Mesenchymal Lineage Heterogeneity Underlies Non-Redundant Functions of Pancreatic Cancer-Associated Fibroblasts

Cancer-associated fibroblast (CAF) heterogeneity is increasingly appreciated, but the origins and functions of distinct CAF subtypes remain poorly understood. The abundant and transcriptionally diverse CAF population in pancreatic ductal adenocarcinoma (PDAC) is thought to arise from a common cell of origin, pancreatic stellate cells (PSCs), with diversification resulting from cytokine and growth factor gradients within the tumor microenvironment. Here we analyzed the differentiation and function of PSCs during tumor progression in vivo. Contrary to expectations, we found that PSCs give rise to a numerically minor subset of PDAC CAFs. Targeted ablation of PSC-derived CAFs within their host tissue revealed non-redundant functions for this defined CAF population in shaping the PDAC microenvironment, including production of specific components of the extracellular matrix. Together, these findings link stromal evolution from distinct cells of origin to transcriptional heterogeneity among PDAC CAFs, and demonstrate unique functions for CAFs of a defined cellular origin.

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ITGA5 inhibition in pancreatic stellate cells attenuates desmoplasia and potentiates efficacy of chemotherapy in pancreatic cancer

Science Advances ◽

10.1126/sciadv.aax2770 ◽

2019 ◽

Vol 5 (9) ◽

pp. eaax2770 ◽

Cited By ~ 7

Author(s):

Praneeth R. Kuninty ◽

Ruchi Bansal ◽

Susanna W. L. De Geus ◽

Deby F. Mardhian ◽

Jonas Schnittert ◽

...

Keyword(s):

Therapeutic Potential ◽

Tumor Stroma ◽

Stellate Cells ◽

Pancreatic Stellate Cells ◽

Ductal Adenocarcinoma ◽

Tumor Penetration ◽

Desmoplastic Stroma ◽

Integrin Α5

Abundant desmoplastic stroma is the hallmark for pancreatic ductal adenocarcinoma (PDAC), which not only aggravates the tumor growth but also prevents tumor penetration of chemotherapy, leading to treatment failure. There is an unmet clinical need to develop therapeutic solutions to the tumor penetration problem. In this study, we investigated the therapeutic potential of integrin α5 (ITGA5) receptor in the PDAC stroma. ITGA5 was overexpressed in the tumor stroma from PDAC patient samples, and overexpression was inversely correlated with overall survival. In vitro, knockdown of ITGA5 inhibited differentiation of human pancreatic stellate cells (hPSCs) and reduced desmoplasia in vivo. Our novel peptidomimetic AV3 against ITGA5 inhibited hPSC activation and enhanced the antitumor effect of gemcitabine in a 3D heterospheroid model. In vivo, AV3 showed a strong reduction of desmoplasia, leading to decompression of blood vasculature, enhanced tumor perfusion, and thereby the efficacy of gemcitabine in co-injection and patient-derived xenograft tumor models.

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Depletion of Macrophages Improves Therapeutic Response to Gemcitabine in Murine Pancreas Cancer

Cancers ◽

10.3390/cancers12071978 ◽

2020 ◽

Vol 12 (7) ◽

pp. 1978

Author(s):

Soeren M. Buchholz ◽

Robert G. Goetze ◽

Shiv K. Singh ◽

Christoph Ammer-Herrmenau ◽

Frances M. Richards ◽

...

Keyword(s):

