RESUMO
The prevailing approach to addressing secondary drug resistance in cancer focuses on treating the resistance mechanisms at relapse. However, the dynamic nature of clonal evolution, along with potential fitness costs and cost compensations, may present exploitable vulnerabilities-a notion that we term "temporal collateral sensitivity." Using a combined pharmacological screen and drug resistance selection approach in a murine model of Ph(+) acute lymphoblastic leukemia, we indeed find that temporal and/or persistent collateral sensitivity to non-classical BCR-ABL1 drugs arises in emergent tumor subpopulations during the evolution of resistance toward initial treatment with BCR-ABL1-targeted inhibitors. We determined the sensitization mechanism via genotypic, phenotypic, signaling, and binding measurements in combination with computational models and demonstrated significant overall survival extension in mice. Additional stochastic mathematical models and small-molecule screens extended our insights, indicating the value of focusing on evolutionary trajectories and pharmacological profiles to identify new strategies to treat dynamic tumor vulnerabilities.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Modelos Biológicos , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Animais , Ensaios de Seleção de Medicamentos Antitumorais , Camundongos , Cromossomo Filadélfia , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Proteínas Proto-Oncogênicas c-bcr/análise , Proteínas Proto-Oncogênicas c-bcr/genéticaRESUMO
Although rarely used in nature, fluorine has emerged as an important elemental ingredient in the design of proteins with altered folding, stability, oligomerization propensities, and bioactivity. Adding to the molecular modification toolbox, here we report the ability of privileged perfluorinated amphiphiles to noncovalently decorate proteins to alter their conformational plasticity and potentiate their dispersion into fluorous phases. Employing a complementary suite of biophysical, in-silico and in-vitro approaches, we establish structure-activity relationships defining these phenomena and investigate their impact on protein structural dynamics and intracellular trafficking. Notably, we show that the lead compound, perfluorononanoic acid, is 106 times more potent in inducing non-native protein secondary structure in select proteins than is the well-known helix inducer trifluoroethanol, and also significantly enhances the cellular uptake of complexed proteins. These findings could advance the rational design of fluorinated proteins, inform on potential modes of toxicity for perfluoroalkyl substances, and guide the development of fluorine-modified biologics with desirable functional properties for drug discovery and delivery applications.
Assuntos
Flúor , Proteínas , Flúor/química , Proteínas/química , Estrutura Secundária de Proteína , TrifluoretanolRESUMO
Small molecules can affect many cellular processes. The disambiguation of these effects to identify the causative mechanisms of cell death is extremely challenging. This challenge impacts both clinical development and the interpretation of chemical genetic experiments. CX-5461 was developed as a selective RNA polymerase I inhibitor, but recent evidence suggests that it may cause DNA damage and induce G-quadraplex formation. Here we use three complimentary data mining modalities alongside biochemical and cell biological assays to show that CX-5461 exerts its primary cytotoxic activity through topoisomerase II poisoning. We then show that acquired resistance to CX-5461 in previously sensitive lymphoma cells confers collateral resistance to the topoisomerase II poison doxorubicin. Doxorubicin is already a frontline chemotherapy in a variety of hematopoietic malignancies, and CX-5461 is being tested in relapse/refractory hematopoietic tumors. Our data suggest that the mechanism of cell death induced by CX-5461 is critical for rational clinical development in these patients. Moreover, CX-5461 usage as a specific chemical genetic probe of RNA polymerase I function is challenging to interpret. Our multimodal data-driven approach is a useful way to detangle the intended and unintended mechanisms of drug action across diverse essential cellular processes.
Assuntos
Antineoplásicos/farmacologia , Benzotiazóis/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Naftiridinas/farmacologia , Proteínas de Ligação a Poli-ADP-Ribose/antagonistas & inibidores , Linhagem Celular Tumoral , DNA Topoisomerases Tipo II/genética , DNA Topoisomerases Tipo II/metabolismo , Relação Dose-Resposta a Droga , Doxorrubicina/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Linfoma , Proteínas de Ligação a Poli-ADP-Ribose/genética , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo , Interferência de RNA , Sensibilidade e EspecificidadeRESUMO
We performed a genome-scale shRNA screen for modulators of B-cell leukemia progression in vivo. Results from this work revealed dramatic distinctions between the relative effects of shRNAs on the growth of tumor cells in culture versus in their native microenvironment. Specifically, we identified many "context-specific" regulators of leukemia development. These included the gene encoding the zinc finger protein Phf6. While inactivating mutations in PHF6 are commonly observed in human myeloid and T-cell malignancies, we found that Phf6 suppression in B-cell malignancies impairs tumor progression. Thus, Phf6 is a "lineage-specific" cancer gene that plays opposing roles in developmentally distinct hematopoietic malignancies.
