Vessel Co-option Research Articles

Dynamic differentiation of F4/80+ tumor-associated macrophage and its role in tumor vascularization in a syngeneic mouse model of colorectal liver metastasis

Tumor-associated macrophages (TAMs) are highly heterogeneous and play vital roles in tumor progression. Here we adopted a C57BL/6 mouse model imitating the late-stage colorectal liver metastasis (CRLM) by Mc38 colorectal cancer cell injection via the portal vein. With serial sections of CRLM biopsies, we defined 7–9 days post-injection as the critical period for tumor neovascularization, which was initiated from the innate liver vessels via vessel cooption and extended by vascular mimicry and thereof growth of CD34+cells. In samples with increasing-sized liver metastases, the infiltrated Ly6C+ CD11b+ F4/80− monocytes steadily gained the expression of F4/80, a Kupffer cell marker, before transformed into Ly6C− CD11bint F4/80+ cells, which, the same phenotype was also adapted by Ly6C− CD11b− F4/80+ Kupffer cells. F4/80+ TAMs showed proximity to neovascularization and tumor vessels, functionally angiogenic in vivo; and greatly promoted the activation of a few key angiogenic markers such as VEGFA, Ki67, etc. in endothelial cells in vitro. Depletion of macrophages or diversion of macrophage polarization during neovascularization impeded tumor growth and vascularization and resulted in greatly reduced F4/80+ TAMs, yet increased CD11b+ cells due to inhibition of TAM differentiation. In summary, our results showed dynamic and spatial–temporal F4/80+ TAM transformation within the tumor microenvironment and strengthened its role as perivascular and angiogenic TAMs in CRLM.

Molecular differences of angiogenic versus vessel co-opting colorectal cancer liver metastases at single-cell resolution

BackgroundColorectal cancer liver metastases (CRCLM) are associated with a poor prognosis, reflected by a five-year survival rate of 14%. Anti-angiogenic therapy through anti-VEGF antibody administration is one of the limited therapies available. However, only a subgroup of metastases uses sprouting angiogenesis to secure their nutrients and oxygen supply, while others rely on vessel co-option (VCO). The distinct mode of vascularization is reflected by specific histopathological growth patterns (HGPs), which have proven prognostic and predictive significance. Nevertheless, their molecular mechanisms are poorly understood.MethodsWe evaluated CRCLM from 225 patients regarding their HGP and clinical data. Moreover, we performed spatial (21,804 spots) and single-cell (22,419 cells) RNA sequencing analyses to explore molecular differences in detail, further validated in vitro through immunohistochemical analysis and patient-derived organoid cultures.ResultsWe detected specific metabolic alterations and a signature of WNT signalling activation in metastatic cancer cells related to the VCO phenotype. Importantly, in the corresponding healthy liver of CRCLM displaying sprouting angiogenesis, we identified a predominantly expressed capillary subtype of endothelial cells, which could be further explored as a possible predictor for HGP relying on sprouting angiogenesis.ConclusionThese findings may prove to be novel therapeutic targets to the treatment of CRCLM, in special the ones relying on VCO.

Histopathological Growth Pattern in Colorectal Liver Metastasis and The Tumor Immune Microenvironment

Almost half of all patients with colorectal cancer present with or eventually develop metastasis, most frequently in the liver. Understanding the histopathological growth patterns and tumor immune microenvironment of colorectal liver metastases may help determine treatment strategies and assess prognosis. A literature search was conducted to gather information on cancer biology, histopathological growth patterns, and the tumor immune microenvironment in colorectal liver metastases, including their mechanisms and their impact on clinical outcomes. A first consensus on histopathological growth patterns emerged in 2017, identifying five growth patterns. Later studies found benefits from a two-tier system, desmoplastic and non-desmoplastic, incorporated into the updated 2022 consensus. Furthermore, the tumor immune microenvironment shows additional characteristic features with relevance to cancer biology. This includes density of T-cells (CD8+), expression of claudin-2, presence of vessel co-option versus angiogenesis, as well as several other factors. The relation between histopathological growth patterns and the tumor immune microenvironment delineates distinct subtypes of cancer biology. The distinct subtypes are found to correlate with risk of metastasis or relapse, and hence to clinical outcome and long-term survival in each patient. In order to optimize personalized and precision therapy for patients with colorectal liver metastases, further investigation into the mechanisms of cancer biology and their translational aspects to novel treatment targets is warranted.

Development of Tumor-Vasculature Interaction on Chip Mimicking Vessel Co-Option of Glioblastoma

Development of Tumor-Vasculature Interaction on Chip Mimicking Vessel Co-Option of Glioblastoma

The molecular mechanisms underlying neutrophil infiltration in vessel co-opting colorectal cancer liver metastases.

Colorectal cancer liver metastases (CRCLMs) have two major histopathological growth patterns (HGPs): desmoplastic (DHGP) and replacement (RHGP). The DHGP tumours derive their vasculature by angiogenesis, while the RHGP tumours use vessel co-option. Various studies have associated RHGP tumours with an unfavourable prognosis, as well as high levels of resistance to anti-angiogenic agents and chemotherapy. Recently, we reported higher numbers of neutrophils in the tumour microenvironment (TME) of vessel co-opting tumours compared to their angiogenic counterparts. However, the molecular mechanisms underlying this phenotype are unclear. Herein, we suggested a positive correlation between the expression of angiopoietin-1 (Ang1) in the hepatocytes and the presence of neutrophils in vessel co-opting tumours. Importantly, upregulation of Ang1 in the hepatocytes is associated with the presence of runt-related transcription factor-1 (RUNX1) in the neighboring cancer cells in vitro and in vivo. Altogether, our data suggest the molecular mechanisms by which neutrophils are infiltrated in vessel co-opting CRCLM lesions. This finding may yield novel therapeutic strategies for CRCLM patients in future.

