Biology subjects

Ting, D. T.

Publications and source records attributed to Ting, D. T..

13 recordsLinked to original sources

bioRxiv2025

The Pdgfd-Pdgfrb axis orchestrates tumor-nerve crosstalk in pancreatic cancer

Nerves are an integral component of the tumor microenvironment, contributing to cancer progression, metastasis, morbidity, and mortality. In pancreatic ductal adenocarcinoma (PDAC), worse clinical outcomes are associated with perineural invasion (PNI), a process by which cancer cells surround and invade nerves. Here, we employed whole-transcriptome and single-cell spatial transcriptomics to identify candidate tumor-nerve interactions that promote PNI. We discovered that Pdgfd signaling promotes key features of nerve invasion. Mechanistically, Pdgfd stimulated cancer cell invasiveness, neurite outgrowth, and direct physical engagement with glia. Pharmacological blockade of this axis reduced each of these processes in vitro as well as PNI in vivo. Thus, Pdgfd-Pdgfrb signaling mediates PNI by coordinating multifaceted cancer-neuron-glia interactions and represents a promising therapeutic strategy aimed at disrupting harmful cancer-nerve crosstalk.

cancer biology↗

bioRxiv2025

Epitope Expression Persists in Circulating Tumor Cells as Breast Cancers Acquire Resistance to Antibody Drug Conjugates

Antibody-drug conjugates (ADCs) target surface proteins on cancer cells, leading to internalization and delivery of a drug payload, thereby enhancing selectivity and minimizing toxicity. ADCs against TROP2 (Sacituzumab govitecan) or HER2 (T-DXd) have demonstrated efficacy in metastatic breast cancer, yet paradoxically, outside of HER2-amplified breast cancers, expression levels of these breast cancer-enriched epitopes in tumor biopsies have not been strongly correlated with clinical response. We undertook serial quantitative imaging of circulating tumor cells (CTCs) in a prospective cohort of 35 patients treated with either of these ADCs. At the single-cell level, expression of TROP2 and HER2 within individual patients is highly heterogeneous in both CTCs and paired tumor biopsies. Measurement of these epitopes on CTCs immediately prior to ADC therapy does not predict depth of clinical response. However, absence of CTCs or >80% reduction in CTC numbers after three weeks of treatment (CTCLow) predicts durable response, compared with CTCHigh cases (TROP2: HR 5.15, P = 0.012; HER2: HR 6.01, P<0.001). Targeted epitopes are not commonly downregulated on CTCs at the time of acquired clinical resistance, and switching between TROP2- and HER2-targeting ADCs sharing similar payloads infrequently leads to second-line response. Thus, while CTC burden is correlated with response to these ADCs, the level of TROP2 or HER2 expression is poorly predictive. These findings point to sensitivity to the drug payload as a potential driver of clinical response to currently approved ADCs in breast cancer. SIGNIFICANCE STATEMENTThe clinical efficacy of ADCs may depend both on differential targeting of cancer cells, using antibodies to tumor-enriched epitopes, and on the cleavage and release of drug payloads to which cancer cells are sensitive. ADCs currently used to treat breast cancer target either of two epitopes, TROP2 or HER2, but they share chemically related payloads. While CTC numbers track with response, we find that epitope expression is not strongly predictive. The limited success of sequentially switching between TROP2- or HER2-targeting ADCs as second-line treatment, following progression on a first-line ADC, highlights the need to incorporate non-cross-resistant drug payloads on ADCs to overcome such acquired resistance.

cancer biology↗

bioRxiv2025

Intraductal Papillary Mucinous Neoplasm Cellular Plasticity linked with Repeat Element Dysregulation