Therapeutic Response ◽

Intraepithelial Neoplasia ◽

Inflammatory Cells ◽

Pancreatic Stellate Cells ◽

Ductal Adenocarcinoma ◽

Scavenging Effect ◽

Liquid Chromatography Tandem Mass ◽

Kpc Mice

Background: The tumor microenvironment (TME) is composed of fibro-inflammatory cells and extracellular matrix (ECM) components. However, the exact contribution of the various TME compartments towards therapeutic response is unknown. Here, we aim to dissect the specific contribution of tumor-associated macrophages (TAMs) towards drug delivery and response in pancreatic ductal adenocarcinoma (PDAC). Methods: The effect of gemcitabine was assessed in human and murine macrophages, human pancreatic stellate cells (hPSCs), and tumor cells (L3.6pl, BxPC3 and KPC) in vitro. The drug metabolism of gemcitabine was analyzed by liquid chromatography–tandem mass spectrometry (LC–MS/MS). Preclinical studies were conducted using KrasG12D;p48-Cre and KrasG12D;p53172H;Pdx-Cre mice to investigate gemcitabine delivery at different stages of tumor progression and upon pharmacological TAM depletion. Results: Gemcitabine accumulation was significantly increased in murine PDAC tissue compared to pancreatic intraepithelial neoplasia (PanIN) lesions and healthy control pancreas tissue. In vitro, macrophages accumulated and rapidly metabolized gemcitabine resulting in a significant drug scavenging effect for gemcitabine. Finally, pharmacological TAM depletion enhanced therapeutic response to gemcitabine in tumor-bearing KPC mice. Conclusion: Macrophages rapidly metabolize gemcitabine in vitro, and pharmacological depletion improves the therapeutic response to gemcitabine in vivo. Our study supports the notion that TAMs might be a promising therapeutic target in PDAC.

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Pancreatic tumor organoids for modeling in vivo drug response and discovering clinically-actionable biomarkers

10.1101/513267 ◽

2019 ◽

Cited By ~ 6

Author(s):

Ling Huang ◽

Bruno Bockorny ◽

Indranil Paul ◽

Dipikaa Akshinthala ◽

Omar Gandarilla ◽

...

Keyword(s):

Cell Biology ◽

Drug Response ◽

Biomarker Discovery ◽

Surface Proteins ◽

Ductal Adenocarcinoma ◽

Core Set ◽

And Function ◽

Pdx Models ◽

Tumor Organoids

AbstractPatient-derived models are transforming translational cancer research. It is not clear if the emergence of patient-derived organoid (PDO) models can extend the utility of the widely used patient-derived xenograft (PDX). In addition, the utility of PDO models for serum biomarker discovery is not known. Here, we demonstrate that PDO models recapitulate the genomics, cell biology, glycomics and drug responses observed in PDX models. Furthermore, we demonstrate the applicability of PDO models for identification of N-glycans that are enriched in the glycome of pancreatic ductal adenocarcinoma (PDAC). Surprisingly, among all the glycans observed in PDX and PDOs, a core set of 57 N-glycans represent 50-94% of the relative abundance of all N-glycans detected, suggesting that only a subset of glycans dominate the cell surface landscape in PDAC. In addition, we outline a tumor organoid-based pipeline to identify surface proteins in extracellular vesicles (EV) from media supernatant of PDO cultures. When combined with the affinity-based validation platform, the EV surface proteins discovered in PDOs are effective in differentiating patients with PDAC from those with benign pancreatitis in the clinic, identifying PDO as powerful discovery platform for serum biomarkers. Thus, PDOs extend the utility of the archival collections of PDX models for translational research and function as a powerful platform for identification of clinically-actionable biomarkers in patients blood.Significance statementTumor organoids extend the utility of PDX models as platforms for investigating drug response, glycosylation changes and function as new platforms for discovering blood-based biomarkers

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Interplay of RFX transcription factors 1, 2 and 3 in motile ciliogenesis

Nucleic Acids Research ◽

10.1093/nar/gkaa625 ◽

2020 ◽

Vol 48 (16) ◽

pp. 9019-9036

Author(s):

Sylvain Lemeille ◽

Marie Paschaki ◽

Dominique Baas ◽

Laurette Morlé ◽

Jean-Luc Duteyrat ◽

...

Keyword(s):

Transcription Factors ◽

Transcriptional Control ◽

Ex Vivo ◽

Ependymal Cells ◽

Clear Understanding ◽

Animal Development ◽

C Elegans ◽

And Function ◽

Redundant Functions

Abstract Cilia assembly is under strict transcriptional control during animal development. In vertebrates, a hierarchy of transcription factors (TFs) are involved in controlling the specification, differentiation and function of multiciliated epithelia. RFX TFs play key functions in the control of ciliogenesis in animals. Whereas only one RFX factor regulates ciliogenesis in C. elegans, several distinct RFX factors have been implicated in this process in vertebrates. However, a clear understanding of the specific and redundant functions of different RFX factors in ciliated cells remains lacking. Using RNA-seq and ChIP-seq approaches we identified genes regulated directly and indirectly by RFX1, RFX2 and RFX3 in mouse ependymal cells. We show that these three TFs have both redundant and specific functions in ependymal cells. Whereas RFX1, RFX2 and RFX3 occupy many shared genomic loci, only RFX2 and RFX3 play a prominent and redundant function in the control of motile ciliogenesis in mice. Our results provide a valuable list of candidate ciliary genes. They also reveal stunning differences between compensatory processes operating in vivo and ex vivo.

