Pub Date : 2024-06-10DOI: 10.1158/1538-8514.synthleth24-a012
Haijiao Zhang, Basil Allen, D. Bottomly, P. Ryabinin, Schannon K Mcweeney
Monosomy 7 is among the most frequent cytogenetic abnormalities in hematological disorders and is associated with short survival and drug resistance. Despite its high prevalence and detrimental impact, the therapeutic vulnerabilities underlying monosomy 7-associated blood disorders remain largely elusive, impeding progress toward improved patient care. The homeostatic cellular requirement for a normal dosage of essential genes creates an opportunity to target vulnerabilities that arise due to reduced levels of proteins encoded by a haploinsufficient essential gene. Briefly, the loss of one copy of a dosage-sensitive essential gene (gene X), in combination with the inhibition of itself or a related gene (gene Y), or an associated pathway results in lethal consequences for cells. The remarkable and selective clinical efficacy of lenalidomide for the treatment of del(5q) MDS has demonstrated how allelic haploinsufficiency underlies the sensitivity to this drug by synthetic lethality. Differential expression analysis of gene and protein expression in primary AML samples with monosomy 7 revealed significant downregulation of multiple proteasome pathway members at the protein level, but not at the RNA level. Primary AML samples with -7/del(7q) exhibited increased sensitivity (low IC50) to the proteasome inhibitor bortezomib, as evidenced by two independent ex vivo drug screening cohorts (the Beat AML and the FIMM study). Chromosome 7 harbors four proteasome subunits, PSMA2, PSMC2, PSMG3, and SEM1. We performed gene expression, protein expression, copy number analysis, and individual gene knockout experiments. The results have revealed PSMA2 to be a haploinsufficient essential gene on chromosome 7. PSMA2 knockout confers leukemia a growth disadvantage for multiple AML cell clines in both TP53 wild-type and knockout backgrounds. We generated PSMA2 isogenic hemizygous deletion and diploid single-cell clones. PSMA2 hemizygous deletion cells exhibited approximately half the protein expression compared to diploid controls, confirming that PSMA2 is a haploinsufficient gene. PSMA2hemizygous deletion single-cell clones showed significantly enhanced sensitivity to all three evaluated proteasome inhibitors (bortezomib, ixazomib, and carfilzomib), aligning with the sensitivity observed in primary -7/del(7q) leukemia samples. PSMA2 hemizygous deletion cell clones displayed increased p38 and decreased pERK levels upon treatment with proteasome inhibitors, potentially contributing to their increased sensitivity to proteasome inhibitors. Proteomics analysis and in vivo validation is ongoing. As such, we have identified haploinsufficient essential gene PSMA2 mediated proteasome pathway vulnerability in monosomy 7 associated leukemia and further showed that proteasome inhibitors as promising therapeutic approaches for treating hematological disorders associated with monosomy 7. Citation Format: Haijiao Zhang, Basil Allen, Daniel Bottomly, Peter Ryabinin, Schannon K.
单体 7 是血液病中最常见的细胞遗传学异常之一,与存活期短和耐药性有关。尽管单体 7 基因异常发病率高、影响恶劣,但与单体 7 基因异常相关的血液疾病的治疗漏洞在很大程度上仍然难以捉摸,阻碍了改善患者护理的进程。细胞对必需基因正常剂量的平衡性要求,为针对单倍体必需基因编码的蛋白质水平降低而产生的薄弱环节提供了机会。简而言之,剂量敏感的重要基因(X 基因)的一个拷贝缺失,再加上自身或相关基因(Y 基因)或相关途径受到抑制,就会给细胞带来致命的后果。来那度胺在治疗 del(5q) MDS 方面卓越的选择性临床疗效证明了等位基因单倍体缺乏症是如何通过合成致死性对这种药物产生敏感性的。对单体7型原发性急性髓细胞样本中基因和蛋白质表达的差异表达分析表明,蛋白酶体通路的多个成员在蛋白质水平上显著下调,但在RNA水平上没有下调。两个独立的体外药物筛选队列(Beat AML 和 FIMM 研究)证明,-7/del(7q)的原发性急性髓细胞样本对蛋白酶体抑制剂硼替佐米的敏感性增加(IC50 低)。染色体 7 上有四个蛋白酶体亚基:PSMA2、PSMC2、PSMG3 和 SEM1。我们进行了基因表达、蛋白表达、拷贝数分析和单个基因敲除实验。结果表明,PSMA2是7号染色体上的一个单倍体必需基因。在 TP53 野生型和基因敲除背景下,PSMA2 基因敲除会使多个急性髓性白血病细胞系处于生长劣势。我们生成了 PSMA2 同源基因半杂合子缺失和二倍体单细胞克隆。与二倍体对照组相比,PSMA2半杂合子缺失细胞的蛋白表达量约为对照组的一半,这证实了PSMA2是一个单倍体基因。PSMA2半杂合子缺失单细胞克隆对所评估的三种蛋白酶体抑制剂(硼替佐米、伊沙佐米和卡非佐米)的敏感性都明显增强,这与在原发性-7/del(7q)白血病样本中观察到的敏感性一致。PSMA2半杂合子缺失细胞克隆在接受蛋白酶体抑制剂治疗后,p38水平升高,pERK水平降低,这可能是它们对蛋白酶体抑制剂更敏感的原因。蛋白质组学分析和体内验证正在进行中。因此,我们发现了单倍体必需基因PSMA2介导的蛋白酶体通路在7号单体相关白血病中的脆弱性,并进一步表明蛋白酶体抑制剂是治疗7号单体相关血液病的有前景的治疗方法:Haijiao Zhang, Basil Allen, Daniel Bottomly, Peter Ryabinin, Schannon K. Targeting proteasome vulnerabilities for the treatment of monosomy 7 associated blood disorders [abstract].In:AACR 癌症研究特别会议论文集:扩展和转化癌症合成漏洞;2024 年 6 月 10-13 日;加拿大魁北克省蒙特利尔。费城(宾夕法尼亚州):AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr A012.