Assuntos
Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Leucemia/genética , Linhagem da Célula , Proliferação de Células/genética , Genoma Humano/genética , Humanos , Leucemia/fisiopatologia , Mutação/genética , RNA Interferente Pequeno/genética , Proteínas RepressorasRESUMO
Metastatic cancer has a poor prognosis, because it is broadly disseminated and associated with both intrinsic and acquired drug resistance. Critical unmet needs in effectively killing drug resistant cancer cells include overcoming the drug desensitization characteristics of some metastatic cancers/lesions, and tailoring therapeutic regimens to both the tumor microenvironment and the genetic profiles of the resident cancer cells. Bioengineers and materials scientists are developing technologies to determine how metastatic sites exclude therapies, and how extracellular factors (including cells, proteins, metabolites, extracellular matrix, and abiotic factors) at metastatic sites significantly affect drug pharmacodynamics. Two looming challenges are determining which feature, or combination of features, from the tumor microenvironment drive drug resistance, and what the relative impact is of extracellular signals vs. intrinsic cell genetics in determining drug response. Sophisticated systems biology tools that can de-convolve a crowded network of signals and responses, as well as controllable microenvironments capable of providing discrete and tunable extracellular cues can help us begin to interrogate the high dimensional interactions governing drug resistance in patients.
Assuntos
Neoplasias , Resistência a Medicamentos , Matriz Extracelular/metabolismo , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/genética , Microambiente TumoralRESUMO
BACKGROUND: When three SARS-CoV-2 vaccines came to market in Europe and North America in the winter of 2020-2021, distribution networks were in a race against a major epidemiological wave of SARS-CoV-2 that began in autumn 2020. Rapid and optimized vaccine allocation was critical during this time. With 95% efficacy reported for two of the vaccines, near-term public health needs likely require that distribution is prioritized to the elderly, health care workers, teachers, essential workers, and individuals with comorbidities putting them at risk of severe clinical progression. METHODS: We evaluate various age-based vaccine distributions using a validated mathematical model based on current epidemic trends in Rhode Island and Massachusetts. We allow for varying waning efficacy of vaccine-induced immunity, as this has not yet been measured. We account for the fact that known COVID-positive cases may not have been included in the first round of vaccination. And, we account for age-specific immune patterns in both states at the time of the start of the vaccination program. Our analysis assumes that health systems during winter 2020-2021 had equal staffing and capacity to previous phases of the SARS-CoV-2 epidemic; we do not consider the effects of understaffed hospitals or unvaccinated medical staff. RESULTS: We find that allocating a substantial proportion (>75%) of vaccine supply to individuals over the age of 70 is optimal in terms of reducing total cumulative deaths through mid-2021. This result is robust to different profiles of waning vaccine efficacy and several different assumptions on age mixing during and after lockdown periods. As we do not explicitly model other high-mortality groups, our results on vaccine allocation apply to all groups at high risk of mortality if infected. A median of 327 to 340 deaths can be avoided in Rhode Island (3444 to 3647 in Massachusetts) by optimizing vaccine allocation and vaccinating the elderly first. The vaccination campaigns are expected to save a median of 639 to 664 lives in Rhode Island and 6278 to 6618 lives in Massachusetts in the first half of 2021 when compared to a scenario with no vaccine. A policy of vaccinating only seronegative individuals avoids redundancy in vaccine use on individuals that may already be immune, and would result in 0.5% to 1% reductions in cumulative hospitalizations and deaths by mid-2021. CONCLUSIONS: Assuming high vaccination coverage (>28%) and no major changes in distancing, masking, gathering size, hygiene guidelines, and virus transmissibility between 1 January 2021 and 1 July 2021 a combination of vaccination and population immunity may lead to low or near-zero transmission levels by the second quarter of 2021.
