Biology subjects

Andrechek, E. R.

Publications and source records attributed to Andrechek, E. R..

6 recordsLinked to original sources

bioRxiv2024

Unravelling biological processess and EGFR pathway regulation by the receptor-like protein tyrosine phosphatase PTPRH in non-small cell lung cancer

The balance of protein phosphorylation is often disrupted in cancer, with hyperactivity of kinases and inactivation of phosphatases driving cell proliferation and survival pathways. PTPRH, a protein tyrosine phosphatase, is mutated in [~]5% of non-small cell lung cancers (NSCLC). However, how PTPRH contributes to biological processes and tumorigenesis was unknown. We uncovered PTPRHs candidate interactors and associated pathways by applying a proximity-dependent biotinylation assay (BioID) and generating a signature transcriptome in two NSCLC cell lines derived from the primary tumor (NCI-H23) or a metastatic site (NCI-H2023), followed by functional validation. Candidate interactors included signaling molecules and structural proteins linked to integrins and focal adhesions, adherens junctions, migration, and the cytoskeleton, in addition to interactions with the receptor tyrosine kinases EGFR, EPHA2, and ROR2, and the phosphatases PTPN3 and PTPRJ. Considering the importance of EGFR in lung cancers and the role of EPHA2 in regulating cell adhesion, we examined how PTPRH regulates their signaling. Overexpression of PTPRH decreased EGFR phosphorylation at tyrosine 1173. It also reduced phospho-EPHA2, with one of the target tyrosine residues identified as the ligand-dependent Y588. At the cellular level, PTPRH and EPHA2 colocalize, with PTPRH gain inducing morphological alterations, such as increased eccentricity, smaller size and changes in the cytoskeleton organization in NCI-H23 cells. These changes are accompanied by increased FAK Y397 phosphorylation, but reduced cell adhesion to the ECM. Additionally, pathway enrichment analysis revealed downregulation of multiple oncogenic, metabolic, and cell adhesion signaling pathways, with increased levels of PTPRH leading to reduced migration in vitro, suppressed tumor growth and lung colonization and tumor differentiation in vivo. Interestingly, some alterations may be independent of PTPRH catalytic activity and tailored to a cell lines site of origin and genetic background. These results indicate that PTPRH regulates key signaling, structural networks, and tumor behavior with loss facilitating NSCLC progression.

cancer biology↗

bioRxiv2024

Atypical Cell Cycle Regulation Promotes Mammary Stem Cell Expansion and Therapeutic Resistance

BackgroundThe cell cycle of mammary stem cells must be tightly regulated to ensure normal homeostasis of the mammary gland to prevent abnormal proliferation and susceptibility to tumorigenesis. The atypical cell cycle regulator, Spy1 can override cell cycle checkpoints, including those activated by the tumour suppressor p53 which mediates mammary stem cell homeostasis. Spy1 has also been shown to promote expansion of select stem cell populations in other developmental systems. Spy1 protein is elevated during proliferative stages of mammary gland development, is found at higher levels in human breast cancers, and promotes susceptibility to mammary tumourigenesis when combined with loss of p53. We hypothesized that Spy1 cooperates with loss of p53 to increase susceptibility to tumour initiation due to changes in susceptible mammary stem cell populations during development and drives the formation of more aggressive stem like tumours. MethodsUsing a transgenic mouse model driving expression of Spy1 within the mammary gland, mammary development and stemness were assessed. These mice were intercrossed with p53 null mice to study the tumourigenic properties of Spy1 driven p53 null tumours, as well as global changes in signaling via RNA sequencing analysis. ResultsWe show that elevated levels of Spy1 leads to expansion of mammary stem cells, even in the presence of p53, and an increase in mammary tumour formation. Spy1-driven tumours have an increased cancer stem cell population, decreased checkpoint signaling, and demonstrate an increase in therapy resistance. Loss of Spy1 decreases tumor onset and reduces the cancer stem cell population. ConclusionsThis data demonstrates the potential of Spy1 to expand mammary stem cell populations and contribute to the initiation and progression of aggressive, drug resistant breast cancers with increased cancer stem cell populations.

cancer biology↗

bioRxiv2021

Insight into mammary gland development and tumor prevention in a newly developed metastatic mouse model of breast cancer

The development of breast cancer has been observed due to altered regulation of mammary gland developmental processes. Thus, a better understand of the normal mammary gland development can reveal possible mechanism in how normal cells are re-programmed to become malignant cells. E2F1-4 are part of the E2F transcription factor family with varied roles in mammary development. However, little is known about the role of E2F5 in mammary gland development. A combination of scRNAseq and predictive signature tools demonstrate the presence of E2F5 in the mammary gland and showed altered activity during the various phases of mammary gland development and function. Testing the hypothesis that E2F5 regulates mammary function, we generated a mammary-specific E2F5 knockout mouse model, resulting in modest mammary gland development changes. However, after a prolonged latency the E2F5 conditional knockout mice developed highly metastatic mammary tumors with metastases in both the lung and liver. Transplantation of the tumors revealed metastases to lymph nodes that was enriched through serial transplantation. Through whole genome sequencing and RNAseq analysis we identified, and then confirmed in vivo, that Cyclin D1 was dysregulated in E2F5 conditional knockout mammary glands and tumors. Based on these findings, we propose that loss of E2F5 leads altered regulation of Cyclin D1, which facilitates the development of mammary tumors.

