Pub Date : 2024-09-13DOI: 10.1101/2024.09.09.609499
Katharine J Herbert, Rosie Upstill-Goddard, Stephan B Dreyer, Selma Rebus, Australian Pancreatic Cancer Genome Initiative, Christian Pilarsky, Debabrata Mukhopadhyay, Christopher J. Lord, Genomics Innovation Alliance, Andrew V Biankin, Fieke E.M. Froeling, David K Chang
Pancreatic ductal adenocarcinoma (PDAC) remains the most lethal cancer and will soon be the second most common cause of cancer related death. While regimens containing DNA damaging agents such as FOLFIRINOX and PARP inhibitors have derived clinical benefits for some patients, their efficacy invariably fails over time. This presents a significant clinical challenge, and thus there is an urgent need for novel therapeutic strategies which are able to overcome the acquisition of resistance in PDAC. Clinically relevant models of treatment resistance were generated from patient-derived cell lines by extended exposure to chemotherapy agents. Synergy scoring, clonogenicity assays, flow cytometry, immunofluorescence and transcriptomic analysis were used to investigate the efficacy of combined ATR and PARP inhibition in re-sensitising resistant PDAC to treatment. Acquisition of resistance was associated with transcriptomic shifts in cell cycle checkpoint regulation, metabolic control, DNA damage response (DDR), programmed cell death and the replication stress response. Additionally, combined treatment with the ATR inhibitor (ceralasertib), and the PARP inhibitor (olaparib) was synergistic in all models of acquired resistance. Sequential treatment using ceralasertib prior to olaparib was highly effective at low dose for DDR proficient cell lines, whereas DDR deficient models responded better when treated with olaparib first. We provide in vitro evidence of a novel therapeutic strategy to overcome acquired PARP inhibitor and platinum resistance in PDAC by using sequential exposure to ceralasertib and olaparib. A sequential regimen may be more tolerable and should be investigated clinically to circumvent dose limiting toxicity in concurrent combinations.
胰腺导管腺癌（PDAC）仍然是致死率最高的癌症，很快将成为癌症相关死亡的第二大常见原因。虽然含有 DNA 损伤剂（如 FOLFIRINOX 和 PARP 抑制剂）的治疗方案为一些患者带来了临床疗效，但随着时间的推移，这些方案的疗效总会出现衰退。这给临床治疗带来了巨大挑战，因此迫切需要能够克服 PDAC 耐药性的新型治疗策略。通过协同作用评分、克隆生成测定、流式细胞术、免疫荧光和转录组学分析，研究了联合抑制ATR和PARP对耐药PDAC治疗再敏感的疗效。耐药性的获得与细胞周期检查点调控、代谢控制、DNA损伤反应（DDR）、程序性细胞死亡和复制应激反应的转录组学转变有关。此外，在所有获得性耐药模型中，ATR抑制剂（ceralasertib）和PARP抑制剂（olaparib）的联合治疗具有协同作用。我们通过体外实验证明了一种新的治疗策略，即通过连续暴露于ceralasertib和奥拉帕利来克服PDAC中获得性PARP抑制剂和铂类耐药性。序贯疗法可能更耐受，应在临床上进行研究，以规避同时联合用药时的剂量限制毒性。
{"title":"Sequential ATR and PARP Inhibition Overcomes Acquired DNA Damaging Agent Resistance in Pancreatic Ductal Adenocarcinoma","authors":"Katharine J Herbert, Rosie Upstill-Goddard, Stephan B Dreyer, Selma Rebus, Australian Pancreatic Cancer Genome Initiative, Christian Pilarsky, Debabrata Mukhopadhyay, Christopher J. Lord, Genomics Innovation Alliance, Andrew V Biankin, Fieke E.M. Froeling, David K Chang","doi":"10.1101/2024.09.09.609499","DOIUrl":"https://doi.org/10.1101/2024.09.09.609499","url":null,"abstract":"Pancreatic ductal adenocarcinoma (PDAC) remains the most lethal cancer and will soon be the second most common cause of cancer related death. While regimens containing DNA damaging agents such as FOLFIRINOX and PARP inhibitors have derived clinical benefits for some patients, their efficacy invariably fails over time. This presents a significant clinical challenge, and thus there is an urgent need for novel therapeutic strategies which are able to overcome the acquisition of resistance in PDAC.\u0000Clinically relevant models of treatment resistance were generated from patient-derived cell lines by extended exposure to chemotherapy agents. Synergy scoring, clonogenicity assays, flow cytometry, immunofluorescence and transcriptomic analysis were used to investigate the efficacy of combined ATR and PARP inhibition in re-sensitising resistant PDAC to treatment.\u0000Acquisition of resistance was associated with transcriptomic shifts in cell cycle checkpoint regulation, metabolic control, DNA damage response (DDR), programmed cell death and the replication stress response. Additionally, combined treatment with the ATR inhibitor (ceralasertib), and the PARP inhibitor (olaparib) was synergistic in all models of acquired resistance. Sequential treatment using ceralasertib prior to olaparib was highly effective at low dose for DDR proficient cell lines, whereas DDR deficient models responded better when treated with olaparib first.\u0000We provide in vitro evidence of a novel therapeutic strategy to overcome acquired PARP inhibitor and platinum resistance in PDAC by using sequential exposure to ceralasertib and olaparib. A sequential regimen may be more tolerable and should be investigated clinically to circumvent dose limiting toxicity in concurrent combinations.","PeriodicalId":501233,"journal":{"name":"bioRxiv - Cancer Biology","volume":"74 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142210601","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Although a lot of effort has been dedicated to ovarian cancer (OC) research, the mortality rate is still among the highest in female gynecologic malignancies. The effects of the aged tumor microenvironment are still being undermined despite age being the highest risk factor in ovarian cancer development and progression. In this study, we have conducted RNA sequencing and lipidomics analysis of gonadal adipose tissues from young and aged rat xenografts before and after ovarian cancer formation. We have found significantly higher tumor formation rates and volumes in aged OC xenograft rat models compared to their young counterparts (p<0.05), suggesting the aged adipose microenvironment (AME) is more susceptible to OC outgrowth. We have revealed significant shifts in the gene expression enrichment from groups of young vs. aged rats before