Biology subjects

Bui, J. D.

Publications and source records attributed to Bui, J. D..

4 recordsLinked to original sources

bioRxiv2025

Identification of Protein Cargo in Extracellular Vesicles from Macrophages in Progressing and Regressing Tumors

Exosomes are extracellular vesicles (EVs) that carry bioactive molecules from a cell of origin and may alter gene expression of the acceptor cell via binding to cell surface receptors and/or delivering their cargo into the cell. Tumor-associated macrophages (TAMs) can increase or decrease tumor growth through various mechanisms, but the impact of TAM-EVs on tumors has been difficult to elucidate due to challenges in isolating and culturing TAMs for EV purification. In this study, we set out to uncover the protein identities within EVs from TAMs in progressing and regressing tumors, thereby uncovering EV proteins roles in TAM-tumor crosstalk. TAMs were purified from tumors using magnetic bead isolation and cultured for up to 9 days. TAMs from regressing tumors were found to be more M1-like by high expression of major histocompatibility complex (MHC) class II, while TAMs from progressing tumors are more M2-like (low expression of MHC class II). Long-term in vitro culture of TAMs resulted in reduced expression of MHC class II. EVs were harvested from plated TAMs, and EV proteins were identified by mass spectrometry and compared to EV proteins from bone marrow-derived macrophages (BMDMs). Unsupervised hierarchical clustering studies revealed that proteins from TAM-EVs converge based on their in vitro culture duration rather than their tumor origin. Signature proteins in TAM-EVs were characterized, with the identification of galectin-3-binding protein as a signature protein present in EVs from M1-like BMDMs and regressor macrophages. Finally, immunostimulatory pathways were identified in progressor-TAM-EVs through protein-protein interaction network analysis.

immunology↗

bioRxiv2025

Derivation of functional early gestation decidual natural killer cell subtypes from induced pluripotent stem cells

Abnormal decidual natural killer cell (dNK) function is linked to pregnancy complications occurring in both early and late gestation, including recurrent pregnancy loss, preeclampsia, and preterm birth. Exploration of dNK heterogeneity as it relates to function is an active area of research; however, most of this work has focused on early gestation. Using flow cytometric and transcriptomic single-cell definitions of dNK subtypes, we characterized dNK heterogeneity in term dNK within both chorioamniotic membranes and basal plate. We also applied aptamer-based secretome profiling to first trimester and term dNK, and found dNK-specific proteins - VEGF and PLGF - to be reduced at term. We further determined that, compared to first trimester dNK, term dNK have reduced cytoxicity against target cells. Finally, we applied this knowledge to establish a protocol for differentiation of induced pluripotent stem cells (iPSC) into functional dNK. We found that treatment with TGF{beta} enriched for dNK2 subtype, while inducing dNK markers, CD9 and CD103. We evaluated function using cytokine and degranulation assays, aptamer-based secretome profiling, and cytotoxicity assays. We found that iPSC-dNK are functionally most similar to primary dNK. Further, TGF{beta} iPSC-dNK had reduced GM-CSF in response to PMA/I and increased secretion of VEGF and other first trimester-specific proteins - supportive of a shift towards an early gestation, dNK2-dominant, phenotype. We conclude that changes in dNK function across gestation reflect shifts in dNK subtypes that can be reproducibly derived from iPSC, providing a new method for modeling dNK and laying the foundation for cell-based therapeutics for reproductive disease. Significance StatementAlterations in maternal decidual natural killer cells (dNK) are associated with pregnancy complications - from recurrent pregnancy loss to preeclampsia and preterm birth. We found dNK from different regions of the term placenta to be distinct from peripheral blood NK and early gestation dNK, based on gene and surface marker expression, subtype composition, secretome, and cytotoxicity. We report a novel, reproducible protocol to generate dNK resembling the most abundant dNK subtype in early gestation from induced pluripotent stem cells. Our study lays the foundation for in vitro modeling of the maternal-fetal interface and therapeutic development for reproductive disease.

cell biology↗

bioRxiv2024

Evolutionary fingerprint in rodent PD1 confers weakened activity and enhanced tumor immunity compared to human PD1

Mechanistic understanding of the immune checkpoint receptor PD1 is largely based on mouse models, but human and mouse PD1 orthologs exhibit only 59.6% identity in amino acid sequences. Here we show that human PD1 is more inhibitory than mouse PD1 due to stronger interactions with the ligands PDL1 and PDL2 and with the effector phosphatase Shp2. A novel motif highly conserved among PD1 orthologs in vertebrates except in rodents is primarily responsible for the differential Shp2 recruitment. Evolutionary analysis suggested that rodent PD1 orthologs uniquely underwent functional relaxation, particularly during the K-Pg boundary. Humanization of the PD1 intracellular domain disrupted the anti-tumor activity of mouse T cells while increasing the magnitude of anti-PD1 response. Together, our study uncovers species-specific features of the PD1 pathway, with implications to PD1 evolution and differential anti-PD(L)1 responses in mouse models and human patients.

immunology↗

bioRxiv2022

Lymphatic-Preserving Treatment Sequencing with Immune Checkpoint Inhibition Unleashes cDC1-Dependent Antitumor Immunity in HNSCC

Immune checkpoint inhibition (ICI) with anti-CTLA-4 and anti-PD-1 has revolutionized oncology; however, response rates remain limited in most cancer types, highlighting the need for more effective immune oncology (IO) treatment strategies. Paradoxically, head and neck squamous cell carcinoma (HNSCC), which bears a mutational burden and immune infiltrate commensurate with cancers that respond robustly to ICI, has demonstrated no response to anti- CTLA-4 in any setting or to anti-PD-1 for locally-advanced disease. Scrutiny of the landmark clinical trials defining current IO treatments in HNSCC reveals that recruited patients necessarily received regional ablative therapies per standard of care, prompting us to hypothesize that standard therapies, which by design ablate locoregional lymphatics, may compromise host immunity and the tumor response to ICI. To address this, we employed tobacco-signature HNSCC murine models in which we mapped tumor-draining lymphatics and developed models for regional lymphablation with surgery or radiation. Remarkably, we found that lymphablation eliminates the tumor ICI response, significantly worsening overall survival and repolarizing the tumor- and peripheral-immune compartments. Mechanistically, within tumor-draining lymphatics, we observed an upregulation of cDC1 cells and IFN-I signaling, showed that both are necessary for the ICI response and lost with lymphablation. Ultimately, we defined rational IO sequences that mobilize peripheral immunity, achieve optimal tumor responses, confer durable immunity and control regional lymphatic metastasis. In sum, we provide a mechanistic understanding of how standard regional, lymphablative therapies impact the response to ICI, which affords insights that can be applied to define rational, lymphatic-preserving IO treatment sequences for cancer. One Sentence SummaryDespite the promise of immune checkpoint inhibition, therapeutic responses remain limited, raising the possibility that standard of care treatments delivered in concert may compromise the tumor response; here, we provide a mechanistic understanding of how standard oncologic therapies targeting regional lymphatics impact the tumor response to immune-oncology therapy in order to define rational treatment sequences that mobilize systemic antitumor immunity, achieve optimal tumor responses, confer durable antitumor immunity, and control regional metastatic disease. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=107 SRC="FIGDIR/small/478744v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@542d71org.highwire.dtl.DTLVardef@737ad0org.highwire.dtl.DTLVardef@183f519org.highwire.dtl.DTLVardef@505ce2_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