{"title":"Abstract A012: Targeting proteasome vulnerabilities for the treatment of monosomy 7 associated blood disorders","authors":"Haijiao Zhang, Basil Allen, D. Bottomly, P. Ryabinin, Schannon K Mcweeney","doi":"10.1158/1538-8514.synthleth24-a012","DOIUrl":"https://doi.org/10.1158/1538-8514.synthleth24-a012","url":null,"abstract":"\u0000 Monosomy 7 is among the most frequent cytogenetic abnormalities in hematological disorders and is associated with short survival and drug resistance. Despite its high prevalence and detrimental impact, the therapeutic vulnerabilities underlying monosomy 7-associated blood disorders remain largely elusive, impeding progress toward improved patient care. The homeostatic cellular requirement for a normal dosage of essential genes creates an opportunity to target vulnerabilities that arise due to reduced levels of proteins encoded by a haploinsufficient essential gene. Briefly, the loss of one copy of a dosage-sensitive essential gene (gene X), in combination with the inhibition of itself or a related gene (gene Y), or an associated pathway results in lethal consequences for cells. The remarkable and selective clinical efficacy of lenalidomide for the treatment of del(5q) MDS has demonstrated how allelic haploinsufficiency underlies the sensitivity to this drug by synthetic lethality. Differential expression analysis of gene and protein expression in primary AML samples with monosomy 7 revealed significant downregulation of multiple proteasome pathway members at the protein level, but not at the RNA level. Primary AML samples with -7/del(7q) exhibited increased sensitivity (low IC50) to the proteasome inhibitor bortezomib, as evidenced by two independent ex vivo drug screening cohorts (the Beat AML and the FIMM study). Chromosome 7 harbors four proteasome subunits, PSMA2, PSMC2, PSMG3, and SEM1. We performed gene expression, protein expression, copy number analysis, and individual gene knockout experiments. The results have revealed PSMA2 to be a haploinsufficient essential gene on chromosome 7. PSMA2 knockout confers leukemia a growth disadvantage for multiple AML cell clines in both TP53 wild-type and knockout backgrounds. We generated PSMA2 isogenic hemizygous deletion and diploid single-cell clones. PSMA2 hemizygous deletion cells exhibited approximately half the protein expression compared to diploid controls, confirming that PSMA2 is a haploinsufficient gene. PSMA2hemizygous deletion single-cell clones showed significantly enhanced sensitivity to all three evaluated proteasome inhibitors (bortezomib, ixazomib, and carfilzomib), aligning with the sensitivity observed in primary -7/del(7q) leukemia samples. PSMA2 hemizygous deletion cell clones displayed increased p38 and decreased pERK levels upon treatment with proteasome inhibitors, potentially contributing to their increased sensitivity to proteasome inhibitors. Proteomics analysis and in vivo validation is ongoing. As such, we have identified haploinsufficient essential gene PSMA2 mediated proteasome pathway vulnerability in monosomy 7 associated leukemia and further showed that proteasome inhibitors as promising therapeutic approaches for treating hematological disorders associated with monosomy 7.\u0000 Citation Format: Haijiao Zhang, Basil Allen, Daniel Bottomly, Peter Ryabinin, Schannon K. ","PeriodicalId":18791,"journal":{"name":"Molecular Cancer Therapeutics","volume":null,"pages":null},"PeriodicalIF":5.7,"publicationDate":"2024-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141360653","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-10DOI: 10.1158/1538-8514.synthleth24-ia005
Zuzana Tothova
Splicing modulation is a promising treatment strategy pursued to date only in splicing-factor mutant cancers; however, its therapeutic potential is poorly understood outside of this context. Like splicing factors, genes encoding components of the cohesin complex are frequently mutated in myeloid malignancies, including 15-20% of myelodysplastic syndromes (MDS) and secondary acute myeloid leukemia (AML), where they are associated with poor outcomes. I will discuss our recent findings identifying cohesin mutations as biomarkers of sensitivity to drugs targeting splicing-factor SF3B1 (H3B-8800 and E-7107) and describe the mechanism by which drug-induced alterations in splicing of DNA repair genes, such as BRCA1 and BRCA2, underlie this sensitivity. We have demonstrated that treatment of cohesin-mutant cells with SF3B1 modulators results in impaired DNA damage response, accumulation of DNA damage, and increased sensitivity to PARP inhibitors and a panel of chemotherapeutic agents in vitro and in vivo, using AML cell lines and patient-derived xenograft models. Furthermore, we identified RAD51 foci formation as a predictive biomarker of sensitivity to SF3B1 splicing modulation alone or followed by sequential treatment with PARP inhibition and chemotherapy, and have identified additional non-cohesin mutant subtypes of MDS/AML and ovarian and breast cancer which are sensitive to this therapeutic approach. Our findings expand the potential therapeutic benefits of SF3B1 splicing modulators to include cohesin-mutant MDS/AML and we propose this as broader strategy for therapeutic targeting of other DNA damage-repair deficient cancers. Citation Format: Zuzana Tothova. Therapeutic vulnerabilities of cohesin-mutant myeloid malignancies [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr IA005.
剪接调节是一种很有前景的治疗策略,迄今为止只在剪接因子突变的癌症中使用;然而,在这种情况之外,人们对其治疗潜力还知之甚少。与剪接因子一样,编码凝聚素复合物成分的基因也经常在髓系恶性肿瘤中发生突变,包括15-20%的骨髓增生异常综合征(MDS)和继发性急性髓系白血病(AML),它们与不良预后有关。我将讨论我们最近的研究发现,凝聚素突变是靶向剪接因子 SF3B1(H3B-8800 和 E-7107)的药物敏感性的生物标志物,并描述药物诱导的 DNA 修复基因(如 BRCA1 和 BRCA2)剪接改变是这种敏感性的基础机制。我们利用急性髓细胞性白血病细胞系和患者衍生的异种移植模型证明,用 SF3B1 调节剂处理凝聚素突变细胞会导致 DNA 损伤反应受损、DNA 损伤积累以及对 PARP 抑制剂和一系列化疗药物的体外和体内敏感性增加。此外,我们还发现 RAD51 病灶的形成是对单独使用 SF3B1 剪接调节或随后使用 PARP 抑制剂和化疗进行连续治疗的敏感性的预测性生物标志物,并发现了对这种治疗方法敏感的其他非内聚酶突变亚型 MDS/AML、卵巢癌和乳腺癌。我们的研究结果扩大了 SF3B1 剪接调节剂的潜在治疗效果,将粘连蛋白突变的 MDS/AML 包括在内,并建议将其作为针对其他 DNA 损伤修复缺陷癌症的更广泛的治疗策略。引用格式:Zuzana Tothova.粘合素突变型髓系恶性肿瘤的治疗弱点[摘要]。In:AACR 癌症研究特别会议论文集:癌症合成脆弱性的扩展与转化》,2024 年 6 月 10-13 日,加拿大魁北克省蒙特利尔。费城(宾夕法尼亚州):AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr IA005.