tumor formation, groups of young vs. aged rats when the tumor formed, and groups of aged rats before and after tumor formation. We also observed shifts in the lipid components of the gonadal adipose tissues between young and aged rat xenografts when tumors were generated. Additionally, we found that the aged AME was associated with age-related changes in the immune cell composition, especially inflammation-related cells. The top hits showing the most differences between aged and young adipose tissues were eight genes including S100a8, S100a9, Il1rl1, Lcn2, C3, Hba-a1, Fcna, and Pnpla3, 22 lipids including multiple isoforms of free fatty acids (FFA) and triglyceride (TG), as well as four immune cells including neutrophil, myeloid dendritic cell, T cell CD4+ (non-regulatory), and mast cell activation. The functional correlation among S100a8, S100a9, neutrophil, and FFA (18:3) was also determined. Furthermore, FFA (18:3), which was shown to be downregulated in aged xenograft rats, was capable of inhibiting OC cell proliferation. In conclusion, our study suggested that aging promoted OC proliferation through changes in genes/pathways, lipid metabolism, and immune cells. Targeting the aging adipose microenvironment, particularly lipid metabolism reprogramming, holds promise as a therapeutic strategy for OC, which warrants further investigation.
{"title":"Regulation of Age-Related Lipid Metabolism in Ovarian Cancer","authors":"Jihua Feng, Clay Douglas Rouse, Isabella Coogan, Olivia Byrd, Zhiqing Huang","doi":"10.1101/2024.09.06.611709","DOIUrl":"https://doi.org/10.1101/2024.09.06.611709","url":null,"abstract":"Although a lot of effort has been dedicated to ovarian cancer (OC) research, the mortality rate is still among the highest in female gynecologic malignancies. The effects of the aged tumor microenvironment are still being undermined despite age being the highest risk factor in ovarian cancer development and progression. In this study, we have conducted RNA sequencing and lipidomics analysis of gonadal adipose tissues from young and aged rat xenografts before and after ovarian cancer formation. We have found significantly higher tumor formation rates and volumes in aged OC xenograft rat models compared to their young counterparts (p<0.05), suggesting the aged adipose microenvironment (AME) is more susceptible to OC outgrowth. We have revealed significant shifts in the gene expression enrichment from groups of young vs. aged rats before tumor formation, groups of young vs. aged rats when the tumor formed, and groups of aged rats before and after tumor formation. We also observed shifts in the lipid components of the gonadal adipose tissues between young and aged rat xenografts when tumors were generated. Additionally, we found that the aged AME was associated with age-related changes in the immune cell composition, especially inflammation-related cells. The top hits showing the most differences between aged and young adipose tissues were eight genes including S100a8, S100a9, Il1rl1, Lcn2, C3, Hba-a1, Fcna, and Pnpla3, 22 lipids including multiple isoforms of free fatty acids (FFA) and triglyceride (TG), as well as four immune cells including neutrophil, myeloid dendritic cell, T cell CD4+ (non-regulatory), and mast cell activation. The functional correlation among S100a8, S100a9, neutrophil, and FFA (18:3) was also determined. Furthermore, FFA (18:3), which was shown to be downregulated in aged xenograft rats, was capable of inhibiting OC cell proliferation. In conclusion, our study suggested that aging promoted OC proliferation through changes in genes/pathways, lipid metabolism, and immune cells. Targeting the aging adipose microenvironment, particularly lipid metabolism reprogramming, holds promise as a therapeutic strategy for OC, which warrants further investigation.","PeriodicalId":501233,"journal":{"name":"bioRxiv - Cancer Biology","volume":"409 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142210606","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-11DOI: 10.1101/2024.09.06.611647
Jose Teles-Reis, Ashish Jain, Dan Liu, Rojyar Khezri, Sofia Michelli, Alicia Alfonso Gomez, Caroline DIllard, Tor E Rusten
Cell biology and genetic analysis of intracellular, intercellular and inter-organ interaction studies in animal models are key for understanding development, physiology, and disease. The MARCM technique can emulate tumor development by simultaneous clonal tumor suppressor loss-of-function generation coupled with GAL4-UAS-driven oncogene and marker expression, but the utility is limited for studying tumor-host interactions due to genetic constraints. To overcome this, we introduce EyaHOST, a novel system that replaces MARCM with the QF2-QUAS binary gene expression system under the eya promoter control, unleashing the fly community genome-wide GAL4-UAS driven tools to manipulate any host cells or tissue at scale. EyaHOST generates epithelial clones in the eye epithelium similar to MARCM. EyaHOST-driven RasV12 oncogene overexpression coupled with scribble tumor suppressor knockdown recapitulates key cancer features, including systemic catabolic switching and organ wasting. We demonstrate effective tissue-specific manipulation of host compartments such as neighbouring epithelial cells, immune cells, fat body, and muscle using fly avatars with tissue-specific GAL4 drivers. Organ-specific inhibition of autophagy or stimulation of growth-signaling through PTEN knockdown in fat body or muscle prevents cachexia-like wasting. Additionally, we show that RasV12, scribRNAi tumors induce caspase-driven apoptosis in the epithelial microenvironment. Inhibition of apoptosis by p35 expression in the microenvironment promotes tumor growth. EyaHOST offers a versatile modular platform for dissecting tumor-host interactions and other mechanisms involving intercellular and inter-organ communication in Drosophila.
