Mechanisms Of Treatment Resistance Research Articles

Impact of Antimicrobial Resistance in Health and Economic Outcomes: A Review

Background: Antimicrobial resistance (AMR) observed when microorganism adapt and multiply in the presence of drugs to which they are exposed. Antibiotic resistance is a global health problem, with the current trend of inappropriate and overuse of antibiotics predicted to cause 10 million deaths by 2050. However, resistant bacteria are multiplying worldwide, infecting healthy people, animals, and human environment, affecting countries with low- and middle-income countries. Although the discovery of antibiotics has saved many lives in recent years, increasing bacterial resistance to antibiotics threatens clinical and agricultural progress, and even treatable bacterial infections can be life-threatening. Strategies: In the present review we discuss the global burden of social and economic impact of antibiotic resistance, common mechanisms of antibiotic resistance, microbiological health and treatment resistance, patient attitudes, hospital and cancer outcomes. Summarize the impact of AMR. Potential strategies to overcome AMR such as Targeting antimicrobial-resistant enzymes, Targeting antimicrobial-resistant bacteria, Drug delivery systems, Physiochemical methods, Unconventional strategies. Conclusion: The devastating effect of antibiotic resistance is clear by way of the sudden boom inside the quantity of cases in which microorganism are immune to typically prescribed antibiotics in clinical practice. Although the superiority of bacterial resistance is growing, there is limited statistics at the impact of this hassle on the fitness and economic consequences of the clinical network.

Abstract PS12-02: Exploration of ctDNA Dynamics in the PACE Trial: A Randomized Phase II Study of Fulvestrant, Palbociclib, and Avelumab for HR+/HER2- Metastatic Breast Cancer

Abstract Background The PACE trial for HR+/HER2- metastatic breast cancer (MBC) prospectively evaluated whether continuation of the CKD4/6 inhibitor (CDK4/6i) palbociclib (P) beyond progression on prior CDK4/6i and aromatase inhibitor (AI), with a change in endocrine therapy (ET) to fulvestrant (F), improved outcomes beyond change to F alone, as well as explored the activity of a palbociclib, fulvestrant, and avelumab (P+F+A) triplet. The primary analysis did not show improvement in progression-free survival (PFS) with F+P versus F alone. In this analysis, an evaluation of serial translational PACE samples was performed to gain insights into the mutational landscape of HR+/HER2- MBC after prolonged exposure to CDK4/6i, evaluate early mutational dynamics with ongoing therapy, and identify potential genetic markers predictive of treatment response. Additionally, the identification of acquired mutations at the time of tumor progression could provide valuable information about mechanisms of treatment resistance. Methods PACE is a multicenter randomized open-label investigator-initiated phase II trial. Eligible patients (pts) had HR+/HER2- evaluable MBC with prior progression on AI and any CDK4/6i after > 6 months (mo) of therapy in the MBC setting, or during/within 12 mo in the adjuvant setting, with no more than 1 prior line of chemotherapy for MBC. Pts were randomized 1:2:1 to F alone, F+P, or F+P+A, with tumor assessments every 8 weeks. Correlative samples including circulating tumor DNA (ctDNA) were collected at baseline (BL), times of tumor assessments (C3D1), and end of study/progression (EOT). Targeted NGS (Guardant360) to evaluate mutations (somatic single nucleotide variants and indels) was performed. Kaplan-Meier survival curves were used for assessing PFS and the log-rank test for comparisons. McNemar’s test with continuity correction was used to test the changes in the frequency of mutations. Results Of the 220 randomized pts, 211 contributed ctDNA samples (200 at BL, 129 at C3D1, 146 at EOT). Characteristics of the correlative science population were similar to the intention to treat population. At BL, the most common mutations observed were ESR1 (57%), TP53 (38%), PIK3CA (36%), GATA3 (20%) ATM (12%) and Rb1 (12%). The presence of a PIK3CA mutation vs WT-PIK3CA at BL was associated with shorter PFS in the F alone arm (p=0.001) but not in the F+P or F+P+A arms. ESR1 mutations present at baseline were D538G (34%), Y537S (21%), Y537N (15%), E380Q (10%) and L536H (4%). At C3D1, the most common mutations observed were ESR1 (47%), TP53 (33%), PIK3CA (26.1), GATA3 (18.1), ATM (13%) and PTEN (8.6%). Including only BL and C3D1 matched samples (N=124), the prevalence of ESR1 mutations decreased after 2 cycles of treatment (57% at BL vs 44% at C3D1), predominantly observed in the F+P+A arm. For those in any arm with at least 6 months of disease stability on study (N=53), the presence of an ESR1 D538G mutation decreased during therapy, with 38% having a mutation at BL, 24% at C3D1, and no pts gaining ESR1 D538G at C3D1. The prevalence of PIK3CA mutations also decreased on treatment (32% at BL to 24% at C3D1), with the greatest decrease in the F+P arm. A predictive model correlating early genomic dynamics and associations with outcome will be presented. Conclusions The PACE trial offers one of the largest clinical trial cohorts describing the genomic landscape of HR+/HER2- MBC post-CDK4/6 exposure. Results from this correlative study provide a comprehensive mutational landscape post-CDK4/6i, demonstrate associations between mutations and clinical outcomes, and suggest the potential value of early ctDNA dynamics in this setting. Citation Format: Rinath Jeselsohn, Jingxin Fu, Yue Ren, Reshma Mahtani, Cynthia Ma, Angela DeMichele, Massimo Cristofanilli, Jane Meisel, Kathy Miller, Yara Abdou, Elizabeth Riley, Rubina Qamar, Priyanka Sharma, Sonya Reid, Naomi Ko, Yuan Liu, Harold Burstein, Michelle DeMeo, Sara Tolaney, Erica Mayer. Exploration of ctDNA Dynamics in the PACE Trial: A Randomized Phase II Study of Fulvestrant, Palbociclib, and Avelumab for HR+/HER2- Metastatic Breast Cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS12-02.

