Fibrotic Stroma Research Articles

Therapeutic Strategies to Overcome Fibrotic Barriers to Nanomedicine in the Pancreatic Tumor Microenvironment.

Pancreatic cancer is notorious for its dismal prognosis. The enhanced permeability and retention (EPR) effect theory posits that nanomedicines (therapeutics in the size range of approximately 10-200 nm) selectively accumulate in tumors. Nanomedicine has thus been suggested to be the "magic bullet"-both effective and safe-to treat pancreatic cancer. However, the densely fibrotic tumor microenvironment of pancreatic cancer impedes nanomedicine delivery. The EPR effect is thus insufficient to achieve a significant therapeutic effect. Intratumoral fibrosis is chiefly driven by aberrantly activated fibroblasts and the extracellular matrix (ECM) components secreted. Fibroblast and ECM abnormalities offer various potential targets for therapeutic intervention. In this review, we detail the diverse strategies being tested to overcome the fibrotic barriers to nanomedicine in pancreatic cancer. Strategies that target the fibrotic tissue/process are discussed first, which are followed by strategies to optimize nanomedicine design. We provide an overview of how a deeper understanding, increasingly at single-cell resolution, of fibroblast biology is revealing the complex role of the fibrotic stroma in pancreatic cancer pathogenesis and consider the therapeutic implications. Finally, we discuss critical gaps in our understanding and how we might better formulate strategies to successfully overcome the fibrotic barriers in pancreatic cancer.

Histomorphological Evaluation of Desmoplastic Tumor Stroma in Malignant Ovarian Surface Epithelial Tumors.

Ovarian cancer is the 8th most common cancer in women worldwide. Tumor budding is defined as a type of invasive growth in carcinomas with either a single tumor cell or a cluster of up to four cells at the invasive tumor front and is associated with epithelial-mesenchymal transition. A reactive stroma rich in cancer-associated fibroblasts is associated with higher tumor grade and poorer prognosis in breast, colorectal, and oral cancers. The present study was conducted to highlight the prognostic significance of tumor budding and fibrotic cancer stroma in malignant ovarian surface epithelial tumors with known prognostic parameters. This was a retrospective cross-sectional study conducted over a 2-year period, in which all histologically diagnosed cases of malignant ovarian surface epithelial tumors who underwent surgery were included. The fibrotic stroma was classified into three distinct categories - mature, intermediate, and immature. The number of tumor buds was counted at the invasive front of the tumor and graded based on the number of buds - 0-5, 5-9, and ≥10 buds. Among the 50 cases, 32% (16 cases) had mature stroma, whereas 30% (15 cases) and 38% (19 cases) had intermediate and immature stroma, respectively. Although a significant association could not be established between tumor budding and stroma grade, a fair agreement was established between them. A significant association could be established between histological grade with both tumor budding (P = 0.03) and fibrotic stroma grade (P = 0.02). The study highlighted the role of stromal response in malignant surface epithelial tumors of the ovary since a higher-grade tumor was associated with an immature stroma, whereas a lower-grade tumor was associated with a mature stroma.

RBPJ Deficiency Sensitizes Pancreatic Acinar Cells to KRAS-Mediated Pancreatic Intraepithelial Neoplasia Initiation

Development of pancreatic ductal adenocarcinoma (PDAC) is a multistep process intensively studied; however, precocious diagnosis and effective therapy still remain unsatisfactory. The role for Notch signaling in PDAC has been discussed controversially, as both cancer-promoting and cancer-antagonizing functions have been described. Thus, an improved understanding of the underlying molecular mechanisms is necessary. Here, we focused on RBPJ, the receiving transcription factor in the Notch pathway, examined its expression pattern in PDAC, and characterized its function in mouse models of pancreatic cancer development and in the regeneration process after acute pancreatitis. Conditional transgenic mouse models were usedfor functional analysis of RBPJ in the adult pancreas, initiation of PDAC precursor lesions, and pancreatic regeneration. Pancreata and primary acinar cells were tested for acinar-to-ductal metaplasia together with immunohistology and comprehensive transcriptional profiling by RNA sequencing. We identified reduced RBPJ expression in a subset of human PDAC specimens. Ptf1α-CreERT-driven depletion of RBPJ in transgenic mice revealed that its function is dispensable for the homeostasis and maintenance of adult acinar cells. However, primary RBPJ-deficient acinar cells underwent acinar-to-ductal differentiation in exvivo. Importantly, oncogenic KRAS expression in the context of RBPJ deficiency facilitated the development of pancreatic intraepithelial neoplasia lesions with massive fibrotic stroma formation. Interestingly, RNA-sequencing data revealed a transcriptional profile associated with the cytokine/chemokine and extracellular matrix changes. In addition, lack of RBPJ delays the course of acute pancreatitis and critically impairs it in the context of KRASG12D expression. Our findings imply that downregulation of RBPJ in PDAC patients derepresses Notch targets and promotes KRAS-mediated pancreatic acinar cells transformation and desmoplasia development.

