Biology subjects

Bild, A. H.

Publications and source records attributed to Bild, A. H..

11 recordsLinked to original sources

bioRxiv2024

A novel combination therapy for ER+ breast cancer suppresses drug resistance via an evolutionary double-bind

Chemotherapy remains a commonly used and important treatment option for metastatic breast cancer. A majority of ER+ metastatic breast cancer patients ultimately develop resistance to chemotherapy, resulting in disease progression. We hypothesized that an "evolutionary double-bind", where treatment with one drug improves the response to a different agent, would improve the effectiveness and durability of responses to chemotherapy. This approach exploits vulnerabilities in acquired resistance mechanisms. Evolutionary models can be used in refractory cancer to identify alternative treatment strategies that capitalize on acquired vulnerabilities and resistance traits for improved outcomes. To develop and test these models, ER+ breast cancer cell lineages sensitive and resistant to chemotherapy are grown in spheroids with varied initial population frequencies to measure cross-sensitivity and efficacy of chemotherapy and add-on treatments such as disulfiram combination treatment. Different treatment schedules then assessed the best strategy for reducing the selection of resistant populations. We developed and parameterized a game-theoretic mathematical model from this in vitro experimental data, and used it to predict the existence of a double-bind where selection for resistance to chemotherapy induces sensitivity to disulfiram. The model predicts a dose-dependent re-sensitization (a double-bind) to chemotherapy for monotherapy disulfiram.

cancer biology↗

bioRxiv2022

ONC201/TIC10 enhances durability of mTOR inhibitor everolimus in metastatic ER+ breast cancer

The mTOR inhibitor, everolimus, is an important clinical management component of metastatic ER+ breast cancer. However, most patients develop resistance and progress on therapy, highlighting the need to discover strategies that increase mTOR inhibitor effectiveness. We developed ER+ breast cancer cell lines, sensitive or resistant to everolimus, and discovered that combination treatment of ONC201/TIC10 with everolimus inhibited cell growth in 2D/3D in vitro studies. We confirmed increased therapeutic response in primary patient cells progressing on everolimus, supporting clinical relevance. We show ONC201/TIC10, in metastatic ER+ breast cancer cells, mechanistically involves oxidative phosphorylation inhibition and stress response activation. Transcriptomic analysis in everolimus resistant breast patient tumors and mitochondrial functional assays in resistant cell lines demonstrated increased mitochondrial respiration dependency, contributing to ONC201/TIC10 sensitivity. We propose that ONC201/TIC10 and modulation of mitochondrial function may provide an effective add-on therapy strategy for patients with metastatic ER+ breast cancers resistant to mTOR inhibitors.

cancer biology↗

bioRxiv2022

Phylogenetic inference from single-cell RNA-seq data

Tumors are comprised of subpopulations of cancer cells that harbor distinct genetic profiles and phenotypes that evolve over time and during treatment. By reconstructing the course of cancer evolution, we can understand the acquisition of the malignant properties that drive tumor progression. Unfortunately, recovering the evolutionary relationship of individual cancer cells linked to their phenotypes remains a difficult challenge. To address this issue, we have developed PhylinSic, a method that reconstructs the phylogenetic relationships among cells linked to their gene expression profiles from single cell RNA-sequencing (scRNA-Seq) data, and showed that it was robust to the low read depth, drop-out, and noisiness of scRNA-Seq data. This method called nucleotide bases from scRNA-Seq reads using a probabilistic smoothing approach, and then estimated a phylogenetic tree using a Bayesian modeling algorithm. We evaluated PhylinSic and showed that it identified evolutionary relationships resulting from selective events such as drug selection and metastasis and was sensitive enough to identify subclones from genetic drift. Finally, we applied methods of phylogenetic inference and found that breast tumors resistant to chemotherapies harbored two genetic lineages that independently manifested high predicted activity of K-Ras and {beta}-catenin, potentially acquired by distinct mechanisms through convergent evolution. This suggested that therapeutic strategies may need to target multiple lineages to be durable. Taken together, these results demonstrated that PhylinSic provides a framework to model the evolution and link the genotypes and phenotypes of cells within a tumor or cohort of monophyletic tumors using scRNA-Seq.

bioinformatics↗

bioRxiv2022

Cancer cells communicate with macrophages to prevent T cell activation during development of cell cycle therapy resistance

