Biology subjects

Hachey, S. J.

Publications and source records attributed to Hachey, S. J..

8 recordsLinked to original sources

bioRxiv2025

PHLDA2 promotes breast cancer metastasis by co-opting a developmental program for placental vascular remodeling

Identifying drivers of metastasis is essential for developing new treatments for patients with advanced disease. Here, we identify PHLDA2 as a robust driver of breast cancer metastasis. Previous work established PHLDA2 as an imprinted gene expressed by trophoblasts which are critical for vascular remodeling during placental development. We find that hypomethylation of PHLDA2 in breast tumors correlates with increased gene expression, which is associated with metastasis and poor survival in breast cancer patients. RNA-sequencing showed that PHLDA2 overexpression results in upregulation of genes that control invasion, extracellular matrix assembly, and vascular remodeling, consistent with trophoblast functions in placental development. Using an in vitro vascularized microtumor (VMT) system, we find that PHLDA2 functions through SPARC, which promotes metastasis by inducing vascular permeability and enhancing tumor dissemination. These data suggest that increased expression of PHLDA2 through hypomethylation promotes metastasis by ectopic expression of a developmental program for vascular remodeling.

cancer biology↗

bioRxiv2025

ImmuniT Platform for Improved Neoantigen Prediction in Lung Cancer

11.1 IntroductionLung cancer remains the leading cause of cancer-related deaths, with most patients presenting with advanced, treatment-resistant disease. While immunotherapy has improved outcomes for some, most patients fail to mount an effective immune response due to inadequate tumor recognition. Neoantigen-based therapies offer a promising approach to personalized immunotherapy, but current discovery methods can miss immunogenic targets, particularly those with low or heterogeneous expression. To address this, we developed the ImmuniT platform, which enhances neoantigen identification by amplifying patient-specific targets from primary tumor samples, improving prediction accuracy for more precise immunotherapy. 1.2 MethodsPatients with lung cancer were recruited under an IRB-approved protocol, and freshly resected tumor tissue and matched blood samples were collected. Tumors were processed into single-cell suspensions, enriched for EpCAM+ epithelial cells, and treated to enhance neoantigen expression. Peripheral blood and tumor-infiltrating lymphocytes were co-cultured with cancer cells to expand neoantigen-reactive T cells. The nextneopi pipeline integrated tumor mutational burden (TMB), HLA typing, and transcriptomic data to predict immunogenic targets. MHC:epitope complexes were validated via tetramer staining to identify patient-derived, neoantigen-specific T cells. 1.3 ResultsThe ImmuniT platform demonstrated superior neoantigen prediction and T cell activation in vitro compared to conventional methods across five NSCLC patients. In one patient, it identified two neoantigens missed by standard approaches, which were validated based on their ability to stimulate tumor-infiltrating and peripheral blood lymphocytes. Across all tested samples, the platform identified a broader spectrum of immunogenic targets. These findings highlight its potential to enhance neoantigen discovery and improve personalized immunotherapy strategies. 1.4 ConclusionOur findings indicate that the ImmuniT platform improves neoantigen detection in NSCLC by identifying a wider range of tumor-specific antigens, including those over-looked by conventional methods. By expanding the pool of targetable neoantigens, this technology has the potential to enhance T cell activation and optimize immunotherapy. The ImmuniT platform represents a promising advancement towards more effective, personalized treatment strategies for lung cancer patients, particularly those who do not respond to current immunotherapies.

immunology↗

bioRxiv2025

Methods for Processing and Analyzing Images of Vascularized Micro-Organ and Tumor Systems

Our group has developed and validated an advanced microfluidic platform to improve preclinical modeling of healthy and disease states, enabling extended culture and detailed analysis of tissue-engineered miniaturized organ constructs, or "organs-on-chips." Within this system, diverse cell types self-organize into perfused microvascular networks under dynamic flow within tissue chambers, effectively mimicking the structure and function of native tissues. This setup facilitates physiological intravascular delivery of nutrients, immune cells, and therapeutic agents, and creates a realistic microenvironment to study cellular interactions and tissue responses. Known as the vascularized micro-organ (VMO), this adaptable platform can be customized to represent various organ systems or tumors, forming a vascularized micro-tumor (VMT) for cancer studies. The VMO/VMT system closely simulates in vivo nutrient exchange and drug delivery within a 3D microenvironment, establishing a high-fidelity model for drug screening and mechanistic studies in vascular biology, cancer, and organ-specific pathologies. Furthermore, the optical transparency of the device supports high-resolution, real-time imaging of fluorescently labeled cells and molecules within the tissue construct, providing key insights into drug responses, cell interactions, and dynamic processes such as epithelial-mesenchymal transition. To manage the extensive imaging data generated, we created standardized, high-throughput workflows for image analysis. This manuscript presents our image processing and analysis pipeline, utilizing a suite of tools in Fiji/ImageJ to streamline data extraction from the VMO/VMT model, substantially reducing manual processing time. Additionally, we demonstrate how these tools can be adapted for analyzing imaging data from traditional in vitro models and microphysiological systems developed by other researchers.

bioengineering↗

bioRxiv2024

CDC42 Inhibitors Alter Patterns of Vessel Arborization in Skin and Tumors in vivo