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Pub Date : 2024-06-10DOI: 10.1158/1538-8514.synthleth24-a023
J. P. Shen, Yue Gu, S. Chowdhury
Background For decades double knockout (KO) perturbation screens were limited to model organisms such as S. pombe and S. cerevisiae. CRISPR technology has revolutionized genetic interaction discovery by allowing large scale screening in human cell lines, organoids, and mouse models. However, there remains much uncertainty regarding the optimal way to determine the presence of genetic interaction from the raw data generated from these large scale double perturbation experiments. Here we compare two different analysis methods run on the same normalized dataset to determine to what degree does the analysis method influence the determination of genetic interaction. Methods A publicly available genetic interaction dataset containing 24,908 double knock-out constructs across three cell lines (Hela, A549, 293T) in four time points (day 3, 14, 21, 28) and two replicates generated from a pair-wise CRISPR-Cas9 KO screen was used for analysis (Shen et al, Nature Methods, 2017). These data were used to measure single gene fitness scores for 73 known cancer driver genes and all 2628 pair-wise interactions using (1) the numerical Bayesian method from Shen et al, called CTG (Compositional and Time-course-aware Genetic analysis), and (2) the variational Bayesian method GEMINI (Zamanighomi et al, Genome Biology, 2019). Results Single gene KO fitness measurements from CTG and GEMINI were highly correlated for all three cell lines (pearson r 0.678, 0.604, 0.784 for HeLa, A549, and 293T, respectively; p< 0.1 x10-8 for each). In contrast, correlation of genetic interaction scores between the two methods was essentially random: HeLa r= -0.0143, p= 0.47, A549; A549 r= -0.0476, p=0.015; 293T r= -0.0135, p= 0.49. Of 52 synthetic lethal interactions identified by CTG in HeLa at z-score cut off -3, none were identified by GEMINI at same Z cutoff. Conversely of 4 interactions identified by GEMINI, none were identified by CTG. Similarly in A549, of 57 interactions identified by CTG none were identified by GEMINI, of 3 interactions identified by GEMINI none were identified by CTG. Restricting to genetic interactions that were validated in low-throughput drug-drug assays, of 5 synthetic lethal interactions found in HeLa by CTG (CHEK1-MAP2K1, CHEK1-TYMS, ADA-CHEK1, ATM-CHEK1, CDK9-CHEK1) all but CHEK1-TYMS were validated in low-throughput assays. However none of the 5 were scored as hits by GEMINI. Of 3 interactions scored as synthetic lethal in A549 (PRKDC-RRM2, CDK9-PRKDC, CDK4-PRKDC) all but PRKDC-RRM2 were validated, none of the 3 were scored as hits by GEMINI. Conclusions This study highlights dramatic differences in calculated genetic interaction scores from two different computational algorithms applied to the same experimental data. With only 8 of 2628 (0.3%) interactions tested in validation experiments it is not currently possible to know the ground truth in order to assess which method is most accurate. The generation of synthetic genetic interaction data will be a
{"title":"Abstract A023: Determining genetic interaction from double knockout CRISPR screening","authors":"J. P. Shen, Yue Gu, S. Chowdhury","doi":"10.1158/1538-8514.synthleth24-a023","DOIUrl":"https://doi.org/10.1158/1538-8514.synthleth24-a023","url":null,"abstract":"\u0000 Background For decades double knockout (KO) perturbation screens were limited to model organisms such as S. pombe and S. cerevisiae. CRISPR technology has revolutionized genetic interaction discovery by allowing large scale screening in human cell lines, organoids, and mouse models. However, there remains much uncertainty regarding the optimal way to determine the presence of genetic interaction from the raw data generated from these large scale double perturbation experiments. Here we compare two different analysis methods run on the same normalized dataset to determine to what degree does the analysis method influence the determination of genetic interaction. Methods A publicly available genetic interaction dataset containing 24,908 double knock-out constructs across three cell lines (Hela, A549, 293T) in four time points (day 3, 14, 21, 28) and two replicates generated from a pair-wise CRISPR-Cas9 KO screen was used for analysis (Shen et al, Nature Methods, 2017). These data were used to measure single gene fitness scores for 73 known cancer driver genes and all 2628 pair-wise interactions using (1) the numerical Bayesian method from Shen et al, called CTG (Compositional and Time-course-aware Genetic analysis), and (2) the variational Bayesian method GEMINI (Zamanighomi et al, Genome Biology, 2019). Results Single gene KO fitness measurements from CTG and GEMINI were highly correlated for all three cell lines (pearson r 0.678, 0.604, 0.784 for HeLa, A549, and 293T, respectively; p< 0.1 x10-8 for each). In contrast, correlation of genetic interaction scores between the two methods was essentially random: HeLa r= -0.0143, p= 0.47, A549; A549 r= -0.0476, p=0.015; 293T r= -0.0135, p= 0.49. Of 52 synthetic lethal interactions identified by CTG in HeLa at z-score cut off -3, none were identified by GEMINI at same Z cutoff. Conversely of 4 interactions identified by GEMINI, none were identified by CTG. Similarly in A549, of 57 interactions identified by CTG none were identified by GEMINI, of 3 interactions identified by GEMINI none were identified by CTG. Restricting to genetic interactions that were validated in low-throughput drug-drug assays, of 5 synthetic lethal interactions found in HeLa by CTG (CHEK1-MAP2K1, CHEK1-TYMS, ADA-CHEK1, ATM-CHEK1, CDK9-CHEK1) all but CHEK1-TYMS were validated in low-throughput assays. However none of the 5 were scored as hits by GEMINI. Of 3 interactions scored as synthetic lethal in A549 (PRKDC-RRM2, CDK9-PRKDC, CDK4-PRKDC) all but PRKDC-RRM2 were validated, none of the 3 were scored as hits by GEMINI. Conclusions This study highlights dramatic differences in calculated genetic interaction scores from two different computational algorithms applied to the same experimental data. With only 8 of 2628 (0.3%) interactions tested in validation experiments it is not currently possible to know the ground truth in order to assess which method is most accurate. The generation of synthetic genetic interaction data will be a","PeriodicalId":18791,"journal":{"name":"Molecular Cancer Therapeutics","volume":null,"pages":null},"PeriodicalIF":5.7,"publicationDate":"2024-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141362925","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-10DOI: 10.1158/1538-8514.synthleth24-a013
Eleonora Messuti, B. Duso, Alessia Castiglioni, G. Tini, Emanuele Bonetti, Giuseppe Ciossani, Silvia Monzani, Daria Khuntsariya, Marcus Braun, Zdeněk Lánský, Luigi Scietti, Luca Mazzarella