Assuntos
Vacinas contra COVID-19/provisão & distribuição , COVID-19 , Controle de Doenças Transmissíveis/organização & administração , Alocação de Recursos para a Atenção à Saúde/organização & administração , Alocação de Recursos/organização & administração , Cobertura Vacinal , Vacinação , Fatores Etários , COVID-19/epidemiologia , COVID-19/prevenção & controle , Humanos , Incidência , Massachusetts/epidemiologia , Modelos Teóricos , Saúde Pública/métodos , Saúde Pública/normas , Rhode Island/epidemiologia , SARS-CoV-2 , Vacinação/métodos , Vacinação/estatística & dados numéricos , Cobertura Vacinal/estatística & dados numéricos , Cobertura Vacinal/provisão & distribuiçãoRESUMO
Ruthenium (Ru) and osmium (Os) complexes are of sustained interest in cancer research and may be alternative to platinum-based therapy. We detail here three new series of ruthenium and osmium complexes, supported by physico-chemical characterizations, including time-dependent density functional theory, a combined experimental and computational study on the aquation reactions and the nature of the metal-arene bond. Cytotoxic profiles were then evaluated on several cancer cell lines although with limited success. Further investigations were, however, performed on the most active series using a genetic approach based on RNA interference and highlighted a potential multi-target mechanism of action through topoisomerase II, mitotic spindle, HDAC and DNMT inhibition.
Assuntos
Antineoplásicos/farmacologia , Biotina/farmacologia , Complexos de Coordenação/farmacologia , Morfolinas/farmacologia , Osmio/farmacologia , Rutênio/farmacologia , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Biotina/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Cristalografia por Raios X , Teoria da Densidade Funcional , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Camundongos , Modelos Moleculares , Estrutura Molecular , Morfolinas/química , Osmio/química , Rutênio/químicaRESUMO
The somatic co-evolution of tumors and the cellular immune responses that combat them drives the diversity of immune-tumor interactions. This includes tumor mutations that generate neo-antigenic epitopes that elicit cytotoxic T-cell activity and subsequent pressure to select for genetic loss of antigen presentation. Most studies have focused on how tumor missense mutations can drive tumor immunity, but frameshift mutations have the potential to create far greater antigenic diversity. However, expression of this antigenic diversity is potentially regulated by Nonsense Mediated Decay (NMD) and NMD has been shown to be of variable efficiency in cancers. Here we studied how mutational changes influence global NMD and cytolytic immune responses. Using TCGA datasets, we derived novel patient-level metrics of 'NMD burden' and interrogated how different mutation and most importantly NMD burdens influence cytolytic activity using machine learning models and survival outcomes. We find that NMD is a significant and independent predictor of immune cytolytic activity. Different indications exhibited varying dependence on NMD and mutation burden features. We also observed significant co-alteration of genes in the NMD pathway, with a global increase in NMD efficiency in patients with NMD co-alterations. Finally, NMD burden also stratified patient survival in multivariate regression models in subset of cancer types. Our work suggests that beyond selecting for mutations that elicit NMD in tumor suppressors, tumor evolution may react to the selective pressure generated by inflammation to globally enhance NMD through coordinated amplification and/or mutation.
Assuntos
Citotoxicidade Imunológica/genética , Neoplasias/genética , Degradação do RNAm Mediada por Códon sem Sentido/fisiologia , Evolução Biológica , Simulação por Computador , Citosol/metabolismo , Bases de Dados Genéticas , Evolução Molecular , Mutação da Fase de Leitura/genética , Humanos , Aprendizado de Máquina , Mutação/genética , Mutação de Sentido Incorreto/genéticaRESUMO
The identification of biologically significant variants in cancer genomes is critical to therapeutic discovery, but it is limited by the statistical power needed to discern driver from passenger. Independent biological data can be used to filter cancer exomes and increase statistical power. Large genetic databases for inherited diseases are uniquely suited to this task because they contain specific amino acid alterations with known pathogenicity and molecular mechanisms. However, no rigorous method to overlay this information onto the cancer exome exists. Here, we present a computational methodology that overlays any variant database onto the somatic mutations in all cancer exomes. We validate the computation experimentally and identify novel associations in a re-analysis of 7362 cancer exomes. This analysis identified activating SOS1 mutations associated with Noonan syndrome as significantly altered in melanoma and the first kinase-activating mutations in ACVR1 associated with adult tumors. Beyond a filter, significant variants found in both rare cancers and rare inherited diseases increase the unmet medical need for therapeutics that target these variants and may bootstrap drug discovery efforts in orphan indications.