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Targeting integrin alpha5 receptor in pancreatic stellate cells to diminish tumor-promoting effects in pancreatic cancer

10.1101/350678 ◽

2018 ◽

Author(s):

Praneeth R. Kuninty ◽

Ruchi Bansal ◽

Sanne W.L. De Geus ◽

Jonas Schnittert ◽

Joop van Baarlen ◽

...

Keyword(s):

Stellate Cells ◽

Pancreatic Stellate Cells ◽

Ductal Adenocarcinoma ◽

Cancer Associated Fibroblasts ◽

Poor Overall Survival ◽

Fibronectin Receptor ◽

Cancer Aggressiveness ◽

Novel Therapeutic

AbstractPancreatic stellate cells (PSCs) are the main precursors of cancer-associated fibroblasts (CAFs) in pancreatic ductal adenocarcinoma (PDAC), known to induce cancer aggressiveness. Integrin alpha5 (ITGA5), a fibronectin receptor, was found to be overexpressed by CAFs in stroma and linked to poor overall survival (log-rank p=0.022, n=137) of patients with PDAC. In vitro, knockdown of ITGA5 in human PSCs (hPSCs) inhibited their adhesion, migration, and proliferation and also inhibited TGF-β-mediated differentiation. In vivo, co-injection of PANC-1 tumor cells and hPSCs (sh-ITGA5) developed tumors with reduced fibrosis and slower growth rate compared to those composed of PANC-1 and hPSC (sh-Ctrl). Furthermore, we developed a ITGA5-antagonizing peptidomimetic (AV3) which inhibited TGFβ-mediated hPSC differentiation by blocking ITGA5/FAK pathway. In vivo, treatment with AV3 intraperitoneally attenuated tumor fibrosis and thereby enhanced the efficacy of gemcitabine in patient-derived xenografts in mice. Altogether, this study reports the therapeutic importance of ITGA5 in PDAC and provides novel therapeutic peptidomimetic to enhance the effect of chemotherapy.

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Reversal of pancreatic desmoplasia by a tumour stroma-targeted nitric oxide nanogel overcomes TRAIL resistance in pancreatic tumours

Gut ◽

10.1136/gutjnl-2021-325180 ◽

2021 ◽

pp. gutjnl-2021-325180

Author(s):

Hsi-Chien Huang ◽

Yun-Chieh Sung ◽

Chung-Pin Li ◽

Dehui Wan ◽

Po-Han Chao ◽

...

Keyword(s):

Nitric Oxide ◽

Phage Display ◽

Tumour Microenvironment ◽

Pancreatic Stellate Cells ◽

Ductal Adenocarcinoma ◽

Tumour Stroma ◽

Desmoplastic Stroma ◽

Trail Resistance

ObjectiveStromal barriers, such as the abundant desmoplastic stroma that is characteristic of pancreatic ductal adenocarcinoma (PDAC), can block the delivery and decrease the tumour-penetrating ability of therapeutics such as tumour necrosis factor-related apoptosis-inducing ligand (TRAIL), which can selectively induce cancer cell apoptosis. This study aimed to develop a TRAIL-based nanotherapy that not only eliminated the extracellular matrix barrier to increase TRAIL delivery into tumours but also blocked antiapoptotic mechanisms to overcome TRAIL resistance in PDAC.DesignNitric oxide (NO) plays a role in preventing tissue desmoplasia and could thus be delivered to disrupt the stromal barrier and improve TRAIL delivery in PDAC. We applied an in vitro–in vivo combinatorial phage display technique to identify novel peptide ligands to target the desmoplastic stroma in both murine and human orthotopic PDAC. We then constructed a stroma-targeted nanogel modified with phage display-identified tumour stroma-targeting peptides to co-deliver NO and TRAIL to PDAC and examined the anticancer effect in three-dimensional spheroid cultures in vitro and in orthotopic PDAC models in vivo.ResultsThe delivery of NO to the PDAC tumour stroma resulted in reprogramming of activated pancreatic stellate cells, alleviation of tumour desmoplasia and downregulation of antiapoptotic BCL-2 protein expression, thereby facilitating tumour penetration by TRAIL and substantially enhancing the antitumour efficacy of TRAIL therapy.ConclusionThe co-delivery of TRAIL and NO by a stroma-targeted nanogel that remodels the fibrotic tumour microenvironment and suppresses tumour growth has the potential to be translated into a safe and promising treatment for PDAC.