There is great interest in the identification of biomarkers to guide development of antibody-drug conjugates (ADC). Most research has focused on target expression, but key predictors of payload efficacy have not been indeitifed. NF1 is a tumor suppressor classically considered as an inhibitor of RAS signaling, and often mutated in metastatic HER2+ breast cancer (BC). We screened multiple approved drugs for differential sensitivityin CRISPR-engineeredf NF1 KO cells. HER2-targeted agents (small molecules or antibodies) were found to be less effective upon NF1 loss; surprisingly, we identified increased sensitivity to the approved ADC T-DM1, but not to Trastuzumab Deruxtecan (T-Dxd). We then elucidated the underlying molecular cause employing in vivo, in vitro and in vitro reductionist systems. To measure in vivo MT dynamics, we transiently transfected the MT end-binding protein EB3- GFP and reconstructed MT trajectories by live-cell imaging. Upon DM1 treatment, KO cells showed a highly significant reduction in MT speed, demonstrating a direct role for NF1 on MT dynamics in cells. In tubulin polymerization assays, recombinant NF1 greatly accelerated polymerization and completely rescued DM1-induced inhibition. NF1 induced significant MT bundling, a defining feature of many MT-associated proteins, which generates signal indistinguishable from true MT polymerization in turbidity assays. To follow the dynamics of individual microtubules, we applied Total Internal Reflection (TIRF) microscopy on glass-immobilized MTs. As expected, polymerization in the presence of NF1 led to a dose-dependent significant increase in MT dynamics (fraction of elongating MTs, elongation speed, catastrophe rate). Expectedly, DM1 led to significant reduction in the fraction of elongating MTs and speed, but these defects were completely or partially rescued by NF1. Importantly, DM1 did not only lead to MT shortening (as proposed by the current model), but also to clear and frequent MT fracturing, indicating that the drug is not only engaging MT ends but also intra-tubular binding sites. This is consistent with recent models of MT formation which incorporate the frequent presence of areas of discontinuity or damage induced by mechanical stress, exposing intratubular DM1 binding sites. Interestingly, adding NF1 to DM1-treated MTs generated areas of de novo intra-tubular tubulin insertion, coincident with damaged sites, suggesting an entirely novel role for NF1 in MT repair. In conclusion, we provide evidence for a model in which maytansinoids bind not only to soluble tubulin dimers and MT ends, but also to intratubular damaged sites. Thus, the number of binding sites in cells would be proportional to MT damage, suggesting a mechanism for differential efficacy across tumor types and a potential avenue for combinatorial drug development. These results prompt the use of NF1 as a biomarker to select patients for ADC treatment. Funding: FIEO fellowship 2023, AIRC (n25791), Italian
{"title":"Abstract A013: NF1 loss is syntetic lethal with Trastuzumab emtansine","authors":"Eleonora Messuti, B. Duso, Alessia Castiglioni, G. Tini, Emanuele Bonetti, Giuseppe Ciossani, Silvia Monzani, Daria Khuntsariya, Marcus Braun, Zdeněk Lánský, Luigi Scietti, Luca Mazzarella","doi":"10.1158/1538-8514.synthleth24-a013","DOIUrl":"https://doi.org/10.1158/1538-8514.synthleth24-a013","url":null,"abstract":"\u0000 There is great interest in the identification of biomarkers to guide development of antibody-drug conjugates (ADC). Most research has focused on target expression, but key predictors of payload efficacy have not been indeitifed. NF1 is a tumor suppressor classically considered as an inhibitor of RAS signaling, and often mutated in metastatic HER2+ breast cancer (BC). We screened multiple approved drugs for differential sensitivityin CRISPR-engineeredf NF1 KO cells. HER2-targeted agents (small molecules or antibodies) were found to be less effective upon NF1 loss; surprisingly, we identified increased sensitivity to the approved ADC T-DM1, but not to Trastuzumab Deruxtecan (T-Dxd). We then elucidated the underlying molecular cause employing in vivo, in vitro and in vitro reductionist systems. To measure in vivo MT dynamics, we transiently transfected the MT end-binding protein EB3- GFP and reconstructed MT trajectories by live-cell imaging. Upon DM1 treatment, KO cells showed a highly significant reduction in MT speed, demonstrating a direct role for NF1 on MT dynamics in cells. In tubulin polymerization assays, recombinant NF1 greatly accelerated polymerization and completely rescued DM1-induced inhibition. NF1 induced significant MT bundling, a defining feature of many MT-associated proteins, which generates signal indistinguishable from true MT polymerization in turbidity assays. To follow the dynamics of individual microtubules, we applied Total Internal Reflection (TIRF) microscopy on glass-immobilized MTs. As expected, polymerization in the presence of NF1 led to a dose-dependent significant increase in MT dynamics (fraction of elongating MTs, elongation speed, catastrophe rate). Expectedly, DM1 led to significant reduction in the fraction of elongating MTs and speed, but these defects were completely or partially rescued by NF1. Importantly, DM1 did not only lead to MT shortening (as proposed by the current model), but also to clear and frequent MT fracturing, indicating that the drug is not only engaging MT ends but also intra-tubular binding sites. This is consistent with recent models of MT formation which incorporate the frequent presence of areas of discontinuity or damage induced by mechanical stress, exposing intratubular DM1 binding sites. Interestingly, adding NF1 to DM1-treated MTs generated areas of de novo intra-tubular tubulin insertion, coincident with damaged sites, suggesting an entirely novel role for NF1 in MT repair. In conclusion, we provide evidence for a model in which maytansinoids bind not only to soluble tubulin dimers and MT ends, but also to intratubular damaged sites. Thus, the number of binding sites in cells would be proportional to MT damage, suggesting a mechanism for differential efficacy across tumor types and a potential avenue for combinatorial drug development. These results prompt the use of NF1 as a biomarker to select patients for ADC treatment. Funding: FIEO fellowship 2023, AIRC (n25791), Italian ","PeriodicalId":18791,"journal":{"name":"Molecular Cancer Therapeutics","volume":null,"pages":null},"PeriodicalIF":5.7,"publicationDate":"2024-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141363110","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-10DOI: 10.1158/1538-8514.synthleth24-a020
E. Lai, Manuela Neri, E. Sanna, Sonia Nemolato, F. Bardanzellu, M. Scartozzi, Sabrina Giglio, A. Macciò, C. Madeddu