cancer biology↗

bioRxiv2021

Elevated phosphorylation of EGFR in NSCLC due to mutations in PTPRH

The role of EGFR in lung cancer is well described with numerous activating mutations that result in phosphorylation and tyrosine kinase inhibitors that target EGFR. While the role of the EGFR kinase in non-small cell lung cancer (NSCLC) is appreciated, control of EGFR signaling pathways through dephosphorylation by phosphatases is not as clear. In recent work we identified mutations in Protein Tyrosine Phosphatase Receptor Type H (Ptprh, also known as SAP-1) as being associated with elevated phosphorylation of EGFR in a mouse model of breast cancer. We have examined a series of tumors from this mouse model, revealing conserved V483M Ptprh mutations within the FVB background, but a series of varied mutations in other backgrounds. Despite the varied Ptprh mutations in other background strains, matched primary and metastatic tumors largely shared mutational profiles. Profiling the downstream events of Ptprh mutant tumors revealed AKT activation, suggesting a key target of PTPRH was EGFR tyrosine 1197. Given the role of EGFR in lung cancer, we explored TCGA data which revealed that a subset of PTPRH mutant tumors shared gene expression profiles with EGFR mutant tumors, but that EGFR mutations and PTPRH mutations were mutually exclusive. Generation of a PTPRH knockout NSCLC cell line resulted in Y1197 phosphorylation of EGFR, and a rescue with expression of wild type PTPRH returned EGFR phosphorylation to parental line values while a rescue with a D986A catalytically dead mutant PTPRH did not, demonstrating that PTPRH targets EGFR. As expected with active EGFR, the knockout of PTPRH was associated with increased growth rate. Moreover, a dose response curve illustrated that two human NSCLC lines that had naturally occurring PTPRH mutations responded to EGFR tyrosine kinase inhibition. Injection of one of the NSCLC human lines into mice resulted in tumors, and Osimertinib treatment resulted in a reduction of tumor volume relative to vehicle controls. Consistent with prior literature from breast cancer, PTPRH mutation resulted in nuclear pEGFR as seen in immunohistochemistry, suggesting that there may also be a role for EGFR as a transcriptional co-factor. Other roles for PTPRH were explored through a receptor tyrosine kinase array, noting elevated phosphorylation of FGFR1. Knockout of PTPRH in NSCLC cell lines resulted in elevated phosphorylated FGFR1 relative to controls, indicating that PTPRH has a number of targets that may be aberrantly activated in NSCLC with mutations in PTPRH. Together these data suggest that mutations in PTPRH in NSCLC may result in clinically actionable alterations using existing therapies.

cancer biology↗

bioRxiv2019

Low E2F2 activity is associated with high genomic instability and PARPi resistance

The E2F family, classically known for a central role in cell cycle, has a number of emerging roles in cancer including angiogenesis, metabolic reprogramming, metastasis and DNA repair. E2F1 specifically has been shown to be a critical mediator of DNA repair; however, little is known about DNA repair and other E2F family members. Here we present an integrative bioinformatic and high throughput drug screening study to define the role of E2F2 in maintaining genomic integrity in breast cancer. We utilized in vitro E2F2 ChIP-chip and over expression data to identify transcriptional targets of E2F2. This data was integrated with gene expression from E2F2 knockout tumors in an MMTV-Neu background. Finally, this data was compared to human datasets to identify conserved roles of E2F2 in human breast cancer through the TCGA breast cancer, Cancer Cell Line Encyclopedia, and CancerRx datasets. Here we have computationally predicted that E2F2 transcriptionally regulates key mediators of DNA repair. Our gene expression data supports this hypothesis and low E2F2 activity is associated with a highly unstable tumor. In human breast cancer E2F2, status was also correlated with a patients response to PARP inhibition therapy. Taken together this manuscript defines a novel role of E2F2 in cancer progression beyond cell cycle and could be therapeutically relevant.\n\nAuthor SummaryThe E2F family of proteins have been known to regulate cell cycle and have recently been shown to have a number of roles in tumor progression. Here we use a combination of computational techniques and high-throughput drug screening data to establish a novel role of E2F2 in maintaining genomic integrity. We have shown that a number of direct and indirect target genes of E2F2 are involved in multiple classical DNA repair pathways. Importantly, this was shown to be unique to E2F2 and not present with other activator E2Fs like E2F1. We have also shown that E2F2 activity is positively correlated with PARP inhibitor sensitivity regardless of BRCA1/2 status. This is important due to the recent approval of PARP inhibitor therapy in the clinic. Based on our work E2F2 activity could serve as a novel biomarker of response and may identify a new cohort of patients which could benefit from PARPi therapy.

cancer biology↗

bioRxiv2019

E2F1 Drives Breast Cancer Metastasis by Regulating the Target Gene FGF13 and Altering Cell Migration

In prior work we demonstrated that loss of E2F transcription factors inhibits metastasis. Here we address the mechanisms for this phenotype and identify the E2F regulated genes that coordinate tumor cell metastasis. Transcriptomic profiling of E2F1 knockout tumors identified a role for E2F1 as a master regulator of a suite of pro-metastatic genes, but also uncovered E2F1 target genes with an unknown role in pulmonary metastasis. High expression of one of these genes, Fgf13, is associated with early human breast cancer metastasis in a clinical dataset. Together these data led to the hypothesis that Fgf13 is critical for breast cancer metastasis, and that upregulation of Fgf13 may partially explain how E2F1 promotes breast cancer metastasis. To test this hypothesis we ablated Fgf13 via CRISPR. Deletion of Fgf13 in a MMTV-PyMT breast cancer cell line reduces the frequency of pulmonary metastasis. In addition, loss of Fgf13 reduced in vitro cell migration, suggesting that Fgf13 may be critical for tumor cells to invade out of and escape the primary tumor. The significance of this work is twofold: we have both uncovered genomic features by which E2F1 regulates metastasis and we have identified new pro-metastatic functions for the E2F1 target gene Fgf13.

cancer biology↗