Abstract PO5-01-05: An Open-label, Single-center Study Assessing the Efficacy of PH-based Regimen as Neoadjuvant Therapy for HER2-positive Early-stage Breast Cancer: Integrating Genomic Features and Functional Imaging

Abstract Background: Pathological complete response (pCR) has been demonstrated as a surrogate endpoint for disease-free and overall survival in HER2-positive early-stage breast cancer (EBC). Trastuzumab (H) plus pertuzumab (P) improved pCR rates in patients with HER2-positive early-stage breast cancer. Nevertheless, biomarkers to assess the effectiveness of this treatment in the neoadjuvant setting are still unclear. This study aims to evaluate the effectiveness of fluorodeoxyglucose positron emission tomography (18F-FDG-PET) and 68 Ga-Affibody HER2 Imaging PET (HER2-PET) in HER2 EBC neoadjuvant therapy, and to identify novel biomarkers for this regimen. Methods: HER2-positive EBC patients were prospectively recruited at Fudan University Shanghai Cancer Center between April 2020 and March 2022. All patients received the 1 dose of HP, followed by 6 cycles of docetaxel (T), carboplatin (Cb), and HP. We use FoundationOne to evaluate the genomic alterations before treatment. In addition, 18F-FDG-PET and HER2 PET were performed at baseline, after 1 cycle of HP, and after 6 cycles of TCbHP, to assess the effectiveness of these examinations in the prediction of pCR and correlation to disease burden. All patients were received surgery and completed 1 year of HP-based regimen in the adjuvant setting. Results: A total of 61 patients with a median age of 49 years were included. Among them, 93.5% of patients are node positive. 75.8% of the patients reported adverse events (AEs), primarily grade 1 and 2. The incidence of serious AEs was 6.6%. Patients exhibiting a greater than 40% reduction in SUVmax of 18-FDG-PET or HER2-PET scans from baseline to the fifth day after the initial HP treatment were classified as the treatment-sensitive group. The pCR rate for the 18-FDG-PET-sensitive group was 79.3%, while for the insensitive group was 48.1%. For HER2-PET scans, the pCR rate was 70.0% in the sensitive group and 59.3% in the non-sensitive group. To evaluate the potential biomarkers, 25 patients (13 pCR and 12 non-pCR) were evaluated with FoundationOne, with the identification of 1,321 somatic alterations. The most frequently altered genes were ERBB2 (100%), TP53 (96%), CDK12 (76%), MYC (56%), CCND1 (44%), PIK3CA (40%), and SRSF2 (36%). In the pCR group, we found PPM1D (38.5%) and RAD21 (38.5%) were altered more frequently than the non-pCR group. PIK3CA alterations were significantly reduced in pCR group, only 23% of cases are affected compared to 58.3% in non-pCR group. Patients were non-pCR had more MDM4 (41.7%) and ZNF217 (41.7%) alterations than pCR patients. Conclusion: The SUVmax value of 18-FDG-PET and HER2-PET scans are correlated well with the tumor burden and the residual disease in the HER2 EBCs. The reduction of SUVmax could predict pCR but need further exploration. Patients with alterations in PPM1D and RAD21 have a favorable response to the TCbHP regimen. Conversely, alterations in PIK3CA, MDM4, and ZNF217 are associated with treatment resistance. These results indicate the significance of 18-FDG-PET and HER2-PET in monitoring treatment effectiveness. The biomarker analysis in this study will help us understand the mechanism of HER2 positive breast cancer treatment resistance and assisting in tailoring personalized therapy. Clinical trial information: NCT04281641. Citation Format: Ming Chen, Benlong Yang, Sun Yuyun, Linxiaoxi Ma, Song Shaoli, Min He, Zhi-Ming Shao, Bingqiu Xiu, Jiong Wu. An Open-label, Single-center Study Assessing the Efficacy of PH-based Regimen as Neoadjuvant Therapy for HER2-positive Early-stage Breast Cancer: Integrating Genomic Features and Functional Imaging [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-01-05.

Immunotherapy drives mesenchymal tumour cell state shift and TME immune response in glioblastoma patients.

Glioblastoma is a highly aggressive type of brain tumour for which there is no curative treatment available. Immunotherapies have shown limited responses in unselected patients, and there is an urgent need to identify mechanisms of treatment resistance to design novel therapy strategies. Here we investigated the phenotypic and transcriptional dynamics at single-cell resolution during nivolumab immune checkpoint treatment of glioblastoma patients. We present the integrative paired single-cell RNA-seq analysis of 76 tumour samples from patients in a clinical trial of the PD-1 inhibitor nivolumab and untreated patients. We identify a distinct aggressive phenotypic signature in both tumour cells and the tumour microenvironment in response to nivolumab. Moreover, nivolumab-treatment was associated with an increased transition to mesenchymal stem-like tumour cells, and an increase in TAMs and exhausted and proliferative T cells. We verify and extend our findings in large external glioblastoma dataset (n = 298), develop a latent immune signature and find 18% of primary glioblastoma samples to be latent immune, associated with mesenchymal tumour cell state and TME immune response. Finally, we show that latent immune glioblastoma patients are associated with shorter overall survival following immune checkpoint treatment (p = 0.0041). We find a resistance mechanism signature in a quarter of glioblastoma patients associated with a tumour-cell transition to a more aggressive mesenchymal-like state, increase in TAMs and proliferative and exhausted T cells in response to immunotherapy. These patients may instead benefit from neuro-oncology therapies targeting mesenchymal tumour cells.