Background and aim of the study: Currently, platinum sensitivity (PS) is a prerequisite for first-line PARP inhibitors (PARPi) in locally advanced and relapsed high grade serous ovarian cancer (HGS-OC). BRCA mutations are recognized as predictive of PS and, therefore, of response to PARPi . Notably, platinum and PARPi cytotoxic action is mainly related by their ability of inducing p53-mediated apoptosis. Therefore, the integrity of p53 machinery is crucial for platinum-related activity whereas the presence of p53 mutation is a fairly frequent event in ovarian cancer, particularly in HGS-OC and in BRCA mutated ones. Patients and methods: We prospectively analyzed 208 women with primary ovarian cancer undergoing surgery at the Department of Gynecologic Oncology, ARNAS G. Brotzu, Cagliari, Italy, between 2019 and 2023. Somatic NGS analysis was performed to detect BRCA and HRD mutations. TP53 mutations were classified according to according to hotspot, structural (missense or nonsense) and functional classification as “gain of function” (GOF) or “loss of function” (LOF), based on the IARC TP53 database. Comparative testing with Fisher's exact test was used to examine TP53 mutation distribution and associations with clinicopathologic factors and PS. The BRCA mutation status was further used to stratify the analysis. Results: Globally, we included 127 adult HGS-OC pts. 84.2% had stage III-IV disease. TP53 mutation was found in 83.4 % of the entire cohort. Somatic BRCA mutations were found in 28.3% pts. Overall, HGS-OC with somatic BRCA mutations had higher TP53 mutation frequency (88.8%) when compared to BRCA WT (81.3%, p=0.1510). Employing the structural classification scheme, most cancers harbored a missense TP53 mutation (76.5%). LOF TP53 mutations were found in 59.4% while GOF in 31.2%. No significant disparity was observed in the distribution of specific TP53 mutations within each classification scheme between cases with BRCA mutations and those without. As for BRCA mutated pts, TP53 WT were all PS. Among those p53 mutated, GOF mutations were associated with PS in 7 pts and platinum resistance in 3 pts; LOF mutations were associated with PS in 7 pts and platinum resistance in 12 pts. The difference in distribution of PS between functional categories of p53 mutations was significant (p=0.0291). As for BRCA WT pts, TP53 WT were all PS. Among TP53 mutated, GOF mutations were associated with PS in 14 pts and platinum resistance in 10 pts; viceversa, LOF mutations were associated with PS in 19 pts and platinum resistance in 25 pts, even if these findings were not statistically significant (p=0.2357). Of relevance, in 5 cases where LOF mutations of p53 was associated with null expression of HIC p53, patients were refractory to platinum-based chemotherapy. Conclusions: Even if preliminary, our data show that HGS-OC harboring p53 null mutations are the poorest prognostic subgroup, especially in terms of PS. Further studies are needed to confirm our find
{"title":"Abstract A020: TP53 mutation and prediction of platinum response in BRCA-mutated ovarian cancer: A prospective case-series analysis","authors":"E. Lai, Manuela Neri, E. Sanna, Sonia Nemolato, F. Bardanzellu, M. Scartozzi, Sabrina Giglio, A. Macciò, C. Madeddu","doi":"10.1158/1538-8514.synthleth24-a020","DOIUrl":"https://doi.org/10.1158/1538-8514.synthleth24-a020","url":null,"abstract":"\u0000 Background and aim of the study: Currently, platinum sensitivity (PS) is a prerequisite for first-line PARP inhibitors (PARPi) in locally advanced and relapsed high grade serous ovarian cancer (HGS-OC). BRCA mutations are recognized as predictive of PS and, therefore, of response to PARPi . Notably, platinum and PARPi cytotoxic action is mainly related by their ability of inducing p53-mediated apoptosis. Therefore, the integrity of p53 machinery is crucial for platinum-related activity whereas the presence of p53 mutation is a fairly frequent event in ovarian cancer, particularly in HGS-OC and in BRCA mutated ones. Patients and methods: We prospectively analyzed 208 women with primary ovarian cancer undergoing surgery at the Department of Gynecologic Oncology, ARNAS G. Brotzu, Cagliari, Italy, between 2019 and 2023. Somatic NGS analysis was performed to detect BRCA and HRD mutations. TP53 mutations were classified according to according to hotspot, structural (missense or nonsense) and functional classification as “gain of function” (GOF) or “loss of function” (LOF), based on the IARC TP53 database. Comparative testing with Fisher's exact test was used to examine TP53 mutation distribution and associations with clinicopathologic factors and PS. The BRCA mutation status was further used to stratify the analysis. Results: Globally, we included 127 adult HGS-OC pts. 84.2% had stage III-IV disease. TP53 mutation was found in 83.4 % of the entire cohort. Somatic BRCA mutations were found in 28.3% pts. Overall, HGS-OC with somatic BRCA mutations had higher TP53 mutation frequency (88.8%) when compared to BRCA WT (81.3%, p=0.1510). Employing the structural classification scheme, most cancers harbored a missense TP53 mutation (76.5%). LOF TP53 mutations were found in 59.4% while GOF in 31.2%. No significant disparity was observed in the distribution of specific TP53 mutations within each classification scheme between cases with BRCA mutations and those without. As for BRCA mutated pts, TP53 WT were all PS. Among those p53 mutated, GOF mutations were associated with PS in 7 pts and platinum resistance in 3 pts; LOF mutations were associated with PS in 7 pts and platinum resistance in 12 pts. The difference in distribution of PS between functional categories of p53 mutations was significant (p=0.0291). As for BRCA WT pts, TP53 WT were all PS. Among TP53 mutated, GOF mutations were associated with PS in 14 pts and platinum resistance in 10 pts; viceversa, LOF mutations were associated with PS in 19 pts and platinum resistance in 25 pts, even if these findings were not statistically significant (p=0.2357). Of relevance, in 5 cases where LOF mutations of p53 was associated with null expression of HIC p53, patients were refractory to platinum-based chemotherapy. Conclusions: Even if preliminary, our data show that HGS-OC harboring p53 null mutations are the poorest prognostic subgroup, especially in terms of PS. Further studies are needed to confirm our find","PeriodicalId":18791,"journal":{"name":"Molecular Cancer Therapeutics","volume":null,"pages":null},"PeriodicalIF":5.7,"publicationDate":"2024-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141365227","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-10DOI: 10.1158/1538-8514.synthleth24-b013
Tyler M. Yasaka, Michael J. Kasper, Li-Ju Wang, Michael Ning, Yufei Huang, S. Monga, Yu-Chiao Chiu
Background: Synthetic essentiality represents a promising therapeutic approach by identifying genes that are necessary for the proliferation and survival of tumors harboring hard-to-target gene alterations. Understanding and accurately predicting synthetic essential genes, through genetic dependencies, may reveal therapeutically effective drug targets in a specific molecular context. Deep learning, as exemplified by our published DeepDEP model, has the potential to capture intricate multi-omic profiles for such prediction tasks. However, the validity of such tools in specific biological contexts remains to be fully examined and presents a major obstacle to adoption by researchers. Materials and Methods: To address this gap, we conducted a case study in which we screened for synthetic essential genes for one of the most frequently mutated and yet undruggable genes, CTNNB1, in hepatocellular carcinoma (HCC). Specifically, we predicted the genetic dependencies of each HCC patient in The Cancer Genome Atlas (TCGA; n=346) by DeepDEP and identified potential dependencies that were intensified with the presence of CTNNB1 mutations. The top 10 genes, ranked by p-value of differential gene-effect scores for CTNNB1-mutated (n=92) versus CTNNB1-WT HCC (n=254), were reviewed in the literature to validate their