{"title":"EyaHOST a modular genetic system for investigation of intercellular and tumor-host interactions in Drosophila melanogaster","authors":"Jose Teles-Reis, Ashish Jain, Dan Liu, Rojyar Khezri, Sofia Michelli, Alicia Alfonso Gomez, Caroline DIllard, Tor E Rusten","doi":"10.1101/2024.09.06.611647","DOIUrl":"https://doi.org/10.1101/2024.09.06.611647","url":null,"abstract":"Cell biology and genetic analysis of intracellular, intercellular and inter-organ interaction studies in animal models are key for understanding development, physiology, and disease. The MARCM technique can emulate tumor development by simultaneous clonal tumor suppressor loss-of-function generation coupled with GAL4-UAS-driven oncogene and marker expression, but the utility is limited for studying tumor-host interactions due to genetic constraints. To overcome this, we introduce EyaHOST, a novel system that replaces MARCM with the QF2-QUAS binary gene expression system under the eya promoter control, unleashing the fly community genome-wide GAL4-UAS driven tools to manipulate any host cells or tissue at scale. EyaHOST generates epithelial clones in the eye epithelium similar to MARCM. EyaHOST-driven RasV12 oncogene overexpression coupled with scribble tumor suppressor knockdown recapitulates key cancer features, including systemic catabolic switching and organ wasting. We demonstrate effective tissue-specific manipulation of host compartments such as neighbouring epithelial cells, immune cells, fat body, and muscle using fly avatars with tissue-specific GAL4 drivers. Organ-specific inhibition of autophagy or stimulation of growth-signaling through PTEN knockdown in fat body or muscle prevents cachexia-like wasting. Additionally, we show that RasV12, scribRNAi tumors induce caspase-driven apoptosis in the epithelial microenvironment. Inhibition of apoptosis by p35 expression in the microenvironment promotes tumor growth. EyaHOST offers a versatile modular platform for dissecting tumor-host interactions and other mechanisms involving intercellular and inter-organ communication in Drosophila.","PeriodicalId":501233,"journal":{"name":"bioRxiv - Cancer Biology","volume":"77 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142210446","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-11DOI: 10.1101/2024.09.09.612140
Robert J Vanner, Suraj Bansal, Marco M Buttigeig, Andy G.X. Zeng, Vincent Rondeau, Darryl Y. Chan, Michelle Chan-Seng-Yue, Liqing Jin, Jessica McLeod, Elisa Donato, Patrick Stelmach, Caitlyn Vlasschaert, Yitong Yang, Aarushi Gupta, Sofia Genta, Enrique Sanz Garcia, Liran Shlush, Mauricio Ribeiro, Marcus O Butler, Sagi Abelson, Mark Minden, Steven M Chan, Michael J Rauh, Andreas Trumpp, John E Dick
Somatic mutations inactivating TET2 are among the most common drivers of clonal hematopoiesis (CH). While TET2 inactivation is associated with monocyte-derived inflammation and improved chimeric antigen-receptor-T cell function, its impact on immunotherapy response is unknown. In our mouse model, hematopoietic Tet2 mutation enhanced immune checkpoint blockade (ICB) response. Enhanced ICB response with Tet2 mutation required phagocytes, CD4 and CD8 T cells. Mechanistically, in Tet2-mutant tumor-infiltrating leukocytes (TILs), ICB preferentially induced anti-tumor states and restricted cell states linked to tumor progression. Tet2-mutant monocytes activated costimulatory programs, while Tet2-mutant T cells showed enhanced T cell memory signatures, lesser exhaustion and decreased regulatory phenotype. Our murine data was clinically relevant, since we found that melanomas from patients with TET2 driver mutation-CH (TET2-CH) showed enhanced immune infiltration, T cell activation, and T cell memory programs. In melanoma patients treated with ICB, TET2-CH was associated with 6-fold greater odds of clinical benefit. Collectively, our data establishes that hematopoietic Tet2 inactivation primes leukocytes for anti-tumor states associated with immunotherapy response and provides a potential biomarker for personalized therapy.
