatment-related deaths, survival data follow-up. Conclusions: EPI-TMZ is potentially effective for treatment of advanced LMS, and the adverse effects appear tolerable. EPI-TMZ may provide better outcomes than reported in previous studies of other treatments for advanced LMS, worth further prospective research.

(CDK) 4/6 inhibitors: An effective treatment for homologous recombination deficient (HRD) luminal breast cancers (BC)?

e13100 Background: Cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) with endocrine therapy (ET) still represents the best 1st-line choice for hormone receptor (HR) positive(+)/HER2 negative(-) metastatic breast cancer (MBC) patients (pts). Approximately 8% of pts with HR+ BC harbors germline BRCA mutation (g BRCAmt). The interaction between Cyclin/CDK pathway, endocrine receptor (ER) and Breast Cancer gene 1/2 (BRCA1/2) is complex and evidence reported so far, await confirmation in the context of future randomized clinical trials. So far, limited data exist about treatment effectiveness of CDK4/6i in homologous recombination deficiency (HRD) Luminal BC. A current frequent question in clinical practice is which additional adjuvant therapy to prioritize for luminal BRCA tumors (tu), considering the approval of both CDK4/6i andpoly ADP ribose polymerase (PARP) inhibitors in this scenario. Considering potential impacts of genetic instability in tu biology, such as potential loss of Rb protein related to Loss of Heterozygosity (LOH) of BRCA2 as a primary resistance mechanism in CDK4/6i, we sought to analyze the clinical results of that drug in tu with genetic instability. Methods: We retrospectively analyzed real-world outcomes of CDK4/6i performance in HR+/HER2- MBC treated in a single institution, with focus in clinical outcomes of HRD BC. germline HR genes status and somatic phosphatidylinositol 3-kinase (PIK3CA) status were also analyzed. Results: A total of 384 consecutive MBC pts treated with CDK4/6i were included in this study. The most used CDK4/6i was palbociclib (40,6%), followed by ribociclib (29,8%) and abemaciclib (29,2%). Considering the whole cohort PFSm 30months (95%IC 26-34) and OSm were 56 months (95%IC 52-60) No differences in PFS or OS was observed among the 3 CDK4/6i. Median age of the cohort was 60 years old. 38.5% were premenopause Pts and 59.9% were postmenopause. A total of 149 (39%) pts performed germline screening. Between 19 pts with gHR mt, 12 were BRCA1/2 mt (just one BRCA1). The other were: 2 PALB2, 2 CHEK2, 1 ATM, 1 RAD51D, 1 RAD51C mt. So far, no differences in PFS ou OS was observed in the presence of BRCA 1/2 or other gHR mt. Nevertheless, PIK3CA mt was tested in 142 pts, 51 were positives. That mt was a predictor of worse OS (55m; 95%IC 45-65) x 63m (95%IC 56-69), p 0,019. Conclusions: The results of this real-world study suggest that treatment outcomes with CDK4/6i may be worse in pts with HRD tu due to potential differences in tu biology. Future exploration in larger samples of patients who have had biomarker testing is ongoing. Patients with Pik3CA mt treated with CDK4/6i presented worse clinical outcomes.

Osimertinib and palbociclib in an EGFR-mutated NSCLC with primary CDK4 amplification after progression under osimertinib

Precision cancer medicine has changed the treatment paradigm of patients with non-small cell lung cancer (NSCLC) with specific molecular aberrations. A major challenge is management of the resistance that tumor cells eventually develop against targeted therapies, either through primary or acquired resistance mechanisms. We report a 61 year-old male patient with metastatic NSCLC harboring an EGFR exon 19 deletion, a PIK3CA mutation, and CDK4 amplification. After an initial partial response to osimertinib as mono-therapy (third-generation EGFR tyrosine kinase inhibitor), the patient had progression of disease after 4 months of treatment and was referred for combined osimertinib and palbociclib (CDK4/6 inhibitor) treatment. Though complicated by transient pneumonitis, the patient has an ongoing partial response for > 10 months and has experienced clinical improvement on this treatment regimen. As amplification of CDK4 occurs in ~ 10% of treatment-naïve patients with EGFR-mutated NSCLC, the successful treatment of our patient with osimertinib and palbociclib may be highly relevant for future patients with NSCLC.

Potential involvement of beta-lactamase homologous proteins in resistance to beta-lactam antibiotics in gram-negative bacteria of the ESKAPEE group

BackgroundEnzymatic degradation mediated by beta-lactamases constitutes one of the primary mechanisms of resistance to beta-lactam antibiotics in gram-negative bacteria. This enzyme family comprises four molecular classes, categorized into serine beta-lactamases (Classes A, C, and D) and zinc-dependent metallo-beta-lactamases (Class B). Gram-negative bacteria producing beta-lactamase are of significant concern, particularly due to their prevalence in nosocomial infections. A comprehensive understanding of the evolution and dissemination of this enzyme family is essential for effective control of these pathogens.In this study, we conducted the prospecting, phylogenetic analysis, and in silico analysis of beta-lactamases and homologous proteins identified in 1827 bacterial genomes with phenotypic data on beta-lactam resistance. These genomes were distributed among Klebsiella pneumoniae (45%), Acinetobacter baumannii (31%), Pseudomonas aeruginosa (14%), Escherichia coli (6%), and Enterobacter spp. (4%). Using an HMM profile and searching for conserved domains, we mined 2514, 8733, 5424, and 2957 proteins for molecular classes A, B, C, and D, respectively. This set of proteins encompasses canonical subfamilies of beta-lactamases as well as hypothetical proteins and other functional groups. Canonical beta-lactamases were found to be phylogenetically distant from hypothetical proteins, which, in turn, are closer to other representatives of the penicillin-binding-protein (PBP-like) and metallo-beta-lactamase (MBL) families. The catalytic amino acid residues characteristic of beta-lactamases were identified from the sequence alignment and revealed that motifs are less conserved in homologous groups than in beta-lactamases. After comparing the frequency of protein groups in genomes of resistant strains with those of sensitive ones applying Fisher’s exact test and relative risk, it was observed that some groups of homologous proteins to classes B and C are more common in the genomes of resistant strains, particularly to carbapenems.We identified the beta-lactamase-like domain widely distributed in gram-negative species of the ESKAPEE group, which highlights its importance in the context of beta-lactam resistance. Some hypothetical homologous proteins have been shown to potentially possess promiscuous activity against beta-lactam antibiotics, however, they do not appear to expressly determine the resistance phenotype. The selective pressure due to the widespread use of antibiotics may favor the optimization of these functions for specialized resistance enzymes.