essentiality in CTNNB1-mutated HCC as well as their potential for pharmacologic inhibition. Survival analysis was performed using published data from the IMBrave150 trial to validate one of the findings. Results: Experimental evidence in the literature supported the essentiality of many of the top 10 predicted genes for CTNNB1-mutated HCC, including one gene with mechanistic evidence of being a transcriptional co-activator of β-catenin target genes. Furthermore, several of these genes have known pharmacologic inhibitors which are either natural compounds or FDA-approved drugs. One example was PDGFB, which encodes a ligand activating the PDGF signaling pathway. PDGF signaling is targeted by sorafenib, an FDA-approved first line drug for HCC. Survival analysis of the sorafenib-treated arm of the IMBrave150 trial showed that patients with mutated CTNNB1 had improved progression-free survival compared to those with wild-type CTNNB1 (p = 0.044). Conclusions: Our study illustrates a potential application of deep learning to identify synthetic essential genes, including genes with readily available pharmacologic inhibitors, for targeting challenging gene alterations. Remarkably, our tool demonstrates the ability to predict cancer dependencies with molecular subtype specificity, suggesting a potential for in silico screening of gene dependencies to facilitate drug discovery and personalized medicine approaches. Our current efforts are focused on optimizing this computational pipeline and making it publicly available for cancer researchers. Citation Format: Tyler M. Yasaka, Michael Kasper, Li-Ju Wang, Michael Ning, Yufei Huang, Satdarshan P Monga, Yu-Chiao C
背景:合成必需基因是一种很有前景的治疗方法,它能确定肿瘤增殖和存活所必需的基因,而这些基因存在难以瞄准的基因改变。通过基因依赖关系了解并准确预测合成必需基因,可以揭示特定分子环境中具有治疗效果的药物靶点。深度学习,如我们已发表的 DeepDEP 模型,有可能捕捉复杂的多基因组图谱来完成此类预测任务。然而,此类工具在特定生物环境中的有效性仍有待充分研究,这也是研究人员采用此类工具的一大障碍。材料与方法:为了填补这一空白,我们进行了一项案例研究,针对肝细胞癌(HCC)中最常见的突变基因之一 CTNNB1 筛选出了合成的重要基因,而 CTNNB1 是最常见的突变基因之一。具体来说,我们通过 DeepDEP 预测了《癌症基因组图谱》(TCGA;n=346)中每位 HCC 患者的基因依赖关系,并确定了 CTNNB1 基因突变会强化的潜在依赖关系。根据 CTNNB1 突变 HCC(n=92)与 CTNNB1-WT HCC(n=254)的差异基因效应得分的 p 值,对排名前 10 位的基因进行了文献回顾,以验证它们在 CTNNB1 突变 HCC 中的重要性以及药物抑制的潜力。利用 IMBrave150 试验的已发表数据进行了生存期分析,以验证其中一项发现。结果:文献中的实验证据支持 CTNNB1 基因突变 HCC 的前 10 个预测基因中的许多基因具有重要意义,其中一个基因的机制证据表明它是β-catenin 靶基因的转录共激活剂。此外,这些基因中还有一些已知的药理抑制剂,它们或是天然化合物,或是美国食品与药物管理局批准的药物。其中一个例子是 PDGFB,它编码一种激活 PDGF 信号通路的配体。索拉非尼是 PDGF 信号通路的靶向药物,索拉非尼是 FDA 批准的治疗 HCC 的一线药物。IMBrave150试验中索拉非尼治疗组的生存期分析表明,与野生型CTNNB1患者相比,突变CTNNB1患者的无进展生存期有所改善(p = 0.044)。结论我们的研究说明了深度学习在识别合成重要基因(包括具有现成药理抑制剂的基因)方面的潜在应用,可用于靶向具有挑战性的基因改变。值得注意的是,我们的工具能够预测具有分子亚型特异性的癌症依赖性,这表明我们有潜力对基因依赖性进行硅学筛选,以促进药物发现和个性化医疗方法。我们目前的工作重点是优化这一计算管道,并将其公开提供给癌症研究人员。引用格式:Tyler M. Yasaka, Michael Kasper, Lii-Ju Wang, Michael Ning, Yufei Huang, Satdarshan P Monga, Yu-Chiao Chiu.基于深度学习的 CTNNB1 突变肝细胞癌合成本质预测 [摘要].In:AACR 癌症研究特别会议论文集:扩展和转化癌症合成脆弱性;2024 年 6 月 10-13 日;加拿大魁北克省蒙特利尔。费城(宾夕法尼亚州):AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr B013.
{"title":"Abstract B013: Deep learning-based prediction of synthetic essentialities in CTNNB1-mutated hepatocellular carcinoma","authors":"Tyler M. Yasaka, Michael J. Kasper, Li-Ju Wang, Michael Ning, Yufei Huang, S. Monga, Yu-Chiao Chiu","doi":"10.1158/1538-8514.synthleth24-b013","DOIUrl":"https://doi.org/10.1158/1538-8514.synthleth24-b013","url":null,"abstract":"\u0000 Background: Synthetic essentiality represents a promising therapeutic approach by identifying genes that are necessary for the proliferation and survival of tumors harboring hard-to-target gene alterations. Understanding and accurately predicting synthetic essential genes, through genetic dependencies, may reveal therapeutically effective drug targets in a specific molecular context. Deep learning, as exemplified by our published DeepDEP model, has the potential to capture intricate multi-omic profiles for such prediction tasks. However, the validity of such tools in specific biological contexts remains to be fully examined and presents a major obstacle to adoption by researchers. Materials and Methods: To address this gap, we conducted a case study in which we screened for synthetic essential genes for one of the most frequently mutated and yet undruggable genes, CTNNB1, in hepatocellular carcinoma (HCC). Specifically, we predicted the genetic dependencies of each HCC patient in The Cancer Genome Atlas (TCGA; n=346) by DeepDEP and identified potential dependencies that were intensified with the presence of CTNNB1 mutations. The top 10 genes, ranked by p-value of differential gene-effect scores for CTNNB1-mutated (n=92) versus CTNNB1-WT HCC (n=254), were reviewed in the literature to validate their essentiality in CTNNB1-mutated HCC as well as their potential for pharmacologic inhibition. Survival analysis was performed using published data from the IMBrave150 trial to validate one of the findings. Results: Experimental evidence in the literature supported the essentiality of many of the top 10 predicted genes for CTNNB1-mutated HCC, including one gene with mechanistic evidence of being a transcriptional co-activator of β-catenin target genes. Furthermore, several of these genes have known pharmacologic inhibitors which are either natural compounds or FDA-approved drugs. One example was PDGFB, which encodes a ligand activating the PDGF signaling pathway. PDGF signaling is targeted by sorafenib, an FDA-approved first line drug for HCC. Survival analysis of the sorafenib-treated arm of the IMBrave150 trial showed that patients with mutated CTNNB1 had improved progression-free survival compared to those with wild-type CTNNB1 (p = 0.044). Conclusions: Our study illustrates a potential application of deep learning to identify synthetic essential genes, including genes with readily available pharmacologic inhibitors, for targeting challenging gene alterations. Remarkably, our tool demonstrates the ability to predict cancer dependencies with molecular subtype specificity, suggesting a potential for in silico screening of gene dependencies to facilitate drug discovery and personalized medicine approaches. Our current efforts are focused on optimizing this computational pipeline and making it publicly available for cancer researchers.\u0000 Citation Format: Tyler M. Yasaka, Michael Kasper, Li-Ju Wang, Michael Ning, Yufei Huang, Satdarshan P Monga, Yu-Chiao C","PeriodicalId":18791,"journal":{"name":"Molecular Cancer Therapeutics","volume":null,"pages":null},"PeriodicalIF":5.7,"publicationDate":"2024-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141365448","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-10DOI: 10.1158/1538-8514.synthleth24-b015
Rand Arafeh, Laura Chang, Lydia Sawyer, Helen Wang, James McFarland, Joshua Dempster, Peter C. DeWeirdt, John G Doench, William C Hahn
This abstract is being presented as a short talk in the scientific program. A full abstract is printed in the Proffered Abstracts section (PR014) of the Conference Program/Proceedings. Citation Format: Rand Arafeh, Laura Chang, Lydia Sawyer, Helen Wang, James McFarland, Joshua Dempster, Peter DeWeirdt, John Doench, William C Hahn. Combinatorial genetic screens to map synthetic lethal interactions and identify new cancer drug targets in KRAS mutant cancers [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr B015.