{"title":"Hematopoietic Tet2 inactivation enhances the response to checkpoint blockade immunotherapy","authors":"Robert J Vanner, Suraj Bansal, Marco M Buttigeig, Andy G.X. Zeng, Vincent Rondeau, Darryl Y. Chan, Michelle Chan-Seng-Yue, Liqing Jin, Jessica McLeod, Elisa Donato, Patrick Stelmach, Caitlyn Vlasschaert, Yitong Yang, Aarushi Gupta, Sofia Genta, Enrique Sanz Garcia, Liran Shlush, Mauricio Ribeiro, Marcus O Butler, Sagi Abelson, Mark Minden, Steven M Chan, Michael J Rauh, Andreas Trumpp, John E Dick","doi":"10.1101/2024.09.09.612140","DOIUrl":"https://doi.org/10.1101/2024.09.09.612140","url":null,"abstract":"Somatic mutations inactivating TET2 are among the most common drivers of clonal hematopoiesis (CH). While TET2 inactivation is associated with monocyte-derived inflammation and improved chimeric antigen-receptor-T cell function, its impact on immunotherapy response is unknown. In our mouse model, hematopoietic Tet2 mutation enhanced immune checkpoint blockade (ICB) response. Enhanced ICB response with Tet2 mutation required phagocytes, CD4 and CD8 T cells. Mechanistically, in Tet2-mutant tumor-infiltrating leukocytes (TILs), ICB preferentially induced anti-tumor states and restricted cell states linked to tumor progression. Tet2-mutant monocytes activated costimulatory programs, while Tet2-mutant T cells showed enhanced T cell memory signatures, lesser exhaustion and decreased regulatory phenotype. Our murine data was clinically relevant, since we found that melanomas from patients with TET2 driver mutation-CH (TET2-CH) showed enhanced immune infiltration, T cell activation, and T cell memory programs. In melanoma patients treated with ICB, TET2-CH was associated with 6-fold greater odds of clinical benefit. Collectively, our data establishes that hematopoietic Tet2 inactivation primes leukocytes for anti-tumor states associated with immunotherapy response and provides a potential biomarker for personalized therapy.","PeriodicalId":501233,"journal":{"name":"bioRxiv - Cancer Biology","volume":"50 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142210604","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Cholangiocarcinoma (CCA) is a diverse collection of malignant tumors that originate in the bile ducts. Mitochondria, the energy converters in eukaryotic cells, contain circular mitochondrial DNA (mtDNA) which has a greater mutation rate than nuclear DNA. Heteroplasmic variations in mtDNA may suggest an increased risk of cancer-related mortality, serving as a potential prognostic marker. In this study, we investigated the mtDNA variations of five CCA cell lines, including KKU-023, KKU-055, KKU-100, KKU213A, and KKU-452 and compared them to the non-tumor cholangiocyte MMNK-1 cell line. We used Oxford Nanopore Technologies (ONT), a long-read sequencing technology capable of synthesizing the whole mitochondrial genome, which facilitates enhanced identification of complicated rearrangements in mitogenomics. The analysis revealed a high frequency of SNVs and INDELs, particularly in the D-loop, MT-RNR2, MT-CO1, MT-ND4, and MT-ND5 genes. Significant mutations were detected in all CCA cell lines, with particularly notable non-synonymous SNVs such as m.8462T C in KKU-023, m.9493G A in KKU-055, m.9172C A in KKU-100, m.15024GC in KKU-213A, m.12994G A in KKU-452, and m.13406G A in MMNK-1, which demonstrated high pathogenicity scores. The presence of these mutations suggests the potential for mitochondrial dysfunction and CCA progression. Analysis of mtDNA structural variants (SV) revealed significant variability among the cell lines. We identified 208 SVs in KKU-023, 185 SVs in KKU-055, 231 SVs in KKU-100, 69 SVs in KKU-213A, 172 SVs in KKU-452, and 217 SVs in MMNK-1. These SVs included deletions, duplications, and inversions, with the highest variability observed in KKU-100 and the lowest in KKU-213A. Our results underscore the diverse mtDNA mutation landscape in CCA cell lines, highlighting the potential impact of these mutations on mitochondrial function and CCA cell line progression. Future research is required to investigate the functional impacts of these variants, their interactions with nuclear DNA in CCA, and their potential as targets for therapeutic intervention.
{"title":"Mitochondrial genomic alterations in cholangiocarcinoma cell lines","authors":"Athitaya Faipan, Sirinya Sitthirak, Arporn Wangwiwatsin, Nisana Namwat, Poramate Klanrit, Attapol Titapan, Apiwat Jareanrat, Vasin Thanasukarn, Natcha Khuntikeo, Luke Boulter, Watcharin Loilome","doi":"10.1101/2024.09.09.611963","DOIUrl":"https://doi.org/10.1101/2024.09.09.611963","url":null,"abstract":"Cholangiocarcinoma (CCA) is a diverse collection of malignant tumors that originate in the bile ducts. Mitochondria, the energy converters in eukaryotic cells, contain circular mitochondrial DNA (mtDNA) which has a greater mutation rate than nuclear DNA. Heteroplasmic variations in mtDNA may suggest an increased risk of cancer-related mortality, serving as a potential prognostic marker. In this study, we investigated the mtDNA variations of five CCA cell lines, including KKU-023, KKU-055, KKU-100, KKU213A, and KKU-452 and compared them to the non-tumor cholangiocyte MMNK-1 cell line. We used Oxford Nanopore Technologies (ONT), a long-read sequencing technology capable of synthesizing the whole mitochondrial genome, which facilitates enhanced identification of complicated rearrangements in mitogenomics. The analysis revealed a high frequency of SNVs and INDELs, particularly in the D-loop, MT-RNR2, MT-CO1, MT-ND4, and MT-ND5 genes. Significant mutations were detected in all CCA cell lines, with particularly notable non-synonymous SNVs such as m.8462T C in KKU-023, m.9493G A in KKU-055, m.9172C A in KKU-100, m.15024GC in KKU-213A, m.12994G A in KKU-452, and m.13406G A in MMNK-1, which demonstrated high pathogenicity scores. The presence of these mutations suggests the potential for mitochondrial dysfunction and CCA progression. Analysis of mtDNA structural variants (SV) revealed significant variability among the cell lines. We identified 208 SVs in KKU-023, 185 SVs in KKU-055, 231 SVs in KKU-100, 69 SVs in KKU-213A, 172 SVs in KKU-452, and 217 SVs in MMNK-1. These SVs included deletions, duplications, and inversions, with the highest variability observed in KKU-100 and the lowest in KKU-213A. Our results underscore the diverse mtDNA mutation landscape in CCA cell lines, highlighting the potential impact of these mutations on mitochondrial function and CCA cell line progression. Future research is required to investigate the functional impacts of these variants, their interactions with nuclear DNA in CCA, and their potential as targets for therapeutic intervention.","PeriodicalId":501233,"journal":{"name":"bioRxiv - Cancer Biology","volume":"4 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142210447","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-11DOI: 10.1101/2024.09.06.611663