Assuntos
Receptores de Ativinas Tipo I/genética , Colágeno Tipo III/genética , Biologia Computacional/métodos , Bases de Dados Genéticas , Neoplasias do Endométrio/genética , Predisposição Genética para Doença , Neoplasias Pulmonares/genética , Melanoma/genética , Síndrome de Noonan/genética , Proteína SOS1/genética , Sequência de Aminoácidos , Linhagem Celular , Exoma/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Variação Genética/genética , Células HEK293 , Humanos , Mutação/genética , FosforilaçãoRESUMO
Transposon-insertion sequencing (TIS) is a powerful approach for deciphering genetic requirements for bacterial growth in different conditions, as it enables simultaneous genome-wide analysis of the fitness of thousands of mutants. However, current methods for comparative analysis of TIS data do not adjust for stochastic experimental variation between datasets and are limited to interrogation of annotated genomic elements. Here, we present ARTIST, an accessible TIS analysis pipeline for identifying essential regions that are required for growth under optimal conditions as well as conditionally essential loci that participate in survival only under specific conditions. ARTIST uses simulation-based normalization to model and compensate for experimental noise, and thereby enhances the statistical power in conditional TIS analyses. ARTIST also employs a novel adaptation of the hidden Markov model to generate statistically robust, high-resolution, annotation-independent maps of fitness-linked loci across the entire genome. Using ARTIST, we sensitively and comprehensively define Mycobacterium tuberculosis and Vibrio cholerae loci required for host infection while limiting inclusion of false positive loci. ARTIST is applicable to a broad range of organisms and will facilitate TIS-based dissection of pathways required for microbial growth and survival under a multitude of conditions.
Assuntos
Elementos de DNA Transponíveis/genética , Interações Hospedeiro-Patógeno/genética , Mutagênese Insercional/genética , Software , Simulação por Computador , Deriva Genética , Sequenciamento de Nucleotídeos em Larga Escala , Cadeias de Markov , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/patogenicidade , Vibrio cholerae/genética , Vibrio cholerae/patogenicidadeRESUMO
Combination chemotherapies have been a mainstay in the treatment of disseminated malignancies for almost 60 y, yet even successful regimens fail to cure many patients. Although their single-drug components are well studied, the mechanisms by which drugs work together in clinical combination regimens are poorly understood. Here, we combine RNAi-based functional signatures with complementary informatics tools to examine drug combinations. This approach seeks to bring to combination therapy what the knowledge of biochemical targets has brought to single-drug therapy and creates a statistical and experimental definition of "combination drug mechanisms of action." We show that certain synergistic drug combinations may act as a more potent version of a single drug. Conversely, unlike these highly synergistic combinations, most drugs average extant single-drug variations in therapeutic response. When combined to form multidrug regimens, averaging combinations form averaging regimens that homogenize genetic variation in mouse models of cancer and in clinical genomics datasets. We suggest surprisingly simple and predictable combination mechanisms of action that are independent of biochemical mechanism and have implications for biomarker discovery as well as for the development of regimens with defined genetic dependencies.
Assuntos
Combinação de Medicamentos , Desenho de Fármacos , Sinergismo Farmacológico , Variação Genética , Neoplasias/tratamento farmacológico , Animais , Biomarcadores/metabolismo , Humanos , Camundongos , Neoplasias/genética , Análise de Componente Principal , Interferência de RNA , Biologia de SistemasRESUMO
The coupling of high-density transposon mutagenesis to high-throughput DNA sequencing (transposon-insertion sequencing) enables simultaneous and genome-wide assessment of the contributions of individual loci to bacterial growth and survival. We have refined analysis of transposon-insertion sequencing data by normalizing for the effect of DNA replication on sequencing output and using a hidden Markov model (HMM)-based filter to exploit heretofore unappreciated information inherent in all transposon-insertion sequencing data sets. The HMM can smooth variations in read abundance and thereby reduce the effects of read noise, as well as permit fine scale mapping that is independent of genomic annotation and enable classification of loci into several functional categories (e.g. essential, domain essential or 'sick'). We generated a high-resolution map of genomic loci (encompassing both intra- and intergenic sequences) that are required or beneficial for in vitro growth of the cholera pathogen, Vibrio cholerae. This work uncovered new metabolic and physiologic requirements for V. cholerae survival, and by combining transposon-insertion sequencing and transcriptomic data sets, we also identified several novel noncoding RNA species that contribute to V. cholerae growth. Our findings suggest that HMM-based approaches will enhance extraction of biological meaning from transposon-insertion sequencing genomic data.