Abstract A014: Targeting TAK1 for intercepting PanIN progression to ductal adenocarcinoma in an inducible KC mouse model

Abstract Despite advances in our knowledge of human pancreatic ductal adenocarcinoma (PDAC), targeted therapies have not yet significantly translated to an improved overall survival for patients. Pancreatic tumor microenvironment enriched by infiltrating immune cells and consisting of dense fibrotic stroma is characterized by desmoplasia, the major contributors of tumor-associated inflammation. Transforming growth factor-β (TGF-β)-activated kinase 1 (TAK1) is widely accepted as a key player in the TNF-α and TGF-β -induced signaling, and principal contributor of tumor fibrosis, inflammation and cell proliferation. Here, we show that pancreatic tumors over- express TAK1, and are associated with fibrotic-stroma and infiltrating immune cells. Also, p-TAK1(ser412) expression is correlated with progression of pancreatic intraepithelial neoplasia (PanIN) lesions to PDAC. Thus, we hypothesized TAK1 as an important target for inhibition of the tumor-associated fibrosis, inflammation and PanIN progression to PDAC. To prove above hypothesis, we tested a selective TAK1 inhibitor, Takinib for its toxicity and efficacy against PanIN progression in the inducible Ptf1CreERT.LSL-KrasG12D (KC) mouse model. Takinib was synthesized and fed to wild type C57BL/6J mice (n=6) at 250 ppm and 500ppm in diet for 6 weeks to determine toxicity. Bodyweight gain, organ weights and serum enzyme analysis did not indicate any toxicity at the tested doses. For efficacy study, LSL-KrasG12D mice were bred with Ptf1-CreERTmice. Pups were genotyped and randomized to groups (n=12). PanINs -PDAC was induced in the KC mice by tamoxifen (oral gavage) followed by 250ppm Takinib administration in diet for 20 weeks. After termination, pancreas tissue sections were evaluated for PanIN multiplicity and PDAC incidence/spread. Administration of Takinib led to significant reduction in PanIN 1 by 54% (p<0.02), PanIN 2 by 77% (p<0.001) and PanIN 3 by 80% (p<0.02) in the KC mice compared to untreated mice. PDAC incidence was also reduced with an increase in normal pancreas in Takinib administered KC mice. Taken together our results suggest that TAK1 is a valuable target for PDAC, and Takinib possesses efficacy against the PanIN and PDAC progression in the KC mouse model warranting further studies. (Supported by PHF Team Science grant and Kerley-Cade Endowed Chair). Citation Format: Venkateshwar Madka, Gopal Pathuri, Gaurav Kuma, Anil Singh, Nicole Stratton, Courtney Houchen, Altaf Mohammed, Chinthalapally V. Rao. Targeting TAK1 for intercepting PanIN progression to ductal adenocarcinoma in an inducible KC mouse model [abstract]. In: Proceedings of the Second Biennial NCI Meeting: Translational Advances in Cancer Prevention Agent Development (TACPAD); 2022 Sep 7-9. Philadelphia (PA): AACR; Can Prev Res 2022;15(12 Suppl_2): Abstract nr A014.

Abstract B060: Investigating the role of ERphagy in pancreatic cancer

Abstract Pancreatic ductal adenocarcinoma (PDAC) harbors mutations in KRAS in greater than 90% of cases and is characterized by a dense, fibrotic stroma that fosters the formation of a hypoxic, nutrient-poor tumor microenvironment. Tumor cells counteract these stresses through the hyperactivation of multiple nutrient scavenging pathways, including macropinoytosis and macroautophagy. Whereas the role of the latter in the regulation of PDAC growth has been extensively studied, much less is known about the role of endoplasmic reticulum (ER)-specific autophagy (ERphagy), a homeostatic mechanism involving ER membrane remodeling which promotes the clearance of misfolded protein aggregates, thereby supporting proteostasis. Using an ERphagy reporter system, we found that pancreatic cancer cells display mutant KRas-dependent suppression of ERphagy both at baseline and in the context of nutrient deprivation, which was partially dependent on the ER-resident receptor CCPG1. Furthermore, direct perturbation of the 26S proteasome with bortezomib enhanced ERphagy, suggesting that the attenuated ERphagy capacity in pancreatic cancer cells may reflect a greater dependence on proteasomal activity for the maintenance of proteostasis. Significantly, the knockdown of CCPG1 but not other ERphagy receptors augmented the growth-suppressive effects of bortezomib in vitro, an effect likely caused by unresolved proteotoxic stress. Overall, our findings implicate CCPG1-mediated ERphagy in pancreatic cancer cell survival and identify a targetable cell-intrinsic vulnerability that can potentiate the cytotoxic effects of bortezomib, which alone is clinically ineffective in treating PDAC. Citation Format: Sandra L. Vogt, Dafna Bar-Sagi. Investigating the role of ERphagy in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr B060.