BackgroundIntraductal papillary mucinous neoplasms (IPMNs) are preinvasive pancreatic lesions with a spectrum of histologic phenotypes and variable risk in progressing to invasive cancer. Aberrant repetitive element expression has been shown to be functionally linked to cell state changes in pancreatic cancer. ObjectiveThis study utilized spatial transcriptomics with customized repeat element probes to better understand the relationship of histologic subtypes, repeat element dysregulation, and molecular profiles of different cell populations in IPMN. DesignA total of 52 lesions from 18 patients with resected IPMNs of different histologies and degrees of dysplasia were analyzed with whole transcriptome spatial analysis (GeoMx). Of these, 50 lesions from 17 patients were also processed for single-cell spatial molecular imaging (CosMx). Repeat element probes for LINE1, HSATII, HERVK, and HERVH were used for GeoMx and CosMx. ResultsPancreaticobiliary-type IPMN was enriched for basal-like epithelium and infiltration of Treg cells. Intestinal-type IPMN was enriched for classical epithelium and macrophage infiltrates. Gastric-type IPMN was found to have equal basal-like and classical epithelium with a diverse immune infiltrate. Repeat RNAs were expressed at high levels across IPMN phenotypes and enriched in high-grade dysplasia. Single-cell transcriptional trajectory analysis revealed a phylogeny starting from gastric toward intestinal and pancreaticobiliary branches associated with higher-grade dysplasia and repeat RNA expression. ConclusionSpatial transcriptomics of IPMN identified a molecular continuum between histological subtypes supporting a common gastric-type origin that transitions to intestinal and pancreatobiliary phenotypes. This cell state plasticity is linked with repeat element expression that can be a potential biomarker for IPMN progression. What is already known on this topicO_LIMolecular characterization of IPMN subtypes using regional spatial transcriptomics has described the differences between histologies C_LIO_LIRepeat element expression is associated with cell state changes in pancreatic ductal neoplasm C_LI What this study addsO_LISingle cell spatial molecular imaging of IPMN subtypes reveals a cellular and molecular continuum starting from gastric histology with distinct branches to Intestinal and pancreatobiliary subtypes C_LIO_LIRepeat element expression is associated with higher grade IPMN histology C_LIO_LIRepeat element expression is elevated in the histologic and molecular continuum from gastric to intestinal and pancreatobiliary-type IPMN C_LI How this study might affect research, practice, or policyO_LIThese results support a common gastric histology origin of IPMN subtypes with different molecular trajectories towards higher grade disease C_LIO_LIOur findings suggest that repeat RNA expression can be used in conjunction with other transcriptional markers of cell state as a biomarker for IPMN progression C_LI

cancer biology↗

bioRxiv2024

A TROP2/Claudin Program Mediates Immune Exclusion to Impede Checkpoint Blockade in Breast Cancer

BackgroundImmune exclusion inhibits anti-tumor immunity and response to immunotherapy, but its mechanisms remain poorly defined. In triple-negative breast cancer (TNBC), an aggressive and generally immune-rich subtype, an immune-cold microenvironment predicts poor prognosis due to a limited response to chemotherapy and immune checkpoint inhibitors. This study aimed to identify mechanisms regulating immune infiltration in TNBC. MethodsWe performed spatial transcriptomic analysis comparing immune-enriched versus immune-cold treatment-naive TNBCs. Functional analyses, including loss-of-function and reconstitution experiments, were conducted to investigate the role of Trophoblast Cell-Surface Antigen 2 (TROP2), a key target of anti-cancer Antibody Drug Conjugates (ADCs), in promoting TNBC progression. A humanized TROP2 syngeneic TNBC model was used to assess the effects of TROP2-targeting in combination with anti-PD1 therapy. Additionally, data from patients treated with immune checkpoint blockade were used to test hypotheses from the preclinical findings. ResultsWe reveal that TROP2 controls barrier-mediated immune exclusion in TNBC through Claudin 7 association and tight junction regulation. TROP2 expression is inversely correlated with T cell infiltration and predicts poor outcomes in TNBC. We demonstrate that TROP2 is sufficient to drive tumor progression in vivo in a CD8 T cell-dependent manner, while its loss deregulates expression and localization of multiple tight junction proteins, enabling T cell infiltration. We show that TROP2 targeting via hRS7, the antibody component of the ADC Sacituzumab govitecan (SG), enhances the anti-PD1 response and improves T cell accessibility and effector function. Correspondingly, TROP2 expression is highly associated with lack of response to anti-PD1 therapy in human breast cancer. ConclusionsThis study defines a new mechanism of barrier-mediated immune exclusion in cancer controlled by TROP2-dependent tight junctions. This mechanism drives tumor progression but can be targeted via TROP2-directed therapy to activate anti-tumor immunity and enhance immunotherapy response. What is already known on this topicBarrier-mediated immune exclusion is emerging as a mechanism of immune evasion in cancer such as triple-negative breast cancer (TNBC), but its molecular underpinnings remain poorly understood. TROP2 is a surface glycoprotein targeted by antibody-drug conjugates such as Sacituzumab govitecan, yet its functional role beyond drug delivery has been unclear. What this study addsThis study identifies TROP2 as a key regulator of tight junction-mediated immune exclusion in TNBC, independent of its intracellular signaling function. TROP2 promotes an immune-cold tumor microenvironment by enforcing mechanical barriers that limit T cell infiltration. How this study might affect research, practice or policyThese findings establish TROP2 as a functional driver of immune evasion and provide a mechanistic rationale for combining TROP2-targeting therapies with immune checkpoint inhibitors to overcome resistance in immune-excluded tumors.