Assuntos
Elementos de DNA Transponíveis , Genes Bacterianos , Sequenciamento de Nucleotídeos em Larga Escala , Análise de Sequência de DNA , Vibrio cholerae/genética , Regiões 5' não Traduzidas , Escherichia coli/genética , Biblioteca Gênica , Genes Essenciais , Loci Gênicos , Cadeias de Markov , RNA não Traduzido/genética , Vibrio cholerae/crescimento & desenvolvimentoRESUMO
BACKGROUND: Mycobacterium avium subsp. paratuberculosis (MAP) is an obligate intracellular pathogen that infects many ruminant species. The acquisition of foreign genes via horizontal gene transfer has been postulated to contribute to its pathogenesis, as these genetic elements are absent from its putative ancestor, M. avium subsp. hominissuis (MAH), an environmental organism with lesser pathogenicity. In this study, high-throughput sequencing of MAP transposon libraries were analyzed to qualitatively and quantitatively determine the contribution of individual genes to bacterial survival during infection. RESULTS: Out of 52384 TA dinucleotides present in the MAP K-10 genome, 12607 had a MycoMarT7 transposon in the input pool, interrupting 2443 of the 4350 genes in the MAP genome (56%). Of 96 genes situated in MAP-specific genomic islands, 82 were disrupted in the input pool, indicating that MAP-specific genomic regions are dispensable for in vitro growth (odds ratio = 0.21). Following 5 independent in vivo infections with this pool of mutants, the correlation between output pools was high for 4 of 5 (R = 0.49 to 0.61) enabling us to define genes whose disruption reproducibly reduced bacterial fitness in vivo. At three different thresholds for reduced fitness in vivo, MAP-specific genes were over-represented in the list of predicted essential genes. We also identified additional genes that were severely depleted after infection, and several of them have orthologues that are essential genes in M. tuberculosis. CONCLUSIONS: This work indicates that the genetic elements required for the in vivo survival of MAP represent a combination of conserved mycobacterial virulence genes and MAP-specific genes acquired via horizontal gene transfer. In addition, the in vitro and in vivo essential genes identified in this study may be further characterized to offer a better understanding of MAP pathogenesis, and potentially contribute to the discovery of novel therapeutic and vaccine targets.
Assuntos
Proteínas de Bactérias/genética , Elementos de DNA Transponíveis , Mycobacterium avium subsp. paratuberculosis/genética , Paratuberculose/microbiologia , Animais , Proteínas de Bactérias/metabolismo , DNA Bacteriano/metabolismo , Regulação Bacteriana da Expressão Gênica , Transferência Genética Horizontal , Genes Essenciais , Aptidão Genética , Sequenciamento de Nucleotídeos em Larga Escala , Camundongos , Camundongos Endogâmicos C57BL , Mycobacterium avium subsp. paratuberculosis/fisiologia , Filogenia , Análise de Sequência de DNARESUMO
Most targeted anticancer therapies fail due to drug resistance evolution. Here we show that tumor evolution can be reproducibly redirected to engineer therapeutic opportunity, regardless of the exact ensemble of pre-existing genetic heterogeneity. We develop a selection gene drive system that is stably introduced into cancer cells and is composed of two genes, or switches, that couple an inducible fitness advantage with a shared fitness cost. Using stochastic models of evolutionary dynamics, we identify the design criteria for selection gene drives. We then build prototypes that harness the selective pressure of multiple approved tyrosine kinase inhibitors and employ therapeutic mechanisms as diverse as prodrug catalysis and immune activity induction. We show that selection gene drives can eradicate diverse forms of genetic resistance in vitro. Finally, we demonstrate that model-informed switch engagement effectively targets pre-existing resistance in mouse models of solid tumors. These results establish selection gene drives as a powerful framework for evolution-guided anticancer therapy.