Abstract PO5-05-05: Clinical outcomes of CDK4/6 inhibitors in patients with bone only metastatic breast cancer

Abstract Background: CDK4/6 inhibitors (CDK 4/6i) have significantly advanced the treatment of hormone receptor-positive (HR+) metastatic breast cancer in the last decade. Several clinical trials including PALOMA, MONALEESA, and MONARCH have shown improved progression-free survival (PFS) rates when CDK4/6i were combined with endocrine therapy (ET). These results led to recommending their use in combination with ET as the first line of therapy in the metastatic settings. Bone metastasis is the most common site of disease in patients with HR+ metastatic breast cancer, about 40% of whom have bone as the only site of metastasis (bone-only disease [BoD]). While previous studies have shown that BoD can have a unique clinical course and a more favorable prognosis compared to non-BoD metastasis, most of the aforementioned CDK 4/6i clinical trials have not shown significant improvements in PFS in BoD subset of patients. Here we aimed to investigate the survival outcomes of BoD patients at a single institution center and evaluate CDK 4/6i activity in preclinical models of bone metastasis in breast cancer. Patients and methods: Patients with HR+, HER2-negative metastatic breast cancer from a retrospective CDK4/6i study at Washington University School of Medicine were included in this analysis. We selected all the patients who received CDK4/6i in the first- or second-line settings from June 2014 to March 2020. Outcomes were collected through June 10, 2023. The Kaplan-Meier method with log rank test and multivariable cox model were used for survival analysis. Results: 114 female patients (n=75 first line, n=39 second line, n=20 African American, n=94 Caucasian) were included in this study. All patients received Palbociclib as the first exposure to CDK4/6 inhibitor in combination with ET for at least two months. Median age was 60 years (range, 28-87) and forty-seven patients (41%) were in the BoD group. Survival outcomes for BoD group were compared to the rest of patients who had either bone and visceral or visceral only metastatic lesions (BV group). Our results demonstrated a significantly reduced median PFS in BoD group compared to BV group (13.1 months [95%CI:8-18.2] vs 19.6 months [95%CI:6.8-32.4], p=0.01) respectively. There was no significant difference in overall survival between BoD and BV groups (36.6 months [95%CI:32.2-41] vs 41.7 months [95%CI 36.1-47.2] p=0.74) respectively. In multivariate analysis regarding the associated risk factors with PFS, after adjusting for age and race, BoD had a HR of 1.62 (95%CI:1.04-2.54) for PFS. Among the 75 patients who received CDK4/6i as their first line of therapy, BoD (n=32 patients [42.7%]) also showed an inferior (p=0.05) median PFS (20 months [95%CI 12.9-27]) compared to BV group (n=43 patients [57.3%], median PFS=28.4 months [95%CI: 16.3-40.4]). To investigate potential treatment resistance mechanisms in bone metastases, we used the triple negative 4T1 and the ER+ luminal B PyMT Bo1 breast tumor cell line derived mammary gland models in immunocompetent mice. Single agent CDK4/6i, ribociclib, decreased primary mammary gland tumor burden. This effect was abrogated when CD8+ T-cells were depleted. In the bone colonization model using these cell lines, ribociclib had little effect on bone tumor burden. Arg1+ immune suppressive macrophages were significantly decreased in the primary mammary tumor but not in the bone tumor after ribociclib administration, suggesting a relative treatment resistance and immune suppression in the bone tumor microenvironment compared to the primary mammary gland. Conclusion: CDK4/6i play a significant role in the treatment of patients with HR+ metastatic breast cancer. Our single institution, retrospective study and preclinical data suggests that CDK4/6i may not be as effective for the treatment of tumors residing in the bone microenvironment, warranting future studies to understand the mechanisms of CDK4/6 inhibitors action in bone versus visceral metastases. Citation Format: Mahta Mardani, Suleyman Noordeen, Katherine Clifton, Cynthia Ma, Jingqin Luo, Jing Xi, Nusayba Bagegni, Foluso Ademuyiwa, Rama Suresh, Ashley Frith, Andrew Davis, Ron Bose, Lindsay Peterson, Shana Thomas, Yu Tao, Takayuki Kobayashi, Jingyu Xiang, Yalin Xu, Xinming Su, Katherine Weilbaecher. Clinical outcomes of CDK4/6 inhibitors in patients with bone only metastatic breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-05-05.

Rapid autopsies to enhance metastatic research: the UPTIDER post-mortem tissue donation program

Research on metastatic cancer has been hampered by limited sample availability. Here we present the breast cancer post-mortem tissue donation program UPTIDER and show how it enabled sampling of a median of 31 (range: 5-90) metastases and 5-8 liquids per patient from its first 20 patients. In a dedicated experiment, we show the mild impact of increasing time after death on RNA quality, transcriptional profiles and immunohistochemical staining in tumor tissue samples. We show that this impact can be counteracted by organ cooling. We successfully generated ex vivo models from tissue and liquid biopsies from distinct histological subtypes of breast cancer. We anticipate these and future findings of UPTIDER to elucidate mechanisms of disease progression and treatment resistance and to provide tools for the exploration of precision medicine strategies in the metastatic setting.