cancer biology↗

bioRxiv2024

Tumor cell-based liquid biopsy using high-throughput microfluidic enrichment of entire leukapheresis product

Circulating Tumor Cells (CTCs), interrogated by sampling blood from patients with cancer, contain multiple analytes, including intact RNA, high molecular weight DNA, proteins, and metabolic markers. However, the clinical utility of tumor cell-based liquid biopsy has been limited since CTCs are very rare, and current technologies cannot process the blood volumes required to isolate a sufficient number of tumor cells for in-depth assays. We previously described a high-throughput microfluidic prototype utilizing high-flow channels and amplification of cell sorting forces through magnetic lenses. Here, we apply this technology to analyze patient-derived leukapheresis products, interrogating a mean blood volume of 5.83 liters from patients with metastatic cancer, with a median of 2,799 CTCs purified per patient. Isolation of many CTCs from individual patients enables characterization of their morphological and molecular heterogeneity, including cell and nuclear size and RNA expression. It also allows robust detection of gene copy number variation, a definitive cancer marker with potential diagnostic applications. High-volume microfluidic enrichment of CTCs constitutes a new dimension in liquid biopsies.

bioengineering↗

bioRxiv2024

CDKN1B (p27kip1) enhances drug tolerant persister CTCs by restricting polyploidy following mitotic inhibitors.

The mitotic inhibitor docetaxel (DTX) is often used to treat endocrine-refractory metastatic breast cancer, but initial responses are mitigated as patients eventually have disease progression. Using a cohort of ex vivo cultures of circulating tumor cells (CTCs) from patients with heavily pretreated breast cancer (n=18), we find two distinct patterns of DTX susceptibility, independent of clinical treatment history. In CTCs cultured from some patients, treatment with a single dose of DTX results in complete cell killing, associated with accumulation of non-viable polyploid ([≥]8N) cells arising from endomitosis. In others, a transient viable drug-tolerant persister (DTP) population emerges, ultimately enabling renewed proliferation of CTCs with preserved parental cell ploidy and DTX sensitivity. In these CTC cultures, efficient cell cycle exit generates a [≤]4N drug-tolerant state dependent on CDKN1B (p27Kip1). Exposure to DTX triggers stabilization of CDKN1B through AKT-mediated phosphorylation at serine 10. Suppression of CDKN1B reduces the number of persister CTCs, increases [≥]8N mitotic cells and abrogates regrowth after DTX exposure. Thus, CDKN1B-mediated suppression of endomitosis contributes to a reversible persister state following mitotic inhibitors in patient-derived treatment refractory breast cancer cells. Summary in bulletsO_LITransient DTX tolerant persister cells emerge in some patient-derived cultured CTCs. C_LIO_LIDTX-tolerant persisters restrict endoreduplication and polyploidy through CDKN1 (p27kip1). C_LIO_LIDTX exposure induces CDKN1B stabilization through AKT mediated phosphorylation at serine 10. C_LIO_LISuppression of polyploidy underlies a drug tolerant persister state specific to mitotic inhibitors. C_LI

cancer biology↗

bioRxiv2023

Dysregulated Repeat Element Viral-like Immune Response in Hepatocellular Carcinoma