本摘要将作为科学会议日程中的简短发言。完整摘要刊载于大会日程/论文集的 "提供摘要 "部分 (PR014)。引用格式:Rand Arafeh, Laura Chang, Lydia Sawyer, Helen Wang, James McFarland, Joshua Dempster, Peter DeWeirdt, John Doench, William C Hahn.通过组合遗传筛选绘制合成致死相互作用图并确定 KRAS 突变癌症的新抗癌药物靶点 [摘要].In:AACR 癌症研究特别会议论文集:扩大和转化癌症合成弱点;2024 年 6 月 10-13 日;加拿大魁北克省蒙特利尔。费城(宾夕法尼亚州):AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr B015.
{"title":"Abstract B015: Combinatorial genetic screens to map synthetic lethal interactions and identify new cancer drug targets in KRAS mutant cancers","authors":"Rand Arafeh, Laura Chang, Lydia Sawyer, Helen Wang, James McFarland, Joshua Dempster, Peter C. DeWeirdt, John G Doench, William C Hahn","doi":"10.1158/1538-8514.synthleth24-b015","DOIUrl":"https://doi.org/10.1158/1538-8514.synthleth24-b015","url":null,"abstract":"\u0000 This abstract is being presented as a short talk in the scientific program. A full abstract is printed in the Proffered Abstracts section (PR014) of the Conference Program/Proceedings.\u0000 Citation Format: Rand Arafeh, Laura Chang, Lydia Sawyer, Helen Wang, James McFarland, Joshua Dempster, Peter DeWeirdt, John Doench, William C Hahn. Combinatorial genetic screens to map synthetic lethal interactions and identify new cancer drug targets in KRAS mutant cancers [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr B015.","PeriodicalId":18791,"journal":{"name":"Molecular Cancer Therapeutics","volume":null,"pages":null},"PeriodicalIF":5.7,"publicationDate":"2024-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141363909","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-10DOI: 10.1158/1538-8514.synthleth24-a011
Wanjun Tang, Karrie Mei Yee Kiang, Gilberto Ka Kit Leung
Chaperone-mediated autophagy (CMA) is a selective form of autophagy that targets proteins with the KFERQ-like motif for lysosomal degradation. This process involves the binding of substrate proteins to the chaperone HSC70 and their subsequent transport to the receptor LAMP2A on the lysosome membrane for degradation. Previous studies have shown a significant increase in the expression level of LAMP2A and the CMA activity in glioblastoma (GBM). However, the comprehensive role of CMA in GBM pathogenesis remains unclear. In this study, we observed a progressive elevation in LAMP2A expression with increasing GBM cell confluence. Notably, inhibition of CMA through LAMP2A knockdown markedly promoted GBM growth in orthotopic mouse models, accompanied by upregulation of positive cell cycle regulators. Conversely, overexpression of LAMP2A delayed GBM cell growth and increased sub-G1 cell percentages. Treatment with CMA activators induced cell death in GBM cells while preserving normal astrocyte viability at equivalent concentrations. Our results suggest that the upregulated expression of LAMP2A in GBM cells may be influenced by the tumor microenvironment rather than being a primary driver of GBM aggressiveness. Furthermore, activating CMA could serve as a potential therapeutic strategy to inhibit GBM growth while safeguarding normal cell viability. Citation Format: Wanjun Tang, Karrie Mei Yee Kiang, Gilberto Ka Kit Leung. Enhancing chaperone-mediated autophagy to impede glioblastoma growth [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr A011.
{"title":"Abstract A011: Enhancing chaperone-mediated autophagy to impede glioblastoma growth","authors":"Wanjun Tang, Karrie Mei Yee Kiang, Gilberto Ka Kit Leung","doi":"10.1158/1538-8514.synthleth24-a011","DOIUrl":"https://doi.org/10.1158/1538-8514.synthleth24-a011","url":null,"abstract":"\u0000 Chaperone-mediated autophagy (CMA) is a selective form of autophagy that targets proteins with the KFERQ-like motif for lysosomal degradation. This process involves the binding of substrate proteins to the chaperone HSC70 and their subsequent transport to the receptor LAMP2A on the lysosome membrane for degradation. Previous studies have shown a significant increase in the expression level of LAMP2A and the CMA activity in glioblastoma (GBM). However, the comprehensive role of CMA in GBM pathogenesis remains unclear. In this study, we observed a progressive elevation in LAMP2A expression with increasing GBM cell confluence. Notably, inhibition of CMA through LAMP2A knockdown markedly promoted GBM growth in orthotopic mouse models, accompanied by upregulation of positive cell cycle regulators. Conversely, overexpression of LAMP2A delayed GBM cell growth and increased sub-G1 cell percentages. Treatment with CMA activators induced cell death in GBM cells while preserving normal astrocyte viability at equivalent concentrations. Our results suggest that the upregulated expression of LAMP2A in GBM cells may be influenced by the tumor microenvironment rather than being a primary driver of GBM aggressiveness. Furthermore, activating CMA could serve as a potential therapeutic strategy to inhibit GBM growth while safeguarding normal cell viability.\u0000 Citation Format: Wanjun Tang, Karrie Mei Yee Kiang, Gilberto Ka Kit Leung. Enhancing chaperone-mediated autophagy to impede glioblastoma growth [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr A011.","PeriodicalId":18791,"journal":{"name":"Molecular Cancer Therapeutics","volume":null,"pages":null},"PeriodicalIF":5.7,"publicationDate":"2024-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141365687","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-10DOI: 10.1158/1538-8514.synthleth24-ia018
Shruti Bhatt
Acute myeloid leukemia (AML) is a complex and genetically diverse with an overall survival rate of less than 32%. Despite the remarkable success of targeted therapy in mediating remission, the emergence of acquired resistance remains a major clinical challenge to overcome. The prevailing understanding of acquired resistance identifies successive genetic changes as the primary cause.TP53 mutations are found in 70-80% of acute myeloid leukemia (AML) patients with complex karyotypes and associated with resistance towards both conventional chemotherapy and newly approved venetoclax plus azacytidine (VEN/AZA) combination. By using CRISPR-Cas9-edited isogenic AML cells harboring, mutation (6 missense mutations) or deletion (KO) of TP53, we found that TP53 mutant/KO cells are less sensitive to etoposide or VEN-AZA induced apoptosis compared to WT without defect in G1 arrest. Surprisingly, we found that TP53-mutant and TP53-wild-type (WT) isogenic AML cells and primary tumors (n=40) had comparable mitochondrial outer membrane permeabilization (MOMP) at baseline, despite the key role of TP53 in transcriptionally activating proapoptotic regulators of MOMP (such as BAX, PUMA, and NOXA). Based on these findings, we hypothesize that the targets downstream of mitochondrial permeabilization drive resistance to HMA/VEN combinations in TP53 mutant disease. By leveraging unbiased bulk RNA-seq and proteomics, and whole genome CRISPR-cas9 screen we identified IAPs as functional vulnerability. Collectively we reveal novel chemoresistance mechanisms in TP53 mutant/KO downstream of MOMP and provide a targeting strategy to improve existing therapy by targeting non-transcriptional function of TP53 in overcoming therapy resistance. Citation Format: Shruti Bhatt. Vulnerabilities of TP53-mutated AML and therapeutic implications [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr IA018.