Abstract 7228: In-vitro efficacy of Dordaviprone/GSK126 combination therapy on castration resistant and neuroendocrine prostate cancer

Abstract Neuroendocrine prostate cancer (NEPC) is an aggressive histologic subtype associated with poor prognosis that commonly arises in later stages as a mechanism of treatment resistance. Activation of oncogenic drivers, in combination with epigenetic changes (such as EZH2 overexpression and DNA methylation) further promotes tumor proliferation and expression of downstream neuroendocrine lineage pathways (in part controlled by transcription factors, including SOX2, ASCL1, and BRN2). Importantly, EZH2 inhibitors (EZH2i) can restore AR expression in CRPC (Ku et al., 2017). Imipridones, including Dordaviprone (ONC201), show promise in treating neuroendocrine tumors by interacting with DRD2 and CLpP (Anderson et al., 2022). These results warrant investigation into the potential for synergism in imipridone/EZH2i combination therapies. In the PCa cell lines LNCap, PC3, 22Rv1, and DU145 the combination of ONC201 and EZH2i GSK126 was assessed. Cell viability data after treatment with ONC201 and GSK126 reveals several dose ranges with potential synergistic activity after 72h. Preliminary western blots after 72 hours of GSK126 treatment indicate modulation of proteins relevant to the mechanism of action of imipridones, specifically DR5 and CLpP, in LNCap, PC3, and 22Rv1. Notably, the decrease in DR5 expression warrants investigation into the effects of EZH2i on TRAIL sensitivity and whether imipridones may rescue DR5 expression to maintain TRAIL sensitivity. Subsequent experiments will investigate dosing and time point-based effects on viability and protein expression, in addition to assessing the synergy of other imipridones and EZH2i’s. Further, while the cell lines PC3, 22Rv1, and DU145 are castration-resistant, the neuroendocrine prostate cell line NCI-H660 will be used as a positive control as it expresses higher neuroendocrine features, including DRD2. Additionally, the modulation in DR5 expression warrants inquiry into the effects of combination therapy on tumor-immune cell interactions. Further, we will manipulate the expression of neuroendocrine transcription factors to assess changes in therapy sensitivity. Citation Format: Connor Purcell, Praveen Srinivasan, Maximilian Pinho-Schwermann, William MacDonald, Elizabeth C. Ding, Vida Tajiknia, Wafik El-Deiry. In-vitro efficacy of Dordaviprone/GSK126 combination therapy on castration resistant and neuroendocrine prostate cancer [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 7228.

Abstract 6243: Rapid autopsy reveals the interplay of tumor heterogeneity at genomic, transcriptomic, and spatial dimensions

Abstract Incomplete understanding of tumor heterogeneity across primary and metastatic sites often lead to ineffective cancer treatments. To comprehensively interrogate tumor heterogeneity at genomic, transcriptomic, and spatial levels, we established a novel rapid autopsy protocol to obtain multiregional tri-omic data consisted of whole exome sequencing, mRNA-sequencing, and multiplexed immunohistochemistry on primary and metastatic tumors and associated microenvironments (TMEs). We applied this protocol to six patients with various primary tumor types: non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), urothelial cancer, and one mixed case with both NSCLC and SCLC. A total of 50 tumor samples with paired tri-omic data were obtained across 16 different primary and metastatic organs to interrogate heterogeneities underlying tumor evolution and treatment resistance mechanisms. We constructed mutation-based phylogeny trees to reveal structural differences in tumor evolution trajectories across patients and infer heterogeneous tumor antigen landscape. We uncovered highly entropic phylogenetic structures having unique variants gained at each metastasis as well as low entropic structures with high sharing across sites. The mixed case had a clonal sweep structure with two metastases developed prior to the rest of the metastasis. In this case, older clones were shown to have strong association with APOBEC mutation signatures, enhanced subclonal mutation burden and loss of human leukocyte antigen heterozygosity as mechanisms of immune evasion. Integrating transcriptome and spatial profiles, we revealed that tumor intrinsic expressions and extrinsic immune infiltration levels covary with tissue types rather than with mutation-based phylogenetic clusters. Across patients, samples that had high similarities in mutational landscape varied in their pro-tumor expression programs and the cellular composition of TMEs. In particular, lung samples across patients had the highest effector T cell population and major histocompatibility complex (MHC) class I, II presentations compared to other tissue types. Contrasting small cell carcinoma (SCC) and adenocarcinoma (Adeno) samples from lung tissues further revealed that SCC had more immunosuppressive TMEs with downregulation of MHC class II. Liver metastasis samples were further characterized to have the most aggressive phenotypes enriched in genomic instability measured by aneuploidy compared to other metastatic sites. In sum, we present a novel technology and computational framework that enables a three-dimensional profiling of tumor heterogeneity. This work suggests that single-site biopsies are insufficient to capture tumor heterogeneity and calls for multi-modal integrations to design treatments that can overcome tumor-specific resistance mechanisms. Citation Format: Ning Yao, Rami Vanguri, Juber Patel, Jayon Lihm, Andrew Fisher, Irene Jarchum, Shigeaki Umeda, Hao Li, Scott Ely, Travis Hollmann, Christine Iacobuzio-Donahue, Benjamin Greenbaum. Rapid autopsy reveals the interplay of tumor heterogeneity at genomic, transcriptomic, and spatial dimensions [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 6243.

Abstract 4638: Integrated proteogenomic characterization of longitudinal glioblastoma

Abstract Glioblastoma multiforme (GBM) is the most aggressive form of primary brain tumor, with no curative treatment options and median patient survival time of almost one year. Despite multi-modal therapy including surgery, irradiation and chemotherapy, all patients experience tumor progression and recurrence. Our group proposed and validated the first single cell guided functional classification of primary GBM in four tumor-intrinsic cell states which informed clinical outcome and delivered therapeutic options. However, recurrent GBM remains therapeutically unresolved due in part to the diffusely invasive nature and in part to marked cellular heterogeneity of the tumor.The evolutionary trajectory of glioblastoma after therapy is a multifaceted biological process that extends beyond discrete genetic alterations alone. Here, we profiled by multi-omics platforms the largest dataset of matched primary and recurrent GBM including 123 longitudinal glioblastoma pairs, temporally separated by standard-of-care treatment. Genomics, proteomics, and phosphoproteomics all independently captured the loss of proliferative-progenitor characteristics and a significant upregulation of specialized neuronal and synaptic signaling programs in recurrent GBM. Proteomic and phosphoproteomic analyses revealed that the molecular transition from proliferative to neuronal state at recurrence is marked by coherent post-translational activation of the WNT/PCP signaling pathway and the BRAF protein kinase. Multi-omic analysis of Patient-Derived Xenograft (PDX) models mimicked the patterns of evolutionary trajectory, consisting of marked activation of neuronal signaling programs in recurrent patients. Inhibition of the BRAF kinase with small molecule inhibitors impaired both neuronal transition and migration capability of recurrent glioblastoma cells, which are the phenotypic hallmarks of glioblastoma progression after therapy. Accordingly, combinatorial treatment of temozolomide with BRAF inhibitor, vemurafenib, significantly prolonged the survival of mouse PDX models. This work provides comprehensive insights into the biological mechanisms of glioblastoma evolution and treatment resistance and highlights new therapeutic opportunities to effectively counter them in the clinic. Citation Format: Simona Migliozzi, Kyung-Hee Kim, Harim Koo, Jun-Hee Hong, Seung Min Park, Hyung Joon Kwon, Luciano Garofano, Young Taek Oh, Fulvio D'Angelo, Chan Il Kim, Anna-Luisa Di Stefano, Franck Bielle, Jinlong Yin, Marc Sanson, Do-Hyun Nam, Jason K. Sa, Anna Lasorella, Jong Bae Park, Antonio Iavarone. Integrated proteogenomic characterization of longitudinal glioblastoma [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 4638.