Abstract C061: Alternative polyadenylation regulates gene expression within the pancreatic cancer tumor microenvironment

Abstract A characteristic of pancreatic ductal adenocarcinoma (PDAC) is that about 90% of its tumor volume can be made up of dense, fibrotic stroma. Multiple studies have shown that this dense stroma strongly influences disease progression and drug delivery. Amongst the cells that make up the stroma, Cancer Associated Fibroblasts (CAFs) are important regulators directly affecting cancer progression and therapeutic resistance. CAFs are a heterogeneous population, including inflammatory, myofibroblastic, and antigen-presenting CAFs, each with specific functional contributions to the cancer phenotype. While we have been able to define CAFs phenotypically, we lack a deeper understanding of the mechanisms underlying their transcriptional heterogeneity and ultimately their contribution in driving PDAC progression. Our lab has previously shown that 3’UTR-Alternative Polyadenylation (APA) is a key mechanism underlying gene dysregulation in PDAC. APA machinery makes use of multiple Poly A Sites (PAS) in a transcript and leads to the formation of distinct isoforms of mRNA with varying lengths, commonly referred to as short or long isoforms. This lengthening and shortening of mRNA modulates mRNA stability, protein expression, and localization, thus directly affecting cellular processes including proliferation, metastasis, and migration. We have now extended these findings to an analysis of APA events within the tumor microenvironment (TME). We find that 3’ UTR shortening is significantly associated with inflammatory CAFs (iCAFs), and that these APA events are correlated with gene expression changes in iCAF marker genes. We hypothesize that APA is a crucial driver of heterogeneity within the tumor microenvironment of PDAC. We have begun to elucidate the mechanisms underlying these gene regulatory changes in vitro using human-derived CAF cell lines via genetic and pharmacological inhibition of APA. Understanding these mechanisms may help us find druggable targets that may eventually affect the overall outcomes of PDAC patients. Citation Format: Aditi H Chaubey, Michael E Feigin. Alternative polyadenylation regulates gene expression within the pancreatic cancer tumor microenvironment [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr C061.

Abstract C008: Intratumoral fibrin as a novel immunomodulatory factor in pancreatic ductal adenocarcinoma

Abstract Introduction: Pancreatic ductal adenocarcinomas (PDACs) are poorly responsive to both chemo- and immunotherapies, mainly because of the presence of highly dense fibrotic stroma and extensive myeloid-derived immune cells, which are known major drivers for an immunosuppressive tumor microenvironment (TME). Another feature of PDACs that may suppress the immune microenvironment is the small ‘fibrin clots’ that are chiefly found in tumor vasculature and interstitial space. PDACs exhibit high levels of fibrinogen and fibrin clots throughout the tumor stroma surrounding the tumor cells. Fibrinogen, a coagulation factor that polymerizes into fibrin, is overrepresented in the stroma from early-to-late stage PDA, suggesting an early activation of coagulation cascade in PDAC. Taken together, we hypothesize that cross-linked fibrin-stroma may suppress the immune microenvironment of PDAC. Methods: Analyzing transcriptome of PDAC patients, we observed the clinical correlation between fibrin-clot relevant genes (FGA, FGB, and FGG) with various immune genes’ expressions and overall survival. We performed multiplex analysis of tumor tissues from PDAC patients (N = 25) with higher and lower fibrin expression, and correlated the abundance of tumor fibrin-clot with plethora of immune cells in the tumor stroma. Using a microfluidic tissue-on-a-chip PDAC model, we determined the role of fibrin in TME immune landscape focusing on macrophage differentiation. Knocking out of FGB in KPC689 cells (KPC689FGB-/-), derived from KPC mice, we determined the role of targeting tumor fibrin in attenuating immunosuppressive TME. Results: Patients with higher expression of FGA, FGB, and FGG (> median) showed significantly lower survival rate in PDAC (p < 0.002) compared to those with lower expression (< median). Markedly, the expression levels of FGB positively correlated with various immune-related genes in PDAC patients, including PD1-L1, PD1-L2, PD-1, FOXP3, macrophage-1 antigen (ITGAM), and CD8β+. Multiplex analysis revealed that Fibrinlow PDAC tissues had higher abundance of CD8+ T-cells but higher number of M2 (CD163+/MCHII-) versus M1 (CD68+/MHCII+/CD163-) macrophages than in Fibrinhigh PDAC tissues. In PDAC-on-a-chip model, human monocytes, when cultured in collagen gels into one compartment along with PDAC in Peak 2 Fibrin (P2F) gels into the side compartment, induced M2 phenotype, infiltrated into the PDAC/P2F compartment, and increased the survival of PDAC cells. Both KPC689FGB+/+ and KPC689FGB-/- cells grew at the same rate, when inoculated into the pancrease of mice, but the abundance of fibrin-laden stroma decreased in KPC689FGB-/- tumors. FACS analysis from explanted tumors showed higher number of activated IFNγ+CD8β+ T cells but lower number of CD11b+Gr1+ MDSCs in KPC689FGB-/- than KPC689FGB+/+ tumors. Conclusion: Our data suggest that in-situ fibrin-formation controls immune microenvironment in PDAC and targeting intratumoral fibrin formation can ‘heat up’ the TME by mitigating the immunosuppression in PDAC. Citation Format: Riajul Wahab, Mazharul Karim, Md Mahedi Hassan, Paul Grippo, Faraz Bisheshari, Taslim Al-Hilal. Intratumoral fibrin as a novel immunomodulatory factor in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr C008.