Keegan A Christensen, MELISSA A Fath, Jordan T Ewald, Claudia Robles-Planells, Stephen A Graves, Spenser S Johnson, Zeb R Zacharias, Jon C. D. Houtman, M. Sue O'Dorisio, Michael K Schultz, Bryan G. Allen, Muhammad Furqan, Yusuf Menda, Dijie Liu, Douglas R Spitz
Introduction: Small cell lung cancer (SCLC) has a 7% 5-year overall survival. C-X-C chemokine receptor 4 (CXCR4), an attractive target for theranostic agents, is highly expressed in SCLCs, and can be targeted with pentixather using the theranostic pair 212Pb/203Pb. The hypothesis that [212Pb/203Pb]-pentixather can be used safely and effectively for imaging and therapy in SCLC in xenograft models was tested. Results: SPECT/CT imaging and biodistribution studies of tumor bearing mice injected with [203Pb]-pentixather demonstrated CXCR4-dependent uptake in tumors and accumulation of radioligand in kidneys and livers. Dosimetry calculations estimated [212Pb]-pentixather uptake in tumor and normal tissue. [212Pb]-pentixather treatment (37-111 kBq/g) of SCLC xenografts (DMS273 and H69AR) significantly prolonged survival and delayed tumor growth. When NSG mice grafted with human hCD34+ bone marrow were treated with [212Pb]-pentixather (37-111 kBq/g), significant cytopenias were observed in peripheral blood complete blood counts (CBCs) at 13-18 days post treatment which resolved by day 28-31. Flow cytometry of bone marrow hematopeotic stem cells in these animals at day 28-31 demonstrated a significantly reduced frequency of the human hematopoietic marker CD45 (hCD45+) and reconstitution of the bone marrow with murine CD45+ (mCD45+) lineages. Conclusions: [203Pb]-pentixather can be used to image CXCR4 expressing SCLC xenografts and treatment with alpha emitter [212Pb]-pentixather significantly prolongs SCLC xenograft median overall survival. Significantly greater mCD45+ bone marrow repopulation was detected in NSG mice engrafted with human bone marrow 28-31 days following [212Pb]-pentixather treatment, relative to hCD45+ bone marrow.
{"title":"Targeting CXCR4 with [212Pb/203Pb]-Pentixather Significantly Increases Overall Survival in Small Cell Lung Cancer","authors":"Keegan A Christensen, MELISSA A Fath, Jordan T Ewald, Claudia Robles-Planells, Stephen A Graves, Spenser S Johnson, Zeb R Zacharias, Jon C. D. Houtman, M. Sue O'Dorisio, Michael K Schultz, Bryan G. Allen, Muhammad Furqan, Yusuf Menda, Dijie Liu, Douglas R Spitz","doi":"10.1101/2024.09.06.611663","DOIUrl":"https://doi.org/10.1101/2024.09.06.611663","url":null,"abstract":"Introduction: Small cell lung cancer (SCLC) has a 7% 5-year overall survival. C-X-C chemokine receptor 4 (CXCR4), an attractive target for theranostic agents, is highly expressed in SCLCs, and can be targeted with pentixather using the theranostic pair <sup>212</sup>Pb/<sup>203</sup>Pb. The hypothesis that [<sup>212</sup>Pb/<sup>203</sup>Pb]-pentixather can be used safely and effectively for imaging and therapy in SCLC in xenograft models was tested. Results: SPECT/CT imaging and biodistribution studies of tumor bearing mice injected with [<sup>203</sup>Pb]-pentixather demonstrated CXCR4-dependent uptake in tumors and accumulation of radioligand in kidneys and livers. Dosimetry calculations estimated [<sup>212</sup>Pb]-pentixather uptake in tumor and normal tissue. [<sup>212</sup>Pb]-pentixather treatment (37-111 kBq/g) of SCLC xenografts (DMS273 and H69AR) significantly prolonged survival and delayed tumor growth. When NSG mice grafted with human hCD34+ bone marrow were treated with [<sup>212</sup>Pb]-pentixather (37-111 kBq/g), significant cytopenias were observed in peripheral blood complete blood counts (CBCs) at 13-18 days post treatment which resolved by day 28-31. Flow cytometry of bone marrow hematopeotic stem cells in these animals at day 28-31 demonstrated a significantly reduced frequency of the human hematopoietic marker CD45 (hCD45+) and reconstitution of the bone marrow with murine CD45+ (mCD45+) lineages. Conclusions: [<sup>203</sup>Pb]-pentixather can be used to image CXCR4 expressing SCLC xenografts and treatment with alpha emitter [<sup>212</sup>Pb]-pentixather significantly prolongs SCLC xenograft median overall survival. Significantly greater mCD45+ bone marrow repopulation was detected in NSG mice engrafted with human bone marrow 28-31 days following [<sup>212</sup>Pb]-pentixather treatment, relative to hCD45+ bone marrow.","PeriodicalId":501233,"journal":{"name":"bioRxiv - Cancer Biology","volume":"177 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142210445","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-11DOI: 10.1101/2024.09.10.612311
Mengxi Zhou, Tito Fojo, Lawrence Schwartz, Susan E Bates, Krastan B. Blagoev