Abstract 1121: Targeting the DNA damage response in high-risk medulloblastoma

Abstract Medulloblastoma (MB) is the most common malignant pediatric brain tumour and is currently treated with surgery, external beam radiation and chemotherapy however, current treatments are woefully inadequate, with survival of only 50% but morbidity approaching 100%. A very-high risk group of SHH-activated MB harboring TP53 mutations (TP53mut-SHH) account for a substantial proportion of treatment failures after radiotherapy with a uniformly fatal outcome. Through optimizing radiation as a selection pressure and employing a functional genetic approach through whole-genome CrisprCas-9 dropout screening, the present study aims to identify drivers of radiation resistance and uncover unique vulnerabilities to sensitize these tumours to therapy. The dropout screen was conducted across a panel of established in vitro models of sporadic murine TP53mut-SHH, which resemble the human disease with profound radio-resistance. Initial results reveal that targeting the DNA damage response confer radiation sensitization. However, consistent hits across all models demonstrate non-homologous end-joining (NHEJ) as being synthetically lethal with radiation specifically in TP53mut-SHH. Genetic knockout experiments confirm genes encoding main proteins involved in NHEJ are synthetically lethal in combination with radiation. A small molecule drug screen targeting NHEJ revealed such vulnerabilities may be therapeutically exploited in TP53mut-SHH and are able sensitize these tumours to radiation. Using our unique approach of leveraging newly established models of TP53mut-SHH with a combination of functional genetic screening and small-molecule drug screening, this study highlights the genetic mechanisms of treatment resistance and reveals novel approaches for radiation sensitization. Here, we develop a novel approach to identify targets that are synthetically lethal with radiotherapy and identify multiple synthetically lethal targets at the genetic level which can be therapeutically targeted, allowing the development of new treatments to overcome current barriers. Citation Format: Alexandria DeCarlo, Carolina Fernandes da Silva, Lily Shen, Vijay Ramaswamy, Uri Tabori. Targeting the DNA damage response in high-risk medulloblastoma [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 1121.

Abstract 3580: Understanding and overcoming innate and acquired resistance to type I and II RAF inhibitors in anaplastic thyroid cancer using translational functional genomics

Abstract Anaplastic thyroid cancer (ATC) is one of the most lethal human cancers, with some patients succumbing to the disease within weeks of diagnosis. Despite a subset of BRAFV600E mutant ATC patients responding to monomeric type I RAF inhibitor (RAFi) dabrafenib in combination with MEK inhibitor (MEKi) trametinib, almost all patients rapidly develop adaptive or acquired resistance. These patients, along with those who do not harbor the BRAFV600E alteration, have limited treatment options. To understand the mechanism of resistance to dabrafenib and trametinib, we utilized multi-region whole genome sequencing and single nuclei profiling of ATC patient tumors to unravel genomic, transcriptomic, and microenvironmental evolution during type I RAFi and MEKi therapy. Single-cell nuclei sequencing of matched primary and resistant ATC patient tumors identified transcriptomic reactivation of MAPK-pathway, along with immunosuppressive macrophage proliferation, underlie the development of acquired resistance. Our translation genomics led us to hypothesize that deeper inhibition of the MAPK-pathway can be efficacious in overcoming treatment resistance. Screening of a panel of type II RAFis revealed that ATC cell lines are exquisitely sensitive to the type II RAFi naporafenib. We further demonstrate that naporafenib in combination with MEKi trametinib can durably and robustly overcome both innate and acquired treatment resistance to dabrafenib and trametinib using ATC cell lines and patient-derived xenograft models, including in a PDX model from an ATC patient who developed acquired resistance to dabrafenib and trametinib. Finally, we describe a novel mechanism of acquired resistance to type II RAF inhibitor and MEK inhibitor through compensatory mutations in MAST1. Taken together, our work using translational and functional genomics have unraveled the differential mechanisms of treatment resistance to type I and type II RAFi in combination with trametinib, and rationalizes the clinical investigation of type II RAFi in the setting of thyroid cancer. Citation Format: Peter Zeng, Jalna Meens, John Barrett, Matthew J. Cecchini, Sarah B. Ryan, Nachuan Pan, Amir Karimi, Laura Jarycki, Alice E. Dawson, Mushfiq Shaikh, David Palma, Eric Winquist, Paul C. Boutros, Laurie Ailles, Anthony C. Nichols. Understanding and overcoming innate and acquired resistance to type I and II RAF inhibitors in anaplastic thyroid cancer using translational functional genomics [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 3580.