PurposeHepatocellular carcinoma (HCC) is a lethal malignancy driven by complex interactions between cancer cells, immune cells, and additional stromal cells in the tumor microenvironment (TME). The LINE1 retrotransposon is a ubiquitous repeat RNA whose de-repression leads to significant cancer cell-intrinsic and TME changes that promote aggressive tumor characteristics. We leveraged single cell spatial transcriptomic profiling to characterize the relationship between LINE1 and differences in the heterogeneous HCC TME. Experimental DesignWe applied our profiling methodology to a cohort of 23 tissue specimens collected from patients who had undergone liver resection or transplantation and validated it in a partially-overlapping similar cohort of 39 specimens using RNA in-situ hybridization (RNA-ISH). ResultsWe found that LINE1-high tumors and LINE1-high single HCC cells exhibited a de-differentiated, stem-like, and inflammatory phenotype. Furthermore, within individual tumors, LINE1 high cancer cells associated spatially with one another and excluded the larger, organized immune cell conglomerates seen in LINE1 low tumors. Finally, we found that LINE1 RNA expression correlated with worse overall survival in the larger expanded retrospective cohort. ConclusionsOur study is the first to show a clearly disorganized immune TME in HCC driven by LINE1 expression, and this observation correlated with poor survival for patients whose tumors expressed large amounts of the LINE1 repeat RNA. These results provide further evidence of how effective anti-tumor immune responses contribute to cures after definitive surgery and may lead to novel biomarkers or drug targets in HCC. TRANSLATIONAL RELEVANCEThe viral-like LINE1 retrotransposon is known to influence tumor cell state and the immune response in a variety of cancer. Here, we have used single cell spatial transcriptomic profiling to resolve repeat and coding gene RNA expression in a cohort of hepatocellular carcinoma (HCC) patients. LINE1 RNA expression in HCC tumor cells was correlated with an undifferentiated stem-like cancer state and a disorganized, sparse immune infiltrate. Using in situ hybridization on an expanded validation cohort, we noted significantly worsened survival in the LINE1 high group. Altogether, LINE1 repeat RNA is a tumor intrinsic biomarker of more aggressive features that can be used for risk stratification and a potential biomarker for response to immunotherapies that merits further investigation.

cancer biology↗

bioRxiv2023

Therapy-associated remodeling of pancreatic cancer revealed by single-cell spatial transcriptomics and optimal transport analysis

In combination with cell intrinsic properties, interactions in the tumor microenvironment modulate therapeutic response. We leveraged high-plex single-cell spatial transcriptomics to dissect the remodeling of multicellular neighborhoods and cell-cell interactions in human pancreatic cancer associated with specific malignant subtypes and neoadjuvant chemotherapy/radiotherapy. We developed Spatially Constrained Optimal Transport Interaction Analysis (SCOTIA), an optimal transport model with a cost function that includes both spatial distance and ligand-receptor gene expression. Our results uncovered a marked change in ligand-receptor interactions between cancer-associated fibroblasts and malignant cells in response to treatment, which was supported by orthogonal datasets, including an ex vivo tumoroid co-culture system. Overall, this study demonstrates that characterization of the tumor microenvironment using high-plex single-cell spatial transcriptomics allows for identification of molecular interactions that may play a role in the emergence of chemoresistance and establishes a translational spatial biology paradigm that can be broadly applied to other malignancies, diseases, and treatments.