Tumors that arise in the epidermis must develop a vascular supply to grow beyond a millimeter in depth. This process requires CDC42 GTPases such as CDC42, RhoJ and RhoQ. Despite this dependence on angiogenesis for growth, melanoma tumors are minimally responsive to current anti-angiogenesis agents, highlighting the need for more effective drugs in this class. Here we integrate antibody infusion, optical tissue clearing, multiphoton imaging, and three-dimensional semi-automated tracing to develop a quantitative approach to measure changes in vascular architecture in skin and skin tumors. This new approach uncovered differences in vessel arborization in the skin of RhoJ KO mice as compared to wild-type mice. Furthermore, novel small molecules that inhibit CDC42 GTPases inhibited both tumor growth and vessel branching within tumors to a similar degree as Braf inhibitors, which are commonly used to treat melanoma. In contrast to Braf inhibitors, however, which only affected tumor vasculature, CDC42 inhibitors affected vascularization in both tumor and normal skin without apparent toxicity to endothelial or stromal cells. These novel CDC42 inhibitors similarly blocked vessel branching in human cell-based micro-physiological models of normal and tumor vessels. RNA sequencing revealed reduced expression of multiple angiogenesis-related genes in drug-treated skin. Taken together, these studies identify a new class of pharmacologic agents that inhibit vessel branching in both normal skin and tumors with potential utility for treating skin cancer and skin diseases characterized by pathologic angiogenesis.

cancer biology↗

bioRxiv2023

Targeting tumor-stromal interactions in triple-negative breast cancer using a human vascularized micro-tumor model

Triple-negative breast cancer (TNBC) is highly aggressive with limited available treatments. Stromal cells in the tumor microenvironment (TME) are crucial in TNBC progression; however, understanding the molecular basis of stromal cell activation and tumor-stromal crosstalk in TNBC is limited. To investigate therapeutic targets in the TNBC stromal niche, we used an advanced human in vitro microphysiological system called the vascularized micro-tumor (VMT). Using single-cell RNA sequencing (scRNA-seq), we revealed that normal breast-tissue stromal cells activate neoplastic signaling pathways in the TNBC TME. By comparing interactions in VMTs with clinical data, we identified therapeutic targets at the tumor-stromal interface with potential clinical significance. Combining treatments targeting Tie2 signaling with paclitaxel resulted in vessel normalization and increased efficacy of paclitaxel in the TNBC VMT. Dual inhibition of Her3 and Akt also demonstrated efficacy against TNBC. These data demonstrate the potential of inducing a favorable TME as a targeted therapeutic approach in TNBC.

cancer biology↗

bioRxiv2022

Engineered vasculature induces functional maturation of pluripotent stem cell-derived islet organoids

Blood vessels play a critical role in pancreatic islet health and function, yet current culture methods to generate islet organoids from human pluripotent stem cells (SC-islets) lack a vascular component. Here, we engineered 3D vascularized SC-islet organoids by assembling SC-islet cells, human primary endothelial cells (ECs) and fibroblasts both in a non-perfused model and a microfluidic device with perfused vessels. Vasculature improved stimulus-dependent Ca2+ influx into SC-{beta}-cells, a hallmark of {beta}-cell function that is blunted in non-vascularized SC-islets. We show that an islet-like basement membrane is formed by vasculature and contributes to the functional improvement of SC-{beta}-cells. Furthermore, cell-cell communication networks based on scRNA-seq data predicted BMP2/4-BMPR2 signaling from ECs to SC-{beta}-cells. Correspondingly, BMP4 augmented the SC-{beta}-cell Ca2+ response and insulin secretion. These vascularized SC-islet models will enable further studies of crosstalk between {beta}-cells and ECs and can serve as in vivo-mimicking platforms for disease modeling and therapeutic testing.

cell biology↗

bioRxiv2021

Structure-based Design of CDC42 Effector Interaction Inhibitors For the Treatment of Cancer

CDC42 family GTPases (RHOJ, RHOQ, CDC42) are upregulated but rarely mutated in cancer and control both the ability of tumor cells to invade surrounding tissues and the ability of endothelial cells to vascularize tumors. Here we use computer-aided drug design to discover a new chemical entity (ARN22089) that targets CDC42 GTPases and blocks CDC42 effector interactions without affecting the binding between closely related GTPases (RAC1, RAS, RAL) and their downstream effectors. Our lead compound has broad activity against a panel of cancer cell lines, inhibits S6 phosphorylation and MAPK activation, activates pro-inflammatory and apoptotic signaling, and blocks tumor growth and angiogenesis in three-dimensional vascularized microtumor models (VMT) in vitro. In addition, ARN22089 has a favorable pharmacokinetic profile and can inhibit the growth of BRAF mutant mouse melanomas and patient-derived xenografts in vivo. Taken together, this work identifies a promising new class of therapeutic agents that influence tumor growth by modulating CDC42 signaling in both the tumor cell and its microenvironment.

molecular biology↗

bioRxiv2020

An In Vitro Vascularized Micro-Tumor Model of Human Colorectal Cancer Recapitulates In Vivo Drug Responses

Around 95% of anti-cancer drugs that show promise during preclinical study fail to gain FDA-approval for clinical use. This failure of the preclinical pipeline highlights the need for improved, physiologically-relevant in vitro models that can better serve as reliable drug-screening tools. The vascularized micro-tumor (VMT) is a novel three-dimensional model system that recapitulates the complex human tumor microenvironment, including perfused vasculature, within a transparent microfluidic device, allowing real-time study of drug responses and tumor-stromal interactions. Here we have validated the VMT platform for the study of colorectal cancer (CRC), the second leading cause of cancer-related deaths, by showing that gene expression, tumor heterogeneity, and treatment response in the VMT more closely model CRC tumor clinicopathology than current standard drug screening modalities, including 2-dimensional (2D) monolayer culture and 3-dimensional (3D) spheroids.

cancer biology↗