Abstract 7057: Validation of OXPHOS gene amplification as a driver of treatment resistance in non-small cell lung cancer

Abstract Lung cancer is the leading cause of cancer deaths in the U.S. and worldwide. Up to 90% of all lung cancers are non-small cell lung cancer (NSCLC). NSCLC is a complex and highly aggressive disease typical for poor prognosis and high rates of treatment resistance. Clinical management of NSCLC varies according to the stage. High-dose stereotactic body radiation therapy (SBRT) is the standard of care for localized non-resectable NSCLC. However, tumor hypoxia is a significant barrier to effective radiation therapy. In the last decade, first line NSCLC treatment has been substantially reinforced with the introduction of immunotherapy targeting immune checkpoints such as programmed cell death 1 (PD-1), yet long-term disease control occurs in less than 25% of NSCLC patients. Therefore, understanding the mechanisms of treatment resistance is essential to address the dire need of introducing novel synergistic therapies to enhance anti-cancer treatment response in refractory NSCLC tumors. The basis of tumor hypoxia has traditionally been attributed to the oxygen supply deficit as malformed tumor vasculature fails to meet the high demand of the rapidly proliferating tumor mass. However, our preliminary analysis of NSCLC patient datasets in the Cancer Genome Atlas (TCGA) PanCancer dataset revealed a significant correlation between high-level expression of nuclear genes encoding mitochondrial subunits essential for oxidative phosphorylation (OXPHOS), and high-level expression of hypoxia-regulated genes (Buffa hypoxia score). Furthermore, we have observed a direct correlation between copy number amplification (CNA) of several OXPHOS genes and hypoxia levels in patient samples. Because mitochondrial function consumes up to 90% of available cellular oxygen, its activity may be indirectly regulating oxygen availability in the tumor microenvironment by rapidly consuming oxygen upon its delivery to the tumor. Moreover, we have identified that almost 40% of NSCLC patients harbor CNA of these essential OXPHOS genes. To provide mechanistic insight into these clinical observations, we have generated mouse syngeneic NSCLC cell lines with modified mRNA expression levels of OXPHOS genes (representing deletion, diploid and amplified number of copies) identified in our screen and observed a direct effect on the rates of mitochondrial oxygen consumption in these cells. Our study investigates the previously unknown role of OXPHOS gene copy number in driving mitochondrial oxygen demand and thus contributing to tumor hypoxia and treatment resistance in NSCLC. Citation Format: Martin Benej, Katarina Benejova, Rebecca Hoyd, Caroline Wheeler, Daniel Spakowicz, Nicholas C. Denko. Validation of OXPHOS gene amplification as a driver of treatment resistance in non-small cell lung cancer [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 7057.

Abstract 6120: Genomic profiling of KRAS and EGFR-altered non-squamous non-small cell lung cancer reveal ancestry-specific co-alterations with therapeutic implications

Abstract Background: Racial and ethnic disparities are highly prevalent in cancer care and impact treatment outcomes, with an underrepresentation of minority populations in clinical studies. The impact of genetic ancestry on the genomic landscape of tumors remains understudied. To address this, we sought to comprehensively characterize the ancestry-based genomic co-alteration landscape and immunotherapy-related biomarkers in KRAS and EGFR-altered tumors, two major driver populations in non-squamous non-small cell lung cancer (non-Sq NSCLC). Methods: Our study consisted of 68,197 adult patients with non-Sq NSCLC who underwent comprehensive genomic profiling using FoundationOne® or FoundationOne®CDx during routine clinical care in the United States. Genetic ancestry was inferred using a SNP-based approach. Tumor mutational burden (TMB) was calculated across 0.8-1.2 megabases. PD-L1 expression was determined by immunohistochemistry using the Dako 22C3 PD-L1 antibody. Results: Overall, 81% of the patients were of European (EUR; n= 55,430), 10% of African (AFR; n=7,062), 5% of East Asian (EAS; n=3,297), 3% of Admixed American (AMR; n=2,011) and less than 1% of South Asian ancestry (SAS; n=497). KRAS was the most frequently altered oncogene in EUR (39%) and AFR (33%), whereas EGFR was most frequently altered in EAS (53%), SAS (36%), and AMR (30%) ancestry groups. While STK11 and KEAP1 alterations co-occurred with KRAS across all ancestry groups, they were significantly (FDR p <= 0.05) less frequent in EAS (16%) and AMR (15%) compared to EUR (28%). Additional ancestry-specific co-alteration patterns in KRAS-altered tumors included co-occurrence with GNAS alterations in AMR (Odds ratio, OR: 3.5, p < 10-5), co-occurrence with ARID1A alterations in SAS (OR: 4.7, p = 0.02), and mutual exclusivity with NF1 alterations in EUR (OR: 0.4, p < 10-5) and AFR (OR: 0.4, p < 10-5). In contrast, EGFR-altered tumors had a more conserved co-alteration landscape across all ancestry groups. Despite the known mutual exclusivity of KRAS and EGFR alterations, 13% of patients of EAS ancestry (compared to 2-4% of other ancestry groups) with KRAS alterations had co-occurring EGFR alterations; notably, a majority of these patients had an amplification in KRAS and/or EGFR, potentially representing treatment resistance mechanisms. Among immunotherapy-associated biomarkers, PD-L1 expression was similar across ancestries. Of note, patients of AFR ancestry with KRAS or EGFR alterations had higher TMB and the SAS ancestry group had the lowest TMB. Conclusion: Our study provides a comprehensive landscape of ancestry-specific patterns in KRAS and EGFR-altered non-Sq NSCLC. These findings can help better understand cancer disparities, aid in the development of new therapeutic strategies and inform more inclusive clinical trials and treatment decisions. Citation Format: Saumya D. Sisoudiya, Armande A. Houle, Tharu M. Fernando, Timothy R. Wilson, Jennifer L. Schutzman, Jessica K. Lee, Alexa B. Schrock, Ethan S. Sokol, Smruthy Sivakumar, Zhen Shi, Gaurav Pathria. Genomic profiling of KRAS and EGFR-altered non-squamous non-small cell lung cancer reveal ancestry-specific co-alterations with therapeutic implications [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 6120.