cancer biology↗

bioRxiv2023

Cancer cells co-evolve with retrotransposons to mitigate viral mimicry

Overexpression of repetitive elements is an emerging hallmark of human cancers1. Diverse repeats can mimic viruses by replicating within the cancer genome through retrotransposition, or presenting pathogen-associated molecular patterns (PAMPs) to the pattern recognition receptors (PRRs) of the innate immune system2-5. Yet, how specific repeats affect tumor evolution and shape the tumor immune microenvironment (TME) in a pro- or anti-tumorigenic manner remains poorly defined. Here, we integrate whole genome and total transcriptome data from a unique autopsy cohort of multiregional samples collected in pancreatic ductal adenocarcinoma (PDAC) patients, into a comprehensive evolutionary analysis. We find that more recently evolved Short Interspersed Nuclear Elements (SINE), a family of retrotransposable repeats, are more likely to form immunostimulatory double-strand RNAs (dsRNAs). Consequently, younger SINEs are strongly co-regulated with RIG-I like receptor associated type-I interferon genes but anti-correlated with pro-tumorigenic macrophage infiltration. We discover that immunostimulatory SINE expression in tumors is regulated by either Long Interspersed Nuclear Elements 1 (LINE1/L1) mobility or ADAR1 activity in a TP53 mutation dependent manner. Moreover, L1 retrotransposition activity tracks with tumor evolution and is associated with TP53 mutation status. Altogether, our results suggest pancreatic tumors actively evolve to modulate immunogenic SINE stress and induce pro-tumorigenic inflammation. Our integrative, evolutionary analysis therefore illustrates, for the first time, how dark matter genomic repeats enable tumors to co-evolve with the TME by actively regulating viral mimicry to their selective advantage.

cancer biology↗

bioRxiv2022

Inflammatory Signaling and Fibroblast-Cancer Cell Interactions Transfer from a Harmonized Human Single-cell RNA Sequencing Atlas of Pancreatic Ductal Adenocarcinoma to Organoid Co-Culture

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy characterized by a heterogeneous tumor microenvironment (TME) that is enriched with cancer associated fibroblasts (CAFs)1. Cell-cell interactions involving these CAFs promote an immunosuppressive phenotype with altered inflammatory gene expression. While single-cell transcriptomics provides a tool to dissect the complex intercellular pathways that regulate cancer-associated inflammation in human tumors, complementary experimental systems for mechanistic validation remain limited. This study integrated single-cell data from human tumors and novel organoid co-cultures to study the PDAC TME. We derived a comprehensive atlas of PDAC gene expression from six published human single-cell RNA sequencing (scRNA-seq) datasets2-7 to characterize intercellular signaling pathways between epithelial tumor cells and CAFs that regulate the inflammatory TME. Analysis of the epithelial cell compartment identified global gene expression pathways that modulate inflammatory signaling and are correlated with CAF composition. We then generated patient-derived organoid-CAF co-cultures to serve as a biological model of the cellular interactions learned from human tissue in the atlas. Transfer learning analysis to additional scRNA-seq data of this co-culture system and mechanistic experiments confirmed the epithelial response to fibroblast signaling. This bidirectional approach of complementary computational and in vitro applications provides a framework for future studies identifying important mechanisms of intercellular interactions in PDAC.

cancer biology↗

bioRxiv2021

Single cell proteomics of tumor compartments identifies differential kinase activities defining sensitivity to mTOR-PI3-kinase inhibition

Cancer therapy often results in heterogeneous responses in different metastatic lesions in the same patient. Inter- and intra-tumor heterogeneity in proteomic signaling within the various tumor compartments and its impact on therapy are not well characterized due to the limited sensitivity of single cell proteomic approaches. To overcome this barrier, we applied single cell mass cytometry with a customized 29-antibody panel [against cell states, receptor tyrosine kinases (RTK) and phosphoinositide 3-kinase/mammalian target of rapamycin (PI3K/mTOR)-, mitogen-activated protein kinase (MAPK)-, and cytokine-signaling] to PTEN-deleted orthotopic prostate cancer xenograft models to measure the evolution of kinase activities in different tumor compartments during metastasis and upon drug treatment. Compared with primary tumors and circulating tumor cells (CTCs), bone metastases but not lung and liver metastases exhibited elevated PI3K/mTOR signaling and RTKs including c-Met protein, which, when suppressed, impaired tumor growth in the bone. Intra-tumoral heterogeneity within tumor compartments also arises from highly proliferative EpCAMhigh epithelial cells with increased PI3K and mTOR kinase activities co-existing with poorly proliferating EpCAMlow mesenchymal populations with reduced kinase activities, findings recapitulated in epithelial and mesenchymal CTC populations in metastatic prostate and breast cancer patients. Increased kinase activity in EpCAMhigh cells rendered them more sensitive to PI3K/mTOR inhibition and drug resistant EpCAMlow populations with reduced kinase activity emerged over time. Taken together, single cell proteomics identified microenvironment- and cell state-dependent activation of kinase networks creating heterogeneity and differential drug sensitivity among and within tumor populations across different sites, defining a new paradigm of drug responses to kinase inhibitors.