Polarization clustering of biological structures with Mueller matrix parameters.

Mueller matrix imaging polarimetry (MMIP) is a promising technique for the characterization of biological tissues, including the classification of microstructures in pathological diagnosis. To expand the parameter space of Mueller matrix parameters, we propose new vector parameters (VPs) according to the Mueller matrix polar decomposition method. We measure invasive bladder cancer (IBC) with extensive necrosis and high-grade ductal carcinoma in situ (DCIS) with MMIP, and the regions of cancer cells and fibrotic stroma are classified with the VPs. Then the proposed and existing VPs are mapped on the Poincaré sphere with 3D visualization, and an indicator of spatial feature is defined based on the minimum enclosing sphere to evaluate the classification capability of the VPs. For both IBC and DCIS, the results show that the proposed VPs exhibit evident contrast between the regions of cancer cells and fibrotic stroma. This study broadens the fundamental Mueller matrix parameters and helps to improve the characterization ability of the MMIP technique.

Exploring the Biology of Cancer-Associated Fibroblasts in Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignancy characterised by a stubbornly low 5-year survival which is essentially unchanged in the past 5 decades. Despite recent advances in chemotherapy and surgical outcomes, progress continues to lag behind that of other cancers. The PDAC microenvironment is characterised by a dense, fibrotic stroma of which cancer-associated fibroblasts (CAFs) are key players. CAFs and fibrosis were initially thought to be uniformly tumour-promoting, however this doctrine is now being challenged by a wealth of evidence demonstrating CAF phenotypic and functional heterogeneity. Recent technological advances have allowed for the molecular profiling of the PDAC tumour microenvironment at exceptional detail, and these technologies are being leveraged at pace to improve our understanding of this previously elusive cell population. In this review we discuss CAF heterogeneity and recent developments in CAF biology. We explore the complex relationship between CAFs and other cell types within the PDAC microenvironment. We discuss the potential for therapeutic targeting of CAFs, and we finally provide an overview of future directions for the field and the possibility of improving outcomes for patients with this devastating disease.

CXCR4-CXCL12-CXCR7 and PD-1/PD-L1 in Pancreatic Cancer: CXCL12 Predicts Survival of Radically Resected Patients.

Pancreatic ductal adenocarcinoma (PDAC) is currently the most deadly cancer. Although characterized by 5-20% of neoplastic cells in the highly fibrotic stroma, immunotherapy is not a valid option in PDAC treatment. As CXCR4-CXCL12 regulates tumor invasion and T-cell access and PD-1/PD-L1 controls immune tolerance, 76 PDACs were evaluated for CXCR4-CXCL12-CXCR7 and PD-1/PD-L1 in the epithelial and stromal component. Neoplastic CXCR4 and CXCL12 discriminated PDACs for recurrence-free survival (RFS), while CXCL12 and CXCR7 discriminated patients for cancer-specific survival (CSS). Interestingly, among patients with radical resection (R0), high tumor CXCR4 clustered patients with worse RFS, high CXCL12 identified poor prognostic patients for both RFS and CSS, while stromal lymphocytic-monocytic PD-L1 associated with improved RFS and CSS. PD-1 was only sporadically expressed (<1%) in focal lymphocyte infiltrate and does not impact prognosis. In multivariate analysis, tumoral CXCL12, perineural invasion, and AJCC lymph node status were independent prognostic factors for RFS; tumoral CXCL12, AJCC Stage, and vascular invasion were independent prognostic factors for CSS. CXCL12's poor prognostic meaning was confirmed in an additional perspective-independent 13 fine-needle aspiration cytology advanced stage-PDACs. Thus, CXCR4-CXCL12 evaluation in PDAC identifies prognostic categories and could orient therapeutic approaches.

Fibroblast activation protein-based theranostics in pancreatic cancer.