Novel prognostic biomarker TBC1D1 is associated with immunotherapy resistance in gliomas.

Glioma, an aggressive brain tumor, poses a challenge in understanding the mechanisms of treatment resistance, despite promising results from immunotherapy. We identified genes associated with immunotherapy resistance through an analysis of The Cancer Genome Atlas (TCGA), Chinese Glioma Genome Atlas (CGGA), and Gene Expression Omnibus (GEO) databases. Subsequently, qRT-PCR and western blot analyses were conducted to measure the mRNA and protein levels of TBC1 Domain Family Member 1 (TBC1D1), respectively. Additionally, Gene Set Enrichment Analysis (GSEA) was employed to reveal relevant signaling pathways, and the expression of TBC1D1 in immune cells was analyzed using single-cell RNA sequencing (scRNA-seq) data from GEO database. Tumor Immune Dysfunction and Exclusion (TIDE) database was utilized to assess T-cell function, while Tumor Immunotherapy Gene Expression Resource (TIGER) database was employed to evaluate immunotherapy resistance in relation to TBC1D1. Furthermore, the predictive performance of molecules on prognosis was assessed using Kaplan-Meier plots, nomograms, and ROC curves. The levels of TBC1D1 were significantly elevated in tumor tissue from glioma patients. Furthermore, high TBC1D1 expression was observed in macrophages compared to other cells, which negatively impacted T cell function, impaired immunotherapy response, promoted treatment tolerance, and led to poor prognosis. Inhibition of TBC1D1 was found to potentially synergistically enhance the efficacy of immunotherapy and prolong the survival of cancer patients with gliomas. Heightened expression of TBC1D1 may facilitate an immunosuppressive microenvironment and predict a poor prognosis. BlockingTBC1D1 could minimize immunotherapy resistance in cancer patients with gliomas.

Abstract B083: Single-cell and spatial transcriptomic characterization of treatment resistance and immune dynamics in high-grade serous ovarian cancer

Abstract B083: Single-cell and spatial transcriptomic characterization of treatment resistance and immune dynamics in high-grade serous ovarian cancer

Exploring androgen receptor signaling pathway in prostate cancer: A path to new discoveries.

Androgen deprivation therapy has achieved significant success in treating prostate cancer through strategies centered on the androgen receptor. However, the emergence of castration-resistant prostate cancer highlights this therapy limitation, underscoring the need to elucidate the mechanisms of treatment resistance. This review aimed to focus on multifaceted resistance mechanisms, including androgen receptor overexpression, splice variants, missense mutations, the involvement of the glucocorticoid receptor, and alterations in coregulators and transcription factors, revealing their roles in castration-resistant prostate cancer progression. These mechanisms promote cell survival and proliferation, depending on the androgen receptor signaling pathway, leading to resistance to conventional therapies. Amplification and mutations in the androgen receptor gene facilitate selective adaptation in treatment-resistant cells, consequently diminishing therapeutic efficacy. Furthermore, the activation of glucocorticoid receptors and aberrant regulation of specific coregulators and transcription factors contribute to the activation of androgen receptor-independent signaling pathways, promoting cell survival and proliferation. These findings hold promise for identifying new targets for treating castration-resistant prostate cancer and developing personalized treatment strategies. The development of future therapies will hinge on precisely targeting the androgen receptor signaling pathway, necessitating a deeper understanding of the molecular targets unique to castration-resistant prostate cancer.

IDHwt glioblastomas can be stratified by their transcriptional response to standard treatment, with implications for targeted therapy

BackgroundGlioblastoma (GBM) brain tumors lacking IDH1 mutations (IDHwt) have the worst prognosis of all brain neoplasms. Patients receive surgery and chemoradiotherapy but tumors almost always fatally recur.ResultsUsing RNA sequencing data from 107 pairs of pre- and post-standard treatment locally recurrent IDHwt GBM tumors, we identify two responder subtypes based on longitudinal changes in gene expression. In two thirds of patients, a specific subset of genes is upregulated from primary to recurrence (Up responders), and in one third, the same genes are downregulated (Down responders), specifically in neoplastic cells. Characterization of the responder subtypes indicates subtype-specific adaptive treatment resistance mechanisms that are associated with distinct changes in the tumor microenvironment. In Up responders, recurrent tumors are enriched in quiescent proneural GBM stem cells and differentiated neoplastic cells, with increased interaction with the surrounding normal brain and neurotransmitter signaling, whereas Down responders commonly undergo mesenchymal transition. ChIP-sequencing data from longitudinal GBM tumors suggests that the observed transcriptional reprogramming could be driven by Polycomb-based chromatin remodeling rather than DNA methylation.ConclusionsWe show that the responder subtype is cancer-cell intrinsic, recapitulated in in vitro GBM cell models, and influenced by the presence of the tumor microenvironment. Stratifying GBM tumors by responder subtype may lead to more effective treatment.