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Identifying mechanisms of drug action remains a fundamental impediment to the development and effective use of chemotherapeutics. Here we describe an RNA interference (RNAi)-based strategy to characterize small-molecule function in mammalian cells. By examining the response of cells expressing short hairpin RNAs (shRNAs) to a diverse selection of chemotherapeutics, we could generate a functional shRNA signature that was able to accurately group drugs into established biochemical modes of action. This, in turn, provided a diversely sampled reference set for high-resolution prediction of mechanisms of action for poorly characterized small molecules. We could further reduce the predictive shRNA target set to as few as eight genes and, by using a newly derived probability-based nearest-neighbors approach, could extend the predictive power of this shRNA set to characterize additional drug categories. Thus, a focused shRNA phenotypic signature can provide a highly sensitive and tractable approach for characterizing new anticancer drugs.
Assuntos
Neoplasias/genética , Humanos , Neoplasias/patologia , Fenótipo , Interferência de RNARESUMO
The current clinical application of combination chemotherapy is guided by a historically successful set of practices that were developed by basic and clinical researchers 50-60 years ago. Thus, in order to understand how emerging approaches to drug development might aid the creation of new therapeutic combinations, it is critical to understand the defining principles underlying classic combination therapy and the original experimental rationales behind them. One such principle is that the use of combination therapies with independent mechanisms of action can minimize the evolution of drug resistance. Another is that in order to kill sufficient cancer cells to cure a patient, multiple drugs must be delivered at their maximum tolerated dose - a condition that allows for enhanced cancer cell killing with manageable toxicity. In light of these models, we aim to explore recent genomic evidence underlying the mechanisms of resistance to the combination regimens constructed on these principles. Interestingly, we find that emerging genomic evidence contradicts some of the rationales of early practitioners in developing commonly used drug regimens. However, we also find that the addition of recent targeted therapies has yet to change the current principles underlying the construction of anti-cancer combinatorial regimens, nor have they made substantial inroads into the treatment of most cancers. We suggest that emerging systems/network biology approaches have an immense opportunity to impact the rational development of successful drug regimens. Specifically, by examining drug combinations in multivariate ways, next generation combination therapies can be constructed with a clear understanding of how mechanisms of resistance to multi-drug regimens differ from single agent resistance.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Simulação por Computador , Dano ao DNA , Análise Mutacional de DNA , Resistência a Múltiplos Medicamentos/genética , Humanos , Leucemia/genética , Terapia de Alvo Molecular , Transdução de SinaisRESUMO
Drug repositioning seeks to leverage existing clinical knowledge to identify alternative clinical settings for approved drugs. However, repositioning efforts fail to demonstrate improved success rates in late-stage clinical trials. Focusing on 11 approved kinase inhibitors that have been evaluated in 139 repositioning hypotheses, we use data mining to characterize the state of clinical repurposing. Then, using a simple experimental correction with human serum proteins in in vitro pharmacodynamic assays, we develop a measurement of a drug's effective exposure. We show that this metric is remarkably predictive of clinical activity for a panel of five kinase inhibitors across 23 drug variant targets in leukemia. We then validate our model's performance in six other kinase inhibitors for two types of solid tumors: non-small cell lung cancer (NSCLC) and gastrointestinal stromal tumors (GISTs). Our approach presents a straightforward strategy to use existing clinical information and experimental systems to decrease the clinical failure rate in drug repurposing studies.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Leucemia , Neoplasias Pulmonares , Humanos , Reposicionamento de Medicamentos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológicoRESUMO
Background: Adult and pediatric tumors display stark differences in their mutation spectra and chromosome alterations. Here, we attempted to identify common and unique gene dependencies and their associated biomarkers among adult and pediatric tumor isolates using functional genetic lethal screens and computational modeling. Methods: We performed CRISRP-Cas9 lethality screens in two adult glioblastoma (GBM) tumor isolates and five pediatric brain tumor isolates representing atypical teratoid rhabdoid tumors (ATRT), diffuse intrinsic pontine glioma, GBM, and medulloblastoma. We then integrated the screen results with machine learning-based gene-dependency models generated from data from >900 cancer cell lines. Results: We found that >50% of candidate dependencies of 280 identified were shared between adult GBM tumors and individual pediatric tumor isolates. 68% of screen hits were found as nodes in our network models, along with shared and tumor-specific predictors of gene dependencies. We investigated network predictors associated with ADAR, EFR3A, FGFR1 (pediatric-specific), and SMARCC2 (ATRT-specific) gene dependency among our tumor isolates. Conclusions: The results suggest that, despite harboring disparate genomic signatures, adult and pediatric tumor isolates share a preponderance of genetic dependences. Further, combining data from primary brain tumor lethality screens with large cancer cell line datasets produced valuable insights into biomarkers of gene dependency, even for rare cancers. Importance of the Study: Our results demonstrate that large cancer cell lines data sets can be computationally mined to identify known and novel gene dependency relationships in adult and pediatric human brain tumor isolates. Gene dependency networks and lethality screen results represent a key resource for neuro-oncology and cancer research communities. We also highlight some of the challenges and limitations of this approach.