Background: We applied 11 mathematical models of tumor growth to clinical trial data available from public and private sources. We have previously described the remarkable capacity for a simple biexponential model of tumor growth to fit thousands of datasets, and to correlate with overall survival. The goal of this study was to extend our analysis to additional tumor types and to evaluate whether alternate growth models could describe the time course of disease burden in the small subset of patients in whom the biexponential model failed. Methods: For this analysis, we obtained data for tumor burden from 17,140 patients with six different tumor types. Imaging data and serum levels of tumor markers were available for 3,346 and 13,794 patients, respectively. Data from patients were first analyzed using the biexponential model to determine rates of tumor growth (g) and regression (d); for those whose data could not be described by this model, fit of their data was assessed using seven alternative models. The model that minimized the Akaike Information Criterion was selected as the best fit. Using the model that best fit an individual patient's data, we estimated the rates of growth (g) and regression (d) of disease burden over time. The rates of tumor growth (g) were examined for association with a traditional endpoint (overall survival). Findings: For each model, the number of patient datasets that fit the model were obtained. As we have previously reported, data from most patients fit a simple model of exponential growth and exponential regression (86%). Data from another 7% of patients fit an alternative model, including 3% fitting to a model of constant or linear regression and exponential growth of tumor on the surface and 3% fitting to model of exponential decay on tumor surface with asymmetric growth. As previously reported, we found that growth rate correlates well with overall survival, remarkably even when data from various histologies are considered together. For patients with multiple timepoints of tumor measurement, the growth rate could often be estimated even during the phase when only net regression of tumor quantity could be discerned. Interpretation: The validation of a simple mathematical model across different cancers and its application to existing clinical data allowed estimation of the rate of growth of a treatment resistant subpopulation of cancer cells. The quantification of available clinical data using the growth rate of tumors in individual patients and its strong correlation with overall survival makes the growth rate an excellent marker of the efficacy of therapy specific to the individual patient.
{"title":"Cancer Interception During Treatment: Using Growth Kinetics to Create a Continuous Variable for Assessing Disease Response","authors":"Mengxi Zhou, Tito Fojo, Lawrence Schwartz, Susan E Bates, Krastan B. Blagoev","doi":"10.1101/2024.09.10.612311","DOIUrl":"https://doi.org/10.1101/2024.09.10.612311","url":null,"abstract":"Background: We applied 11 mathematical models of tumor growth to clinical trial data available from public and private sources. We have previously described the remarkable capacity for a simple biexponential model of tumor growth to fit thousands of datasets, and to correlate with overall survival. The goal of this study was to extend our analysis to additional tumor types and to evaluate whether alternate growth models could describe the time course of disease burden in the small subset of patients in whom the biexponential model failed.\u0000Methods: For this analysis, we obtained data for tumor burden from 17,140 patients with six different tumor types. Imaging data and serum levels of tumor markers were available for 3,346 and 13,794 patients, respectively. Data from patients were first analyzed using the biexponential model to determine rates of tumor growth (g) and regression (d); for those whose data could not be described by this model, fit of their data was assessed using seven alternative models. The model that minimized the Akaike Information Criterion was selected as the best fit. Using the model that best fit an individual patient's data, we estimated the rates of growth (g) and regression (d) of disease burden over time. The rates of tumor growth (g) were examined for association with a traditional endpoint (overall survival).\u0000Findings: For each model, the number of patient datasets that fit the model were obtained. As we have previously reported, data from most patients fit a simple model of exponential growth and exponential regression (86%). Data from another 7% of patients fit an alternative model, including 3% fitting to a model of constant or linear regression and exponential growth of tumor on the surface and 3% fitting to model of exponential decay on tumor surface with asymmetric growth. As previously reported, we found that growth rate correlates well with overall survival, remarkably even when data from various histologies are considered together. For patients with multiple timepoints of tumor measurement, the growth rate could often be estimated even during the phase when only net regression of tumor quantity could be discerned.\u0000Interpretation: The validation of a simple mathematical model across different cancers and its application to existing clinical data allowed estimation of the rate of growth of a treatment resistant subpopulation of cancer cells. The quantification of available clinical data using the growth rate of tumors in individual patients and its strong correlation with overall survival makes the growth rate an excellent marker of the efficacy of therapy specific to the individual patient.","PeriodicalId":501233,"journal":{"name":"bioRxiv - Cancer Biology","volume":"27 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142210448","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The Rad18 E3 ubiquitin ligase is a key regulator of DNA damage tolerance that also functions in repair of DNA double strand breaks. Rad18 is overexpressed in the aggressive brain cancer glioblastoma and its downregulation sensitizes glioblastoma cells to DNA damaging agents. Here we show that Rad18 has an essential role in glioblastoma cells proliferation in the absence of external damage, which is independent of its catalytic activity. Rad18 downregulation leads to cell cycle arrest in the G1 phase of the cell cycle in the absence of apparent DNA damage. We also show that Rad18 is essential for glioblastoma stem cells self-renewal and survival, and the growth of tumor orthotropic xenografts in mice. We also show that increased Rad18 expression enhances the growth of transformed cells and induces features of oncogenic transformation. Altogether, these data propose Rad18, a non-essential gene, as a gene essential for GBM proliferation and a key target to sensitize glioblastoma to therapy.