Cancer cells evolve, acquire resistance to therapy and change their environment. One resistance mechanism involves altering communication with non-malignant cells in the tumor microenvironment (TME). By corrupting the signals for growth and survival, evolving cancer cells can engineer a pro-tumor TME. However, the specific interactions between malignant and non-malignant cells that predispose drug resistance and their changes during treatment remain widely unknown. Here we examine the composition, communication, and phenotypic diversity of tumor-associated cell populations in serial biopsies from early-stage ER+ breast cancers. These patients received either endocrine therapy (letrozole) alone or in combination with a CDK4/6 cell cycle inhibitor ribociclib, and were analyzed using single-cell RNA sequencing (scRNAseq). Our analyses reveal cancer cells from ribociclib resistant tumors stimulate macrophage differentiation towards an immune-suppressive phenotype through upregulation of a broad diversity of cytokines and growth factors. This shift in phenotype leads to reduced macrophage cell IL-15/-18 crosstalk with CD8+ T-cell via IL-2/15RA/18R receptors, resulting in diminished T-cell activation and recruitment. Thus, cancer communication promoting an immune-cold TME predispose tumors to develop CDK4/6 inhibitor resistance, and that the beneficial effects of cell cycle inhibitors through blocking cancer cell proliferation must be balanced against their known inhibitory effect on immune cell division and activation. An optimal treatment strategy will require coupling the prevention of cancer division with activation of an effective cytotoxic T-cell response.

cancer biology↗

bioRxiv2021

Novel temporal and spatial patterns of metastatic colonization from rapid-autopsy tumor biopsies

BackgroundMetastatic breast cancer is a deadly disease with a low 5-year survival rate. Tracking metastatic spread in living patients is difficult, and thus poorly understood. ResultsVia rapid autopsy, we have collected 30 tumor samples over 3 timepoints and across 8 organs from a triple-negative metastatic breast cancer patient. The large number of sites sampled, together with deep whole genome sequencing and advanced computational analysis, allowed us to comprehensively reconstruct the tumors evolution at subclonal resolution. The most unique, previously not reported aspect of the tumors evolution we observed in this patient was the presence of "subclone incubators", i.e. already metastatic sites where substantial tumor evolution occurred before colonization of additional sites and organs by subclones that evolved at the incubator site. Overall, we identified four discrete waves of metastatic expansions, each of which resulted in a number of new, genetically similar metastasis sites that also enriched for particular organs (e.g. abdominal vs bone and brain). The lung played a critical role in facilitating metastatic spread in this patient: the lung was the first site of metastatic escape from the primary breast lesion; subclones at this site were the source of all four subsequent metastatic waves; and multiple sites in the lung acted as subclone incubators. Finally, functional annotation revealed that many known driver or metastasis-promoting tumor mutations in this patient were shared by some, but not all metastatic sites, highlighting the need for more comprehensive surveys of a patients metastases for effective clinical intervention. ConclusionsOur analysis revealed the presence of substantial tumor evolution at metastatic incubator sites, with potentially important clinical implications. Our study demonstrated that sampling of a large number of metastatic sites affords unprecedented detail for studying metastatic evolution.

cancer biology↗

bioRxiv2021

Simplifying the development of portable, scalable, and reproducible workflows

Command-line software plays a critical role in biology research. However, processes for installing and executing software differ widely. The Common Workflow Language (CWL) is a community standard that addresses this problem. Using CWL, tool developers can formally describe a tools inputs, outputs, and other execution details in a manner that fosters use of shared computational methods and reproducibility of complex analyses. CWL documents can include instructions for executing tools inside software containers--isolated, operating-system environments. Accordingly, CWL tools are portable--they can be executed on diverse computers--including personal workstations, high-performance clusters, or the cloud. This portability enables easier adoption of bioinformatics pipelines. CWL supports workflows, which describe dependencies among tools and using outputs from one tool as inputs to others. To date, CWL has been used primarily for batch processing of large datasets, especially in genomics. But it can also be used for analytical steps of a study. This article explains key concepts about CWL and software containers and provides examples for using CWL in biology research. CWL documents are text-based, so they can be created manually, without computer programming. However, ensuring that these documents confirm to the CWL specification may prevent some users from adopting it. To address this gap, we created ToolJig, a Web application that enables researchers to create CWL documents interactively. ToolJig validates information provided by the user to ensure it is complete and valid. After creating a CWL tool or workflow, the user can create "input-object" files, which store values for a particular invocation of a tool or workflow. In addition, ToolJig provides examples of how to execute the tool or workflow via a workflow engine.

bioinformatics↗

bioRxiv2021

Facilitation drives estrogen receptor positive (ER+) breast cancer growth and survival under drug pressure