RESUMO
Introduction: Modern targeted cancer therapies are carefully crafted small molecules. These exquisite technologies exhibit an astonishing diversity of observed failure modes (drug resistance mechanisms) in the clinic. This diversity is surprising because back of the envelope calculations and classic modeling results in evolutionary dynamics suggest that the diversity in the modes of clinical drug resistance should be considerably smaller than what is observed. These same calculations suggest that the outgrowth of strong pre-existing genetic resistance mutations within a tumor should be ubiquitous. Yet, clinically relevant drug resistance occurs in the absence of obvious resistance conferring genetic alterations. Quantitatively, understanding the underlying biological mechanisms of failure mode diversity may improve the next generation of targeted anticancer therapies. It also provides insights into how intratumoral heterogeneity might shape interpatient diversity during clinical relapse. Materials and Methods: We employed spatial agent-based models to explore regimes where spatial constraints enable wild type cells (that encounter beneficial microenvironments) to compete against genetically resistant subclones in the presence of therapy. In order to parameterize a model of microenvironmental resistance, BT20 cells were cultured in the presence and absence of fibroblasts from 16 different tissues. The degree of resistance conferred by cancer associated fibroblasts in the tumor microenvironment was quantified by treating mono- and co-cultures with letrozole and then measuring the death rates. Results and Discussion: Our simulations indicate that, even when a mutation is more drug resistant, its outgrowth can be delayed by abundant, low magnitude microenvironmental resistance across large regions of a tumor that lack genetic resistance. These observations hold for different modes of microenvironmental resistance, including juxtacrine signaling, soluble secreted factors, and remodeled ECM. This result helps to explain the remarkable diversity of resistance mechanisms observed in solid tumors, which subverts the presumption that the failure mode that causes the quantitatively fastest growth in the presence of drug should occur most often in the clinic. Conclusion: Our model results demonstrate that spatial effects can interact with low magnitude of resistance microenvironmental effects to successfully compete against genetic resistance that is orders of magnitude larger. Clinical outcomes of solid tumors are intrinsically connected to their spatial structure, and the tractability of spatial agent-based models like the ones presented here enable us to understand this relationship more completely.
RESUMO
A genetic knockout can be lethal to one human cell type while increasing growth rate in another. This context specificity confounds genetic analysis and prevents reproducible genome engineering. Genome-wide CRISPR compendia across most common human cell lines offer the largest opportunity to understand the biology of cell specificity. The prevailing viewpoint, synthetic lethality, occurs when a genetic alteration creates a unique CRISPR dependency. Here, we use machine learning for an unbiased investigation of cell type specificity. Quantifying model accuracy, we find that most cell type specific phenotypes are predicted by the function of related genes of wild-type sequence, not synthetic lethal relationships. These models then identify unexpected sets of 100-300 genes where reduced CRISPR measurements can produce genome-scale loss-of-function predictions across >18,000 genes. Thus, it is possible to reduce in vitro CRISPR libraries by orders of magnitude-with some information loss-when we remove redundant genes and not redundant sgRNAs.