Rad18 E3 泛素连接酶是 DNA 损伤耐受性的一个关键调节因子，它还具有修复 DNA 双股断裂的功能。Rad18 在侵袭性脑癌胶质母细胞瘤中过度表达，其下调会使胶质母细胞瘤细胞对 DNA 损伤剂敏感。我们在这里发现，在没有外部损伤的情况下，Rad18 对胶质母细胞瘤细胞的增殖起着至关重要的作用，这与它的催化活性无关。在没有明显 DNA 损伤的情况下，Rad18 的下调会导致细胞周期停滞在细胞周期的 G1 阶段。我们还发现，Rad18 对胶质母细胞瘤干细胞的自我更新和存活以及肿瘤在小鼠体内的正交异种移植的生长至关重要。我们还发现，Rad18 表达的增加会促进转化细胞的生长，并诱导致癌转化的特征。总之，这些数据表明 Rad18 这一非基本基因是胶质母细胞瘤增殖所必需的基因，也是使胶质母细胞瘤对治疗敏感的关键靶点。
{"title":"A non-catalytic function for Rad18 in sustaining glioblastoma proliferation","authors":"Nour El-Houda Benbahouche, Chames Kermi, Aurore Siegfried, Lenka Sefancikova, Jean-Marc Pascussi, Julie Pannequin, Jerome Moreaux, Jean-Philippe Hugnot, Marie-Bernadette Delisle, Elizabeth Moyal, Emmanuelle Uro-Coste, Domenico Maiorano","doi":"10.1101/2024.09.05.611406","DOIUrl":"https://doi.org/10.1101/2024.09.05.611406","url":null,"abstract":"The Rad18 E3 ubiquitin ligase is a key regulator of DNA damage tolerance that also functions in repair of DNA double strand breaks. Rad18 is overexpressed in the aggressive brain cancer glioblastoma and its downregulation sensitizes glioblastoma cells to DNA damaging agents. Here we show that Rad18 has an essential role in glioblastoma cells proliferation in the absence of external damage, which is independent of its catalytic activity. Rad18 downregulation leads to cell cycle arrest in the G1 phase of the cell cycle in the absence of apparent DNA damage. We also show that Rad18 is essential for glioblastoma stem cells self-renewal and survival, and the growth of tumor orthotropic xenografts in mice. We also show that increased Rad18 expression enhances the growth of transformed cells and induces features of oncogenic transformation. Altogether, these data propose Rad18, a non-essential gene, as a gene essential for GBM proliferation and a key target to sensitize glioblastoma to therapy.","PeriodicalId":501233,"journal":{"name":"bioRxiv - Cancer Biology","volume":"28 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142210466","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-10DOI: 10.1101/2024.09.08.611898
Isabel Mogollon, Michaela Feodoroff, Pedro Neto, Alba Montedeoca, Vilja Pietiainen, Lassi Paavolainen
Understanding cellular function within 3D multicellular spheroids is essential for advancing cancer research, particularly in studying cell-stromal interactions as potential targets for novel drug therapies. However, accurate single-cell segmentation in 3D cultures is challenging due to dense cell clustering and the impracticality of manual annotations. We present a high-throughput (HT) 3D single-cell analysis pipeline that combines optimized wet-lab conditions, deep learning (DL)-based segmentation models, and Bayesian optimization to address these challenges. By using live-cell nuclear and cytoplasmic dyes, we achieved clear and uniform staining of cell populations in renal cancer and immune T-cell monocultures and cocultures, improving single-cell detection in spheroids. Our pipeline integrates image preprocessing and DL models based on 3DUnet architecture, enabling robust segmentation of densely packed 3D structures. Bayesian optimization, guided by a custom objective function, was employed to refine segmentation parameters and improve quality based on biologically relevant criteria. The pipeline successfully segments cells under various drug treatments, revealing drug-induced changes not detectable by bulk conventional assays. This approach has potential for application to more complex biological samples, including, organoid co-cultures, diverse drug treatments, and integration with additional immunostaining assays, paving the way for detailed HT analyses of single-cell responses.