{"title":"Abstract IA018: Vulnerabilities of TP53-mutated AML and therapeutic implications","authors":"Shruti Bhatt","doi":"10.1158/1538-8514.synthleth24-ia018","DOIUrl":"https://doi.org/10.1158/1538-8514.synthleth24-ia018","url":null,"abstract":"\u0000 Acute myeloid leukemia (AML) is a complex and genetically diverse with an overall survival rate of less than 32%. Despite the remarkable success of targeted therapy in mediating remission, the emergence of acquired resistance remains a major clinical challenge to overcome. The prevailing understanding of acquired resistance identifies successive genetic changes as the primary cause.TP53 mutations are found in 70-80% of acute myeloid leukemia (AML) patients with complex karyotypes and associated with resistance towards both conventional chemotherapy and newly approved venetoclax plus azacytidine (VEN/AZA) combination. By using CRISPR-Cas9-edited isogenic AML cells harboring, mutation (6 missense mutations) or deletion (KO) of TP53, we found that TP53 mutant/KO cells are less sensitive to etoposide or VEN-AZA induced apoptosis compared to WT without defect in G1 arrest. Surprisingly, we found that TP53-mutant and TP53-wild-type (WT) isogenic AML cells and primary tumors (n=40) had comparable mitochondrial outer membrane permeabilization (MOMP) at baseline, despite the key role of TP53 in transcriptionally activating proapoptotic regulators of MOMP (such as BAX, PUMA, and NOXA). Based on these findings, we hypothesize that the targets downstream of mitochondrial permeabilization drive resistance to HMA/VEN combinations in TP53 mutant disease. By leveraging unbiased bulk RNA-seq and proteomics, and whole genome CRISPR-cas9 screen we identified IAPs as functional vulnerability. Collectively we reveal novel chemoresistance mechanisms in TP53 mutant/KO downstream of MOMP and provide a targeting strategy to improve existing therapy by targeting non-transcriptional function of TP53 in overcoming therapy resistance.\u0000 Citation Format: Shruti Bhatt. Vulnerabilities of TP53-mutated AML and therapeutic implications [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr IA018.","PeriodicalId":18791,"journal":{"name":"Molecular Cancer Therapeutics","volume":null,"pages":null},"PeriodicalIF":5.7,"publicationDate":"2024-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141361213","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-10DOI: 10.1158/1538-8514.synthleth24-a026
Ruihan Luo
Deciphering disease progression and the sophisticated tumor ecosystems is imperative for exploring tumorigenesis mechanisms and developing novel prevention strategies. Here, we dissected heterogeneous tissue microenvironments during malignant transitions by leveraging data from 4,972,145 single cells in 1396 samples from 62 datasets spanning 13 major tissue types. Within transitional stem-like subpopulations highly enriched in precancerous lesions and cancers, we identified 30 recurring cellular states, including hypoxia and epithelial senescence, revealing a high degree of plasticity in epithelial stem cells. By characterizing the dynamics of stem-cell crosstalk with the microenvironment along the pseudotime axis, we uncovered distinct roles of ANXA1 at different stages of tumor development. ANXA1 expression levels in stem cells were decreased from the healthy to the precancerous stages, which promoted inflammatory responses by recruiting neutrophils and regulating monocyte differentiation towards M1 macrophages. In contrast, during malignant progression, upregulated ANXA1 fostered M2 macrophage polarization and cancer-associated fibroblast transformation. Our spatiotemporal analysis further provided insights into mechanisms responsible for immunosuppression. Collectively, this study provided a systematic view of cancer origins, and suggested that restoring and maintaining the balance of inflammation and their mediators (e.g., AnxA1/FPRs signaling) may represent a novel approach to control the evolution of precancerous lesions and mitigate the risk for cancer development. Citation Format: Ruihan Luo. Single-cell landscape deciphering cancer cell-of-origin and cellular heterogeneity in malignant transformation of 13 major tissues [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr A026.
{"title":"Abstract A026: Single-cell landscape deciphering cancer Single-cell landscape deciphering cancer cell-of-origin and cellular heterogeneity in malignant transformation of 13 major tissues","authors":"Ruihan Luo","doi":"10.1158/1538-8514.synthleth24-a026","DOIUrl":"https://doi.org/10.1158/1538-8514.synthleth24-a026","url":null,"abstract":"\u0000 Deciphering disease progression and the sophisticated tumor ecosystems is imperative for exploring tumorigenesis mechanisms and developing novel prevention strategies. Here, we dissected heterogeneous tissue microenvironments during malignant transitions by leveraging data from 4,972,145 single cells in 1396 samples from 62 datasets spanning 13 major tissue types. Within transitional stem-like subpopulations highly enriched in precancerous lesions and cancers, we identified 30 recurring cellular states, including hypoxia and epithelial senescence, revealing a high degree of plasticity in epithelial stem cells. By characterizing the dynamics of stem-cell crosstalk with the microenvironment along the pseudotime axis, we uncovered distinct roles of ANXA1 at different stages of tumor development. ANXA1 expression levels in stem cells were decreased from the healthy to the precancerous stages, which promoted inflammatory responses by recruiting neutrophils and regulating monocyte differentiation towards M1 macrophages. In contrast, during malignant progression, upregulated ANXA1 fostered M2 macrophage polarization and cancer-associated fibroblast transformation. Our spatiotemporal analysis further provided insights into mechanisms responsible for immunosuppression. Collectively, this study provided a systematic view of cancer origins, and suggested that restoring and maintaining the balance of inflammation and their mediators (e.g., AnxA1/FPRs signaling) may represent a novel approach to control the evolution of precancerous lesions and mitigate the risk for cancer development.\u0000 Citation Format: Ruihan Luo. Single-cell landscape deciphering cancer cell-of-origin and cellular heterogeneity in malignant transformation of 13 major tissues [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr A026.","PeriodicalId":18791,"journal":{"name":"Molecular Cancer Therapeutics","volume":null,"pages":null},"PeriodicalIF":5.7,"publicationDate":"2024-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141361882","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}