The interplay of positive and negative interactions between drug sensitive and resistant cells influences the effectiveness of treatment in heterogeneous cancer cell populations. In a study of isogenic estrogen receptor positive (ER+) breast cancer cell lineages sensitive and resistant to ribociclib-induced CDK4/6 inhibition in mono- and co-culture, we find that sensitive cells grow and compete more effectively in the absence of treatment. During treatment with ribociclib, sensitive cells survive and proliferate better when grown together with resistant cells than when grown in monoculture, termed facilitation in ecology. Both liquid chromatography-tandem mass spectrometry (LC-MS/MS) assays and single cell RNA-sequencing (scRNAseq) indicate that resistant cell production of estradiol, a highly active estrogen metabolite, is the mechanism of facilitation. Higher estradiol production by resistant cells drives a shift in sensitive cell phenotype to a resistant cell state in coculture. Adding estradiol in monoculture provides sensitive cells with increased resistance to therapy, and cancels facilitation in coculture. Mathematical modeling quantifies the strength of competition and facilitation and predicts that blocking facilitation has the potential to control both resistant and sensitive cell populations and inhibit the emergence of a refractory population.

systems biology↗

bioRxiv2020

Exploiting collateral sensitivity controls growth of mixed culture of sensitive and resistant cells and decreases selection for resistant cells

BackgroundCDK4/6 inhibitors such as ribociclib are becoming widely used targeted therapies in hormone-receptor-positive (HR+) human epidermal growth factor receptor 2-negative (HER2-) breast cancer. However, cancers can advance due to drug resistance, a problem in which tumor heterogeneity and evolution are key features. MethodsRibociclib-resistant HR+/HER2-CAMA-1 breast cancer cells were generated through long-term ribociclib treatment. Characterization of sensitive and resistant cells were performed using RNA sequencing and whole exome sequencing. Lentiviral labeling with different fluorescent proteins enabled us to track the proliferation of sensitive and resistant cells under different treatments in a heterogeneous, 3D spheroid coculture system using imaging microscopy and flow cytometry. ResultsTranscriptional profiling of sensitive and resistant cells revealed the downregulation of the G2/M checkpoint in the resistant cells. Exploiting this acquired vulnerability; resistant cells exhibited collateral sensitivity for the Wee-1 inhibitor, adavosertib (AZD1775). The combination of ribociclib and adavosertib achieved additional antiproliferative effect exclusively in the cocultures compared to monocultures, while decreasing the selection for resistant cells. ConclusionsOur results suggest that optimal antiproliferative effects in heterogeneous cancers can be achieved via an integrative therapeutic approach targeting sensitive and resistant cancer cell populations within a tumor, respectively.

systems biology↗

bioRxiv2020

Topsentinol L Trisulfate, a new Marine Natural Product, that Targets Basal-like and Claudin-low Breast Cancers

BackgroundBreast cancer is a heterogeneous disease. Genomic studies have revealed five different intrinsic subtypes - Luminal A, Luminal B, HER2-enriched, Claudin-low, and Basal-like. Patients diagnosed with Basal-like or Claudin-low breast cancer (BL-CL) suffer from poor prognosis and limited treatment options. Hence, there is an urgent need to identify a new therapeutic lead that can benefit patients with BL-CL breast cancer. MethodsWe used a step-wise screening approach to screen 2778 HP20 fractions from our Marine Invertebrate Compound Library (MICL) to identify compounds that specifically target BL-CL breast cancer. We also performed biochemical investigations to study the effect of the compound in the signaling pathway. Finally, we generated a drug response gene-expression signature and projected it against a human tumor panel of 12 different cancer types to identify other cancer types sensitive to the compound. ResultsWe identified a previously unreported trisulfated sterol, topsentinol L trisulfate (TLT) that exhibits increased efficacy against BL-CL relative to Luminal/HER2+ breast cancer. Biochemical investigation of the effects of TLT on BL-CL revealed its ability to inhibit activation of AMPK and CHK1 and promote activation of p38. The importance of targeting AMPK and CHK1 in BL-CL was validated by treating a panel of breast cancer cell lines with known small molecule inhibitors of AMPK (Dorsomorphin) and CHK1 (Ly2603618) and recording the increased effectiveness against BL-CL compared to Luminal/HER2+ breast cancer. The TLT sensitivity gene-expression signature identified breast and bladder cancer as the most sensitive to TLT while glioblastoma multiforme as least sensitive. ConclusionsOur study identified TLT, a previously uncharacterized trisulfated sterol, as a potential therapeutic selective against BL-CL. Our results also showed that inhibition of AMPK and/or CHK1 might be an effective therapy in BL-CL.