了解三维多细胞球体内的细胞功能对推进癌症研究至关重要，尤其是在研究细胞间相互作用作为新型药物疗法的潜在靶点方面。然而，在三维培养物中进行精确的单细胞分割具有挑战性，因为细胞密集成群，而且人工注释不切实际。我们介绍了一种高通量（HT）三维单细胞分析流水线，它结合了优化的湿实验室条件、基于深度学习（DL）的分割模型和贝叶斯优化来应对这些挑战。通过使用活细胞核和细胞质染料，我们实现了对肾癌和免疫 T 细胞单培养物和共培养物中细胞群的清晰而均匀的染色，提高了球形细胞中的单细胞检测能力。我们的管道集成了图像预处理和基于 3DUnet 架构的 DL 模型，能够对密集的三维结构进行稳健的分割。在自定义目标函数的指导下，我们采用贝叶斯优化法来完善分割参数，并根据生物相关标准提高质量。该管道成功地分割了各种药物治疗下的细胞，揭示了大量传统检测方法无法检测到的药物诱导变化。这种方法有望应用于更复杂的生物样本，包括类器官共培养、不同的药物处理，以及与其他免疫染色检测的整合，为单细胞反应的详细 HT 分析铺平道路。
{"title":"Achieving high-resolution single-cell segmentation in convoluted cancer spheroids via Bayesian optimization and deep-learning","authors":"Isabel Mogollon, Michaela Feodoroff, Pedro Neto, Alba Montedeoca, Vilja Pietiainen, Lassi Paavolainen","doi":"10.1101/2024.09.08.611898","DOIUrl":"https://doi.org/10.1101/2024.09.08.611898","url":null,"abstract":"Understanding cellular function within 3D multicellular spheroids is essential for advancing cancer research, particularly in studying cell-stromal interactions as potential targets for novel drug therapies. However, accurate single-cell segmentation in 3D cultures is challenging due to dense cell clustering and the impracticality of manual annotations. We present a high-throughput (HT) 3D single-cell analysis pipeline that combines optimized wet-lab conditions, deep learning (DL)-based segmentation models, and Bayesian optimization to address these challenges. By using live-cell nuclear and cytoplasmic dyes, we achieved clear and uniform staining of cell populations in renal cancer and immune T-cell monocultures and cocultures, improving single-cell detection in spheroids. Our pipeline integrates image preprocessing and DL models based on 3DUnet architecture, enabling robust segmentation of densely packed 3D structures. Bayesian optimization, guided by a custom objective function, was employed to refine segmentation parameters and improve quality based on biologically relevant criteria. The pipeline successfully segments cells under various drug treatments, revealing drug-induced changes not detectable by bulk conventional assays. This approach has potential for application to more complex biological samples, including, organoid co-cultures, diverse drug treatments, and integration with additional immunostaining assays, paving the way for detailed HT analyses of single-cell responses.","PeriodicalId":501233,"journal":{"name":"bioRxiv - Cancer Biology","volume":"11 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142210465","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-10DOI: 10.1101/2024.09.09.611297
Anil Kumar Kalvala, Ashok Silwal, Bhaumik Patel, Apoorva Kasetti, Kirti Shetty, Jung-Hung Cho, Gerard Lara, Beth Daugherity, Remi Diesler, Venkatesh Pooladanda, Bo R Rueda, Elizabeth P. Henske, Jane J Yu, Maciej M Markiewski, Magdalena Karbowniczek
Pulmonary lymphangioleiomyomatosis (LAM) is metastatic sarcoma but mechanisms regulating LAM metastasis are unknown. Extracellular vesicle (EV) regulate cancer metastasis but their roles in LAM have not yet been investigated. Here, we report that EV biogenesis is increased in LAM and LAM EV cargo is enriched with lung tropic integrins, metalloproteinases, and cancer stem cell markers. LAM-EV increase LAM cell migration and invasion via the ITGα6/β1-c-Src-FAK-AKT axis. Metastable (hybrid) phenotypes of LAM metastasizing cells, pivotal for metastasis, are regulated by EV from primary tumor or metastasizing LAM cells via shuttling ATP synthesis to cell pseudopodia or activation of integrin adhesion complex, respectively. In mouse models of LAM, LAM-EV increase lung metastatic burden through mechanisms involving lung extracellular matrix remodeling. Collectively, these data provide evidence for the role of EV in promoting LAM lung metastasis and identify novel EV-dependent mechanisms regulating metastable phenotypes of tumor cells. Clinical impact of research is that it establishes LAM-EV as novel target for LAM therapy.
{"title":"Extracellular vesicles regulate metastable phenotypes of lymphangioleiomyomatosis cells via shuttling ATP synthesis to pseudopodia and activation of integrin adhesion complexes.","authors":"Anil Kumar Kalvala, Ashok Silwal, Bhaumik Patel, Apoorva Kasetti, Kirti Shetty, Jung-Hung Cho, Gerard Lara, Beth Daugherity, Remi Diesler, Venkatesh Pooladanda, Bo R Rueda, Elizabeth P. Henske, Jane J Yu, Maciej M Markiewski, Magdalena Karbowniczek","doi":"10.1101/2024.09.09.611297","DOIUrl":"https://doi.org/10.1101/2024.09.09.611297","url":null,"abstract":"Pulmonary lymphangioleiomyomatosis (LAM) is metastatic sarcoma but mechanisms regulating LAM metastasis are unknown. Extracellular vesicle (EV) regulate cancer metastasis but their roles in LAM have not yet been investigated. Here, we report that EV biogenesis is increased in LAM and LAM EV cargo is enriched with lung tropic integrins, metalloproteinases, and cancer stem cell markers. LAM-EV increase LAM cell migration and invasion via the ITGα6/β1-c-Src-FAK-AKT axis. Metastable (hybrid) phenotypes of LAM metastasizing cells, pivotal for metastasis, are regulated by EV from primary tumor or metastasizing LAM cells via shuttling ATP synthesis to cell pseudopodia or activation of integrin adhesion complex, respectively. In mouse models of LAM, LAM-EV increase lung metastatic burden through mechanisms involving lung extracellular matrix remodeling. Collectively, these data provide evidence for the role of EV in promoting LAM lung metastasis and identify novel EV-dependent mechanisms regulating metastable phenotypes of tumor cells. Clinical impact of research is that it establishes LAM-EV as novel target for LAM therapy.","PeriodicalId":501233,"journal":{"name":"bioRxiv - Cancer Biology","volume":"12 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142210450","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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