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Cell Type of Pancreatic Ductal Adenocarcinoma Origin: Implications for Prognosis and Clinical Outcomes

Visceral Medicine ◽

10.1159/000520946 ◽

2021 ◽

pp. 1-6

Author(s):

Shilpa Patil ◽

Yan Dou ◽

Janel L. Kopp

Keyword(s):

Pancreatic Ductal Adenocarcinoma ◽

Mouse Models ◽

Pancreatic Injury ◽

Genetically Engineered ◽

Ductal Adenocarcinoma ◽

Cell Of Origin ◽

Cell Type ◽

Genetically Engineered Mouse Models ◽

Cellular Origin ◽

Ductal Cells

Background: Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease that has no effective early detection method or treatment to date. Summary: The normal cell type that initiates PDAC, or its cellular origin, is still unknown. To investigate the contribution of distinct normal epithelial cell types to PDAC tumorigenesis, genetically engineered mouse models were used to show that both acinar and ductal cells are capable of giving rise to PDAC. These studies indicated that genetic mutations and pancreatic injury interact differently with each cellular origin to affect their predilection and process for forming PDAC. In this review, we summarize recent findings using various genetically engineered mouse models in the identification and characterization of the PDAC cell of origin. We also discuss potential implications for cellular origin on tumor development, PDAC transcriptional subtype, and disease prognosis of patients. Key Message: Although it is clear that both ductal and acinar cells have the potential to form PDAC, whether cellular origin can indeed influence patient prognosis and whether knowledge of cellular origin will aid in the diagnosis or treatment of patients in the future will need further study.

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IL-6 and PD-L1 antibody blockade combination therapy reduces tumour progression in murine models of pancreatic cancer

Gut ◽

10.1136/gutjnl-2016-311585 ◽

2016 ◽

Vol 67 (2) ◽

pp. 320-332 ◽

Cited By ~ 159

Author(s):

Thomas A Mace ◽

Reena Shakya ◽

Jason R Pitarresi ◽

Benjamin Swanson ◽

Christopher W McQuinn ◽

...

Keyword(s):

Combination Therapy ◽

Checkpoint Inhibitors ◽

Tumour Progression ◽

Antitumour Activity ◽

Smooth Muscle Actin ◽

Immunohistochemical Analysis ◽

Genetically Engineered ◽

Pancreatic Stellate Cells ◽

Ductal Adenocarcinoma

ObjectiveLimited efficacy of immune checkpoint inhibitors in pancreatic ductal adenocarcinoma (PDAC) has prompted investigation into combination therapy. We hypothesised that interleukin 6 (IL-6) blockade would modulate immunological features of PDAC and enhance the efficacy of anti-programmed death-1-ligand 1 (PD-L1) checkpoint inhibitor therapy.DesignTranscription profiles and IL-6 secretion from primary patient-derived pancreatic stellate cells (PSCs) were analyzed via Nanostring and immunohistochemistry, respectively. In vivo efficacy and mechanistic studies were conducted with antibodies (Abs) targeting IL-6, PD-L1, CD4 or CD8 in subcutaneous or orthotopic models using Panc02, MT5 or KPC-luc cell lines; and the aggressive, genetically engineered PDAC model (KrasLSL−G12D, Trp53LSL−R270H, Pdx1-cre, Brca2F/F (KPC-Brca2 mice)). Systemic and local changes in immunophenotype were measured by flow cytometry or immunohistochemical analysis.ResultsPSCs (n=12) demonstrated prominent IL-6 expression, which was localised to stroma of tumours. Combined IL-6 and PD-L1 blockade elicited efficacy in mice bearing subcutaneous MT5 (p<0.02) and Panc02 tumours (p=0.046), which was accompanied by increased intratumoural effector T lymphocytes (CD62L−CD44−). CD8-depleting but not CD4-depleting Abs abrogated the efficacy of combined IL-6 and PD-L1 blockade in mice bearing Panc02 tumours (p=0.0016). This treatment combination also elicited significant antitumour activity in mice bearing orthotopic KPC-luc tumours and limited tumour progression in KPC-Brca2 mice (p<0.001). Histological analysis revealed increased T-cell infiltration and reduced α-smooth muscle actin cells in tumours from multiple models. Finally, IL-6 and PD-L1 blockade increased overall survival in KPC-Brca2 mice compared with isotype controls (p=0.0012).ConclusionsThese preclinical results indicate that targeted inhibition of IL-6 may enhance the efficacy of anti-PD-L1 in PDAC.