Fibroblast activation protein-α (FAP) is a type II transmembrane serine protease that has specific endopeptidase activity. Given its well-established selective expression in the activated stromal fibroblasts of epithelial cancers, although not in quiescent fibroblasts, FAP has received substantial research attention as a diagnostic marker and therapeutic target. Pancreatic cancer is characterized by an abundant fibrotic or desmoplastic stroma, leading to rapid progression, therapeutic resistance, and poor clinical outcomes. Numerous studies have revealed that the abundant expression of FAP in cancer cells, circulating tumor cells, stromal cells, and cancer-associated fibroblasts (CAFs) of pancreatic adenocarcinoma is implicated in diverse cancer-related signaling pathways, contributing to cancer progression, invasion, migration, metastasis, immunosuppression, and resistance to treatment. In this article, we aim to systematically review the recent advances in research on FAP in pancreatic adenocarcinoma, including its utility as a diagnostic marker, therapeutic potential, and correlation with prognosis. We also describe the functional role of FAP-overexpressing stromal cells, particulary CAFs, in tumor immuno- and metabolic microenvironments, and summarize the mechanisms underlying the contribution of FAP-overexpressing CAFs in pancreatic cancer progression and treatment resistance. Furthermore, we discuss whether targeting FAP-overexpressing CAFs could represent a potential therapeutic strategy and describe the development of FAP-targeted probes for diagnostic imaging. Finally, we assess the emerging basic and clinical studies regarding the bench-to-bedside translation of FAP in pancreatic cancer.

It Melts Like Snow: Steroid Responsiveness of Immunoglobulin G4-Related Disease

Dear Editor, A 35-year-old male presented with insidious onset gradually progressive bilateral painless submandibular swelling, followed by periorbital swelling [Figure 1] for the past 7 years. He had no constitutional or sicca symptoms. He had no history of addiction or autoimmunity in the family. Systemic examination was unremarkable. On evaluation, he had raised inflammatory markers with a negative autoimmune panel. Magnetic resonance imaging orbit showed bilateral orbital extraconal soft-tissue masses with contrast enhancement. Incisional biopsy from the extraconal mass was suggestive of fibrocollagenous stroma with aggregation of lymphocytes, plasma cells, and histiocytes without any necrosis or granuloma. On further evaluation, submandibular gland biopsy showed chronic sialadenitis with immunoglobulin G4 (IgG4)-positive plasma cell >10/hpf and IgG4: IgG ratio >40% [Figure 2] without any major organ involvement on imaging. The diagnosis of IgG4-related disease was kept. He was started on oral steroid 0.5 mg/kg/day and observed a rapid clinical response within 15 days of steroid therapy. Later, he was started on azathioprine and continued with tapering dose of steroid. He was followed up every month for first 3 months and then every 3 monthly upto 16 months without any signs of recurrence. For a long-standing indolent disease of 7 years, drastic steroid response with resolution of clinical symptoms was quite evident in our case.Figure 1: (a) Pretreatment bilateral upper eyelid swelling, (b) pretreatment bilateral submandibular swelling, (c and d) during glucocorticoid therapy (day 15) resolution of orbital and submandibular swelling, respectivelyFigure 2: Salivary gland with seromucinous acini and ducts embedded in mildly fibrotic stroma with numerous lymphoid follicles with germinal centers (a and b: H and E). Clusters of plasma cells are highlighted with CD138 immunohistochemical stain (c). IgG4 positive cells, >10/hpf (d)IgG4-related disease (IgG4-RD) is a chronic, systemic, fibroinflammatory condition with variable organ manifestation. CD4+ T-cells and B-cells are central to the pathogenesis causing organ damage and fibrosis. Marked expansion of IgG4-secreting plasmablast in the blood is usually seen in active and untreated cases.[1] Key to diagnosis is the classical histopathological hallmarks of lymphoplasmacytic infiltrate, storiform fibrosis, and obliterative phlebitis. IgG4 RD usually has a good response to steroids but a variable relapse rate. Factors associated with lesser treatment response and relapse were well analyzed in a Japanese multicenter study. Multivariate analysis from the study revealed that intermediate steroid initial dose (0.4–0.69 mg/day), slow tapering regimen (<0.4 mg/day), and shorter disease duration were associated with a significantly lower rate of disease relapse.[2] The study of isolated IgG4 ophthalmic disease also echoed similar results regarding steroid response and relapse, with more relapse in those with elevated serum IgG4 levels and a shorter duration of initial steroid treatment (<5 months). Low-dose steroid maintenance should be considered to avoid recurrence in patients with elevated serum IgG4 levels.[3] Later on, rituximab therapy showed promising results in inducing remission[4] and steroid-free long-term maintenance.[5] Declaration of patient consent The authors certify that they have obtained all appropriate patient consent forms. In the form, the patient has given his consent for his images and other clinical information to be reported in the journal. The patient understands that his name and initials will not be published and due efforts will be made to conceal identity, but anonymity cannot be guaranteed. Financial support and sponsorship Nil. Conflicts of interest There are no conflicts of interest.