Abstract PR06: Uncovering nuclear expulsion: chromatin-bound signals from dying cancer cells accelerate metastatic outgrowth through S100A4-RAGE pathway

Abstract Tumor cells are highly heterogeneous, exhibiting diverse responses when exposed to different environments. We have recently observed this heterogeneity in tumor cell apoptosis by which some tumor cell die for the benefits of the population’s survival. Majority of tumor cells undergo apoptosis in response to immune surveillance, various treatments, and a hostile microenvironment, both in circulation and at metastatic organ sites. However, the full range of apoptotic signals, or "last words," that dying cells communicate to the surviving cancer cell community remains incompletely understood. While apoptotic cell death is often perceived as a favorable therapeutic outcome, tumor cells can exploit it to enhance their metastatic capabilities, leading to poor prognoses in certain cancers. Here, we present a remarkable discovery of an altered apoptotic program in tumor cells characterized by nuclear expulsion and citrullinated chromatin, promoting metastatic outgrowth in the lungs in a Padi4-dependent manner. We further identified a calcium spike during apoptosis as the trigger for Padi4 activation, resulting in rapid chromatin de-condensation, nuclear content expulsion, followed by the generation of nuclear expulsion products (NEPs). This process was observed in various types of human cancer tissues. Profiling of proteins bound to the NEP complex unveiled an enrichment of receptor for advanced glycation end-products (RAGE) pathway proteins, such as S100a4 in mouse NEPs and high-mobility group-binding (HMGB) proteins in human NEPs. These chromatin-bound RAGE ligands activated RAGE receptors in neighboring surviving tumor cells, leading to Erk pathway activation. This, in turn, promotes proliferation through the RAGE-MAPK pathway. A nuclear expulsion signature was associated with worse prognoses in cancer patients, underscoring the clinical efficacy in the context of apoptosis-inducing treatments. As we continue our investigations, we are particularly interested in understanding how dying tumor cell-derived NEPs from chemotherapy may affect therapy outcomes. Taken together, our studies uncover a novel mechanism by which tumor cells come together as a community and that the apoptotic tumor cells help live tumor cells metastasize. This insight is likely a step forward to understand the mechanisms of metastatic spread, treatment resistance and relapse. Citation Format: Wooyong Park, Justin M Gray, Ronald J Holewinski, Thorkell Andresson, Jae Young So, Carmelo M Carmona-Rivera, Mariana J Kaplan, Steven D Cappell, Li Yang. Uncovering nuclear expulsion: chromatin-bound signals from dying cancer cells accelerate metastatic outgrowth through S100A4-RAGE pathway [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr PR06.

Mechanisms of Melanoma Progression and Treatment Resistance: Role of Cancer Stem-like Cells.

Melanoma is the third most common type of skin cancer, characterized by its heterogeneity and propensity to metastasize to distant organs. Melanoma is a heterogeneous tumor, composed of genetically divergent subpopulations, including a small fraction of melanoma-initiating cancer stem-like cells (CSCs) and many non-cancer stem cells (non-CSCs). CSCs are characterized by their unique surface proteins associated with aberrant signaling pathways with a causal or consequential relationship with tumor progression, drug resistance, and recurrence. Melanomas also harbor significant alterations in functional genes (BRAF, CDKN2A, NRAS, TP53, and NF1). Of these, the most common are the BRAF and NRAS oncogenes, with 50% of melanomas demonstrating the BRAF mutation (BRAFV600E). While the successful targeting of BRAFV600E does improve overall survival, the long-term efficacy of available therapeutic options is limited due to adverse side effects and reduced clinical efficacy. Additionally, drug resistance develops rapidly via mechanisms involving fast feedback re-activation of MAPK signaling pathways. This article updates information relevant to the mechanisms of melanoma progression and resistance and particularly the mechanistic role of CSCs in melanoma progression, drug resistance, and recurrence.

Abstract B021: Clonal heterogeneity in human pancreatic adenocarcinoma and its contribution to therapeutic resistance

Abstract PDAC is a highly aggressive cancer with poor prognosis, largely due to its late diagnosis, rapid progression, and lack of responsiveness to standard therapies. The advent of large-scale genomics, single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics (ST) has illuminated potential genomic targets, the intricate role of the PDAC tumor microenvironment (TME), and gene activity orchestrating cellular arrangements within cancer tissues. In recent work by our group, we characterized the composition of human PDAC at the single-cell level, revealing significant intra-tumoral heterogeneity in transcriptional subtypes and TME composition. We also found that chemotherapy profoundly alters the PDAC TME and may lead to further resistance to immunotherapy by reduced inhibitory checkpoint molecule expression and interactions involving CD8+ T cells. We have since determined that specific clones in human PDAC tumors have unique influences on the biological and clinical features of PDAC. In this study, we tested the hypothesis that the distinct genomic profiles of individual clones within PDAC tumors influence their transcriptional state and their interaction with the TME, ultimately governing disease progression and response to chemotherapy. scRNA-seq analysis was performed on 61 freshly collected human primary PDAC samples from 58 patients, either prior to or following chemotherapy treatment, including 3 matched pre- and post- treatment pairs from the same patient. CNV analysis of PDAC samples revealed significant inter- and intra- tumoral clonal heterogeneity. Using the results of differential gene expression and gene set enrichment analysis, we identified a set of common transcriptional states associated with specific CNV profiles. Further, we found that specific PDAC clones often correlated with classical and basal subtypes. Using a combination of ligand-receptor-interaction (LRI) analysis and spatial transcriptomics, we observed that distinct clones within tumors exhibited heterogeneous interactions with different cell types in the TME, and that the composition of individual clones within tumors and their LRI were significantly and differentially altered in response to chemotherapy. Our findings indicate the presence of distinct clonal neighborhoods in PDAC, and allow interrogation of clonal and cell communication network changes that occur in response to chemotherapy. This data provide new insight into potential mechanisms of treatment resistance and may help identify potential strategies to intercept or more effectively treat metastasis and improve patient outcomes. Citation Format: Despoina Kalfakakou, Emily A. Kawaler, Dylan Tamayo, Lidong Wang, Rosmel Hernandez, Amanda Ackermann, Xiaohong Jing, Cristina Hajdu, Cynthia Loomis, Adrianna Heguy, Amanda W. Lund, Theodore H. Welling, Igor Dolgalev, Aristotelis Tsirigos, Diane M. Simeone. Clonal heterogeneity in human pancreatic adenocarcinoma and its contribution to therapeutic resistance [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr B021.