Biology subjects

Hollern, D.

Publications and source records attributed to Hollern, D..

3 recordsLinked to original sources

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↗

bioRxiv2021

Synergy of chemotherapy and macrophage depletion leads to T cell memory activation and durable triple negative breast cancer regression

Immunosuppressive elements within the tumor microenvironment such as Tumor Associated Macrophages (TAMs) can present a barrier to successful anti-tumor responses by cytolytic T cells. We employed preclinical syngeneic p53 null mouse models of triple negative breast cancer (TNBC) to develop a treatment regimen that harnessed the immunostimulatory effects of low-dose cyclophosphamide coupled with the pharmacologic inhibition of TAMs using either a small molecule CSF1R inhibitor or an anti-CSF1R antibody. This therapeutic combination was used to successfully treat several highly aggressive TNBC murine mammary tumors and lung metastasis. Using this regimen and single cell RNA sequencing we characterized tumor infiltrating lymphocytes (TILs) including helper T cells and antigen-presenting B cells that were highly enriched in good responders to combination therapy. Using high dimensional imaging techniques, we identified the close spatial localization of B220+ CD86+ activated B cells and CD4+ T cells in tertiary lymphoid structures that were present up to 6 weeks post-treatment in one model that also exhibited long-term tumor regression post-treatment. We also characterized the transcriptional and metabolic heterogeneity of TAMS in these two closely related claudin-low/mesenchymal subtype tumor models with differential treatment responses. A murine TAM signature derived from the T12 model is highly expressed and conserved in human claudin-low breast cancers, and high expression of the T12 signature correlated with reduced overall survival. This T12 tumor TAM signature may help identify human claudin-low breast cancer patients that will benefit from the combination of cyclophosphamide and anti-CSF1R therapy. These studies illustrate the complexity of the tumor immune microenvironment and highlight different immune responses that result from rationale combinations of immunotherapy. SignificanceA treatment regimen that harnessed the immunostimulatory effects of cyclophosphamide coupled with the inhibition of CSF1R was used to successfully treat several highly aggressive claudin-low TNBC murine mammary tumors and lung metastasis.

cancer biology↗