Mouse Mammary Gland Whole Mount Density Assessment across Different Morphologies Using a Bifurcated Program for Image Processing

Mammographic density is associated with increased breast cancer risk. Conventional visual assessment of murine mouse models does not include quantified total density analysis. A bifurcated method was sufficient to obtain relative density scores on a broad range of two-dimensional whole mount images that contained both normal and abnormal findings. Image processing techniques, including a ridge operator and a gaussian denoising method, were used to isolate background away from mammary epithelium and use mean pixel intensity to represent mammary density on genetically engineered mouse models for breast cancer in mice 4 to 29 months of age. The bifurcated method allowed for application of an optimal image processing approach for the structural elements present in the whole mount images. Gaussian denoising was the optimal approach when more dense lobular growth and tertiary branching dominate and a ridge operator when epithelial growth was more sparse and secondary branching was the more dominant structural feature. The two processing approaches were combined in a single experimental flow program using an initial image density measurement as the decision point between the two approaches. Higher density was associated with lobular growth, tertiary branching, fibrotic stroma, and presence of cancer. The significance of the study is development of a readily accessible program for digital assessment of mammary gland whole mount density across a range of mammary gland morphologies.

34089 Syringoma of the vulva associated with vulvar hypertrophy

Syringoma is a benign eccrine sweat gland tumor most commonly presenting in young adults as multiple, often clustered, soft 1-2 mm papules on the lower eyelids and cheeks. There is a female predominance and lesions tend to arise after puberty. Syringoma of the vulva is less common with fewer cases reported. Here we describe a case of a 35-year-old woman who presented with multiple pruritic skin-colored papules on the labia majora which had been present for 7 years and were increasing in size and number. The lesions were a source of distress with intimate encounters and the associated vulvar thickening was visible under some clothing. Examination revealed numerous coalescing, 2-4 mm skin-colored, slightly rubbery papules on the bilateral labia majora with associated vulvar hypertrophy. A biopsy showed small comma-shaped ducts and solid cords of epithelium surrounded by a fibrotic stroma compatible with a diagnosis of syringoma. Various treatment options were presented, including excision, carbon dioxide laser ablation, and topical atropine, although surgical excision was favored given the extent of the lesions and resultant hypertrophy. Our case highlights the importance of keeping vulvar syringoma in the differential diagnosis of patients with bilateral vulvar papules, vulvar pruritus and/or vulvar hypertrophy, as well as recognizing the potential impact of this benign condition on patient quality of life.

The Extracellular Matrix: A Key Accomplice of Cancer Stem Cell Migration, Metastasis Formation, and Drug Resistance in PDAC.

Simple SummaryThis review takes the extracellular matrix (ECM) as the starting point, and describes its influence and related mechanisms on the biological behavior of pancreatic ductal carcinoma (PDAC). It focuses on how the ECM regulates cancer stem cells and thus affects the metastasis and drug resistance of PDAC. Finally, current and ongoing treatment strategies for ECM are presented. ECM-related factors and mechanisms are both the focus and difficulty of current therapeutic strategy research, but further exploration is crucial for effectively eradicating the disease and improving patient survival.Pancreatic ductal adenocarcinoma (PDAC) is rich in dense fibrotic stroma that are composed of extracellular matrix (ECM) proteins. A disruption of the balance between ECM synthesis and secretion and the altered expression of matrix remodeling enzymes lead to abnormal ECM dynamics in PDAC. This pathological ECM promotes cancer growth, survival, invasion, and alters the behavior of fibroblasts and immune cells leading to metastasis formation and chemotherapy resistance, which contribute to the high lethality of PDAC. Additionally, recent evidence highlights that ECM, as a major structural component of the tumor microenvironment, is a highly dynamic structure in which ECM proteins establish a physical and biochemical niche for cancer stem cells (CSCs). CSCs are characterized by self-renewal, tumor initiation, and resistance to chemotherapeutics. In this review, we will discuss the effects of the ECM on tumor biological behavior and its molecular impact on the fundamental signaling pathways in PDAC. We will also provide an overview of how the different ECM components are able to modulate CSCs properties and finally discuss the current and ongoing therapeutic strategies targeting the ECM. Given the many challenges facing current targeted therapies for PDAC, a better understanding of molecular events involving the interplay of ECM and CSC will be key in identifying more effective therapeutic strategies to eliminate CSCs and ultimately to improve survival in patients that are suffering from this deadly disease.

Focal adhesion kinase priming in pancreatic cancer, altering biomechanics to improve chemotherapy.