cancer biology↗

bioRxiv2020

Circulating immune cell phenotype dynamics reflect the strength of tumor-immune cell interactions in patients during immunotherapy

The extent that immune cell phenotypes in the peripheral blood reflect within-tumor immune activity prior to and early in cancer therapy is unclear. To address this question, we studied the population dynamics of tumor and immune cells, and immune phenotypic changes, using clinical tumor and immune cell measurements and single cell genomic analyses. These samples were serially obtained from a cohort of advanced gastrointestinal cancer patients enrolled on a trial with chemotherapy and immunotherapy. Using an ecological population model, fitted to clinical tumor burden and immune cell abundance data from each patient, we find evidence of a strong tumor-circulating immune cell interaction in responder patients, but not those patients that progress on treatment. Upon initiation of therapy, immune cell abundance increased rapidly in responsive patients, and once the peak level is reached, tumor burden decreases, similar to models of predator-prey interactions; these dynamic patterns were absent in non-responder patients. To interrogate phenotype dynamics of circulating immune cells, we performed single cell RNA sequencing at serial time points during treatment. These data show that peripheral immune cell phenotypes were linked to the increased strength of patients tumor-immune cell interaction, including increased cytotoxic differentiation and strong activation of interferon signaling in peripheral T-cells in responder patients. Joint modeling of clinical and genomic data highlights the interactions between tumor and immune cell populations and reveals how variation in patient responsiveness can be explained by differences in peripheral immune cell signaling and differentiation soon after the initiation of immunotherapy. One sentence summaryPeripheral immune cell differentiation and signaling, upon initiation of immunotherapy, reflects tumor attacking ability and patient response. Significance statementThe evolution of peripheral immune cell abundance and signaling over time, as well as how these immune cells interact with the tumor, may impact a cancer patients response to therapy. By developing an ecological population model, we provide evidence of a dynamic predator-prey like relationship between circulating immune cell abundance and tumor size in patients that respond to immunotherapy. This relationship is not found either in patients that are non-responsive to immunotherapy or during chemotherapy. Single cell RNA-sequencing (scRNAseq) of serial peripheral blood samples from patients show that the strength of tumor-immune cell interactions is reflected in T-cells interferon activation and differentiation early in treatment. Thus, circulating immune cell dynamics reflect a tumors response to immunotherapy.

cancer biology↗

bioRxiv2020

A 'one-two punch' therapy strategy to target chemoresistance in ER+ breast cancer

BackgroundCancer cell phenotypes evolve over the course of a tumors treatment. The phenotypes that emerge and disappear over time will be specific to each drug regimen and type of cancer. Chemotherapy remains one of the most common and effective treatments for metastatic breast cancer patients; however, resistance to chemotherapy inevitably emerges. Cancer chemotherapy treatment regimens are not designed to target emerging chemo-resistance, despite its clear importance in progressive cancer. This study focuses on finding sequential treatment strategies that target acquired chemo-resistant states and optimize response to chemotherapy. MethodsIn this study, we used heterogeneous tumor samples from patients to identify subclones resistant to chemotherapy. Using flow cytometry for stem cell markers and DNA sequencing to define subclonal population changes, we measured the enrichment of cancer stem cell-like (CSL) phenotypes in subclones that survive chemotherapy. We then analyzed breast cancer patient tumor organoids and cell line acquisition of CSL traits following chemotherapy, as well as the ability of different drugs to reverse acquired resistance, using flow cytometry, mammosphere assays, and single cell RNA-sequencing analysis. ResultsWe show that in progressive estrogen receptor positive (ER+) metastatic breast cancer patients, resistant tumor subclones that emerge following chemotherapy have increased CSL abundance. Further, in vitro organoid growth of ER+ patient cancer cells also shows that chemotherapy treatment leads to increased abundance of ALDH+/CD44+ CSL cells. Chemotherapy induced CSL abundance is blocked by treatment with a pan-HDAC inhibitor, belinostat. Further, belinostat treatment diminished both mammosphere formation and size following chemotherapy, also indicating a decrease in progenitor CSL traits. HDAC inhibitors specific to class IIa (HDAC4, HDAC5) and IIb (HDAC6) were shown to primarily reverse the chemo-resistant CSL state. Single-cell RNA sequencing analysis with patient samples showed that HDAC targets and MYC signaling were promoted by chemotherapy and inhibited upon HDAC inhibitor treatment. ConclusionThese findings indicate that HDAC inhibition can block chemotherapy-induced drug resistant phenotypes with one-two punch strategy in refractory breast cancer cells.

cancer biology↗