cancer biology↗

bioRxiv2020

Single-nucleus and spatial transcriptomics of archival pancreatic cancer reveals multi-compartment reprogramming after neoadjuvant treatment

Pancreatic ductal adenocarcinoma (PDAC) remains a treatment-refractory disease. Characterizing PDAC by mRNA profiling remains particularly challenging. Previously identified bulk expression subtypes were influenced by contaminating stroma and have not yet informed clinical management, whereas single cell RNA-seq (scRNA-seq) of fresh tumors under-represented key cell types. Here, we developed a robust single-nucleus RNA-seq (snRNA-seq) technique for frozen archival PDAC specimens and used it to study both untreated tumors and those that received neoadjuvant chemotherapy and radiotherapy (CRT). Gene expression programs learned across untreated malignant cell and fibroblast profiles uncovered a clinically relevant molecular taxonomy with improved prognostic stratification compared to prior classifications. Moreover, in the increasingly-adopted neoadjuvant treatment context, there was a depletion of classical-like phenotypes in malignant cells in favor of basal-like phenotypes associated with TNF-NFkB and interferon signaling as well as the presence of novel acinar and neuroendocrine classical-like states, which may be more resilient to cytotoxic treatment. Spatially-resolved transcriptomics revealed an association between malignant cells expressing these basal-like programs and higher immune infiltration with increased lymphocytic content, whereas those exhibiting classical-like programs were linked to sparser macrophage-predominant microniches, perhaps pointing to susceptibility to distinct therapeutic strategies. Our refined molecular taxonomy and spatial resolution can help advance precision oncology in PDAC through informative stratification in clinical trials and insights into differential therapeutic targeting leveraging the immune system.

genomics↗

bioRxiv2017

Global Cancer Transcriptome Quantifies Repeat Element Polarization Between Immunotherapy Responsive and T cell Suppressive Classes

Growing evidence indicates that innate immune pathway activation is critical for responses to immunotherapy and overall cancer prognosis. It has been posited that innate immunity in the tumor microenvironment can be driven by derepression of endogenous repetitive element RNA. The ability to characterize these species can potentially provide novel predictive biomarkers for tumor immune responses and a mechanistic basis for elements of innate activation by tumors. We first compared total RNA and poly(A)-capture protocols applied to tumor RNA-sequencing to detect non-RNA coding transcriptomes in the TCGA. While the poly(A) protocol efficiently detects coding, most non-coding genes, and much of the LINE/SINE/ERV repeat repertoire, we found that it fails to capture overall repeat expression and co-expression. The probing of total RNA expression reveals distinct repetitive co-expression subgroups. Secondly, we found that total repeat element expression delivers the most dynamic changes in samples, which may serve as more robust biomarkers of clinical outcomes. Finally, we show that while expression of ERVs, but not other immunostimulatory repeats such as HSATII, is associated with response to immunotherapy in a cohort of patients with urothelial cancer treated with anti-PD-L1 therapy, global repeat derepression strongly correlates with an immunosuppressive phenotype in the microenvironment of colorectal and pancreatic tumors. We validate in situ in human primary tumors, associating the immunosuppressive phenotype with HSATII expression. In conclusion, we demonstrate the importance of analyzing repetitive element RNAs as potential biomarkers of response to immunotherapy and the need to better characterize these features in next generation sequencing protocols.

cancer biology↗