The dense desmoplastic and fibrotic stroma is a characteristic feature of pancreatic ductal adenocarcinoma (PDAC), regulating disease progression, metastasis and response to treatment. Reciprocal interactions between the tumour and stroma are mediated by bidirectional integrin-mediated signalling, in particular by Focal Adhesion Kinase (FAK). FAK is often hyperactivated and overexpressed in aggressive cancers, promoting stromal remodelling and inducing tissue stiffness which can accelerate cancer cell proliferation, survival and chemoresistance. Therapeutic targeting of the PDAC stroma is an evolving area of interest for pre-clinical and clinical research, where a subtle reshaping of the stromal architecture prior to chemotherapy may prove promising in the clinical management of disease and overall patient survival. Here, we describe how transient stromal manipulation (or ‘priming’) via short-term FAK inhibition, rather than chronic treatment, can render PDAC cells exquisitely vulnerable to subsequent standard-of-care chemotherapy. We assess how our priming publication fits with the recent literature and describe in this perspective how this could impact future cancer treatment. This highlights the significance of treatment timing and warrants further consideration of anti-fibrotic therapies in the clinical management of PDAC and other fibrotic diseases.

Cancer Associated Fibroblast (CAF) Regulation of PDAC Parenchymal (CPC) and CSC Phenotypes Is Modulated by ECM Composition.

Simple SummaryHere, we demonstrate for the first time that ECM composition cooperates with CAFs to jointly regulate/modulate the highly dynamic interactions between the CPC and CSC cell lines and establish a continuum between tumor initiation and progression in primary PDAC tumors. Altogether, these findings propose a scenario in which the ECM composition and the cellular secretome of the CAFs cooperate to jointly regulate both growth and morphology of the CPC and CSC cell lines and, by modulating the highly dynamic interactions between them, establishes a continuum between tumor initiation and progression in primary PDAC tumors.Background: Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest of all cancers, having one of the lowest five-year survival rates. One of its hallmarks is a dense desmoplastic stroma consisting in the abnormal accumulation of extracellular matrix (ECM) components, especially Collagen I. This highly fibrotic stroma embeds the bulk cancer (parenchymal) cells (CPCs), cancer stem cells (CSCs) and the main producers of the stromal reaction, the Cancer Associated Fibroblasts (CAFs). Little is known about the role of the acellular ECM in the interplay of the CAFs with the different tumor cell types in determining their phenotypic plasticity and eventual cell fate. Methods: Here, we analyzed the role of ECM collagen I in modulating the effect of CAF-derived signals by incubating PDAC CPCs and CSCs grown on ECM mimicking early (low collagen I levels) and late (high collagen I levels) stage PDAC stroma with conditioned medium from primary cultured CAFs derived from patients with PDAC in a previously described three-dimensional (3D) organotypic model of PDAC. Results: We found that CAFs (1) reduced CPC growth while favoring CSC growth independently of the ECM; (2) increased the invasive capacity of only CPCs on the ECM mimicking the early tumor; and (3) favored vasculogenic mimicry (VM) especially of the CSCs on the ECM mimicking an early tumor. Conclusions: We conclude that the CAFs and acellular stromal components interact to modulate the tumor behaviors of the PDAC CPC and CSC cell types and drive metastatic progression by stimulating the phenotypic characteristics of each tumor cell type that contribute to metastasis.

Reciprocal Interactions between Fibroblast and Pancreatic Neuroendocrine Tumor Cells: Putative Impact of the Tumor Microenvironment.

Simple SummaryTreatment of pancreatic neuroendocrine neoplasms (PNEN) is challenging since a subset of patients will have a metastatic disease at diagnosis and/or will experience resistance to the treatment. Whether the tumor microenvironment (TME) may represent a therapeutic target of interest in these tumors, remains to be elucidated. Our aim was to investigate the role played by stromal fibroblasts (namely, HPF and HFL-1) over the growth of human PNEN cell lines BON-1 and QGP-1, and vice versa. We confirmed a reciprocal stimulatory effect of HPF and HFL-1 over the growth of BON-1 and QGP-1, and the other way around. Likewise, the number of BON-1/QGP-1 colonies and the migration potency of both cell lines, significantly increased in presence of fibroblast-conditioned medium. Finally, the mTOR inhibitor, everolimus, mitigated the fibroblast-induced stimulatory effect over BON-1/QGP-1 cell lines, suggesting that fibroblasts, as an actor of the TME of PNEN, is a promising therapeutic target.Introduction: Pancreatic neuroendocrine neoplasms (PNENs) present with a fibrotic stroma that constitutes the tumor microenvironment (TME). The role played by stromal fibroblasts in the growth of PNENs and their sensitivity to the mTOR inhibitor RAD001 has not yet been established. Methods: We investigated reciprocal interactions between (1) human PNEN cell lines (BON-1/QGP-1) or primary cultures of human ileal neuroendocrine neoplasm (iNEN) or PNEN and (2) human fibroblast cell lines (HPF/HFL-1). Proliferation was assessed in transwell (tw) co-culture or in the presence of serum-free conditioned media (cm), with and without RAD001. Colony formation and migration of BON-1/QGP-1 were evaluated upon incubation with HPFcm. Results: Proliferation of BON-1 and QGP-1 increased in the presence of HFL-1cm, HPFcm, HFL-1tw and HPFtw (BON-1: +46–70% and QGP-1: +42–55%, p < 0.001 vs. controls) and HPFcm significantly increased the number of BON-1 or QGP-1 colonies (p < 0.05). This stimulatory effect was reversed in the presence of RAD001. Likewise, proliferation of human iNEN and PNEN primary cultures increased in the presence of HFL-1 or HPF. Reciprocally, BON-1cm and BONtw stimulated the proliferation of HPF (+90 ± 61% and +55 ± 47%, respectively, p < 0.001 vs. controls), an effect less pronounced with QGP-1cm or QGPtw (+19 to +27%, p < 0.05 vs. controls). Finally, a higher migration potential for BON-1 and QGP-1 was found in the presence of HPFcm (p < 0.001 vs. controls). Conclusions: Fibroblasts in the TME of PNENs represent a target of interest, the stimulatory effect of which over PNENs is mitigated by the mTOR inhibitor everolimus.