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Actin/microtubule crosstalk during platelet biogenesis in mice is critically regulated by Twinfilin1 and Cofilin1

Blood Advances ◽

10.1182/bloodadvances.2019001303 ◽

2020 ◽

Vol 4 (10) ◽

pp. 2124-2134 ◽

Cited By ~ 2

Author(s):

Isabelle C. Becker ◽

Inga Scheller ◽

Lou M. Wackerbarth ◽

Sarah Beck ◽

Tobias Heib ◽

...

Keyword(s):

Regulatory Proteins ◽

Actin Dynamics ◽

Adenomatous Polyposis ◽

Polyposis Coli ◽

And Function ◽

Redundant Functions ◽

H Protein ◽

Functional Defects

Abstract Rearrangements of the microtubule (MT) and actin cytoskeleton are pivotal for platelet biogenesis. Hence, defects in actin- or MT-regulatory proteins are associated with platelet disorders in humans and mice. Previous studies in mice revealed that loss of the actin-depolymerizing factor homology (ADF-H) protein Cofilin1 (Cof1) in megakaryocytes (MKs) results in a moderate macrothrombocytopenia but normal MK numbers, whereas deficiency in another ADF-H protein, Twinfilin1 (Twf1), does not affect platelet production or function. However, recent studies in yeast have indicated a critical synergism between Twf1 and Cof1 in the regulation of actin dynamics. We therefore investigated platelet biogenesis and function in mice lacking both Twf1 and Cof1 in the MK lineage. In contrast to single deficiency in either protein, Twf1/Cof1 double deficiency (DKO) resulted in a severe macrothrombocytopenia and dramatically increased MK numbers in bone marrow and spleen. DKO MKs exhibited defective proplatelet formation in vitro and in vivo as well as impaired spreading and altered assembly of podosome-like structures on collagen and fibrinogen in vitro. These defects were associated with aberrant F-actin accumulation and, remarkably, the formation of hyperstable MT, which appears to be caused by dysregulation of the actin- and MT-binding proteins mDia1 and adenomatous polyposis coli. Surprisingly, the mild functional defects described for Cof1-deficient platelets were only slightly aggravated in DKO platelets suggesting that both proteins are largely dispensable for platelet function in the peripheral blood. In summary, these findings reveal critical redundant functions of Cof1 and Twf1 in ensuring balanced actin/microtubule crosstalk during thrombopoiesis in mice and possibly humans.

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Molecular drivers and cells of origin in pancreatic ductal adenocarcinoma and pancreatic neuroendocrine carcinoma

Experimental Hematology and Oncology ◽

10.1186/s40164-020-00184-0 ◽

2020 ◽

Vol 9 (1) ◽

Author(s):

He-Li Gao ◽

Wen-Quan Wang ◽

Xian-Jun Yu ◽

Liang Liu

Keyword(s):

Pancreatic Cancer ◽

Pancreatic Ductal Adenocarcinoma ◽

Neuroendocrine Carcinoma ◽

Pancreatic Neuroendocrine Tumor ◽

Ductal Adenocarcinoma ◽

Neuroendocrine Neoplasm ◽

Cell Of Origin ◽

Cells Of Origin ◽

Pancreatic Neuroendocrine Neoplasm ◽

Pancreatic Neuroendocrine Carcinoma

Abstract Pancreatic cancer is one of the most common causes of cancer-related deaths worldwide. The two major histological subtypes of pancreatic cancer are pancreatic ductal adenocarcinoma (PDAC), accounting for 90% of all cases, and pancreatic neuroendocrine neoplasm (PanNEN), which makes up 3–5% of all cases. PanNEN is classified into well-differentiated pancreatic neuroendocrine tumor and poorly-differentiated pancreatic neuroendocrine carcinoma (PanNEC). Although PDAC and PanNEN are commonly thought to be different diseases with distinct biology, cell of origin, and genomic abnormalities, the idea that PDAC and PanNEC share common cells of origin has been gaining support. This is substantiated by evidence that the molecular profiling of PanNEC is genetically and phenotypically related to PDAC. In the current review, we summarize published studies pointing to common potential cells of origin and speculate about how the distinct paths of differentiation are determined by the genomic patterns of each disease. We also discuss the overlap between PDAC and PanNEC, which has been noted in clinical observations.

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