Histopathology and Immunohistochemical profile of mediastinal lymphoma in Dr. Soetomo General Hospital

Mediastinal lymphoma is a rare tumor. Approximately 10% of primary mediastinal lymphoma occur in mediastinum. In this study we look for sex and age characteristics, histopathology and immunohistochemical profile of mediastinal lymphoma. Data were collected from the Anatomical Pathology laboratory Dr. Soetomo archives from January 2016 to August 2018. Seventeen cases of mediastinal lymphoma were found predominantly in males (76.47 %). The age range was between 17 and 64 years old. Distribution of tumor type was as follows: Non-Hodgkin’s Lymphoma, B-cell type high grade (Ki67 &gt; 30%) 17.65 % and Non-Hodgkin’s Lymphoma, B-cell type low grade (Ki67 &lt;30%) 11.76%, Non-Hodgkin’s Lymphoma, T-cell type high grade 11.76%, Non-Hodgkin’s Lymphoma unknown subtype 35.29%, Classical Hodgkin’s Lymphoma (CHL) 17.65%, T-cell Lymphoblastic Lymphoma (T-LBL) 5.88%. Histologically, Non-Hodgkin’s Lymphoma showed a diffuse pattern consisting of anaplastic lymphoid cells, pleomorphic nuclei, coarse chromatin, thin cytoplasm between fibrotic stroma and the immunohistochemistry profile showed positive for CD20 in NHL B-cell type. CD3 were positive in Peripheral T-cell Lymphoma and T-Lymphoblastic Lymphoma (T-LBL). Classical Hodgkin’s Lymphoma showed large cells (reed-stenberg cells) and Hodgkin’s cells dispersed between an inflammatory background, CD30 were positive in CHL cases. Tdt was positive in T-LBL cases.

Good and Bad Stroma in Pancreatic Cancer: Relevance of Functional States of Cancer-Associated Fibroblasts.

Simple SummaryRecent progress in research on the biology of cancer-associated fibroblasts (CAFs) in pancreatic ductal adenocarcinoma (PDAC) indicates their diverse states and plasticity, which may lead to good and bad stroma, suppressing and promoting cancer progression, respectively. The characteristics of the stroma differ spatially, even within the same tumors, based on the balance between cancer-restraining CAF and cancer-promoting CAF proliferation at the site. These heterogeneous CAFs also influence the sensitivity of PDAC to anticancer therapeutics. Further preclinical and clinical studies will advance our understanding of the roles of CAFs in disease progression and aid the development of therapeutics that modulate or ameliorate the tumor microenvironment in PDAC.A well-known feature of human pancreatic ductal adenocarcinoma (PDAC) is the extensive proliferation of cancer-associated fibroblasts (CAFs) and highly fibrotic stroma. Recent evidence, based mainly on single-cell analyses, has identified various subsets of CAFs in PDAC mouse models. However, we do not know how these CAF subsets are involved in the progression and drug resistance of human PDAC. Additionally, it remains unclear whether these diverse CAFs have distinct origins and are indicators of genuinely distinct CAF lineages or reflect different states of the same CAFs depending on the tumor microenvironment. Interestingly, recent preclinical studies have started to characterize the nature of cancer-restraining CAFs and have identified their markers Meflin and collagen type I alpha 1. These studies have led to the development of strategies to induce changes in CAF phenotypes using chemical reagents or recombinant viruses, and some of them have been tested in clinical studies. These strategies have the unique potential to convert the so-called bad stroma to good stroma and may also have therapeutic implications for non-cancer diseases such as fibrotic diseases. Together with recently developed sophisticated strategies that specifically target distinct CAF subsets via adoptive cell transfer therapy, vaccination, and antibody–drug conjugates, any future findings arising from these clinical efforts may expand our understanding of the significance of CAF diversity in human PDAC.