Hematogenous metastasis and tumor dormancy as concepts or dogma? The continuum of vessel co-option and angiotropic extravascular migratory metastasis as an alternative.

It has been accepted for many years that tumor cells spread via the circulation to distant sites. The latency period between treatment and tumor recurrence has been attributed to dormant cells in distant organs that emerge and grow as metastatic tumors. These processes are accepted with an incomplete demonstration of their existence. Challenging such a well-established accepted paradigm is not easy as history as shown. An alternative or co-existing mechanism involving tumor cell migration along the outside of the vessels and co-option of the blood vessel has been studied for over 25 years and is presented. Several lines of data support this new mechanism of tumor spread and metastatic growth and is termed angiotropic extravascular migratory metastasis or EVMM. This slow migration along the outside of the vessel wall may explain the latency period between treatment and metastatic tumor growth. The reader is asked to be open to this possible new concept in how tumors spread and grow and the reason for this latency period. A full understanding of how tumors spread and grow is fundamental for the targeting of new therapeutics.

Targeting FAPα-expressing hepatic stellate cells overcomes resistance to antiangiogenics in colorectal cancer liver metastasis models.

Vessel co-option has been demonstrated to mediate colorectal cancer liver metastasis (CRCLM) resistance to antiangiogenic therapy. The current mechanisms underlying vessel co-option have mainly focused on “hijacker” tumor cells, whereas the function of the “hijackee” sinusoidal blood vessels has not been explored. Here, we found that the occurrence of vessel co-option in bevacizumab-resistant CRCLM xenografts was associated with increased expression of fibroblast activation protein α (FAPα) in the co-opted hepatic stellate cells (HSCs), which was dramatically attenuated in HSC-specific conditional Fap-knockout mice bearing CRCLM allografts. Mechanistically, bevacizumab treatment induced hypoxia to upregulate the expression of fibroblast growth factor–binding protein 1 (FGFBP1) in tumor cells. Gain- or loss-of-function experiments revealed that the bevacizumab-resistant tumor cell–derived FGFBP1 induced FAPα expression by enhancing the paracrine FGF2/FGFR1/ERK1/-2/EGR1 signaling pathway in HSCs. FAPα promoted CXCL5 secretion in HSCs, which activated CXCR2 to promote the epithelial-mesenchymal transition of tumor cells and the recruitment of myeloid-derived suppressor cells. These findings were further validated in tumor tissues derived from patients with CRCLM. Targeting FAPα+ HSCs effectively disrupted the co-opted sinusoidal blood vessels and overcame bevacizumab resistance. Our study highlights the role of FAPα+ HSCs in vessel co-option and provides an effective strategy to overcome the vessel co-option–mediated bevacizumab resistance.

New insights into antiangiogenic therapy resistance in cancer: Mechanisms and therapeutic aspects.

Angiogenesis is a hallmark of cancer and is required for tumor growth and progression. Antiangiogenic therapy has been revolutionarily developing and was approved for the treatment of various types of cancer for nearly two decades, among which bevacizumab and sorafenib continue to be the two most frequently used antiangiogenic drugs. Although antiangiogenic therapy has brought substantial survival benefits to many cancer patients, resistance to antiangiogenic drugs frequently occurs during clinical treatment, leading to poor outcomes and treatment failure. Cumulative evidence has demonstrated that the intricate interplay among tumor cells, bone marrow-derived cells, and local stromal cells critically allows for tumor escape from antiangiogenic therapy. Currently, drug resistance has become the main challenge that hinders the therapeutic efficacies of antiangiogenic therapy. In this review, we describe and summarize the cellular and molecular mechanisms conferring tumor drug resistance to antiangiogenic therapy, which was predominantly associated with redundancy in angiogenic signaling molecules (e.g., VEGFs, GM-CSF, G-CSF, and IL17), alterations in biological processes of tumor cells (e.g., tumor invasiveness and metastasis, stemness, autophagy, metabolic reprogramming, vessel co-option, and vasculogenic mimicry), increased recruitment of bone marrow-derived cells (e.g., myeloid-derived suppressive cells, tumor-associated macrophages, and tumor-associated neutrophils), and changes in the biological functions and features of local stromal cells (e.g., pericytes, cancer-associated fibroblasts, and endothelial cells). We also review potential biomarkers to predict the response to antiangiogenic therapy in cancer patients, which mainly consist of imaging biomarkers, cellular and extracellular proteins, a certain type of bone marrow-derived cells, local stromal cell content (e.g., pericyte coverage) as well as serum or plasma biomarkers (e.g., non-coding RNAs). Finally, we highlight the recent advances in combination strategies with the aim of enhancing the response to antiangiogenic therapy in cancer patients and mouse models. This review introduces a comprehensive understanding of the mechanisms and biomarkers associated with the evasion of antiangiogenic therapy in cancer, providing an outlook for developing more effective approaches to promote the therapeutic efficacy of antiangiogenic therapy.

Tumor vessel co-option: The past & the future.

Tumor vessel co-option (VCO) is a non-angiogenic vascularization mechanism that is a possible cause of resistance to anti-angiogenic therapy (AAT). Multiple tumors are hypothesized to primarily rely on growth factor signaling-induced sprouting angiogenesis, which is often inhibited during AAT. During VCO however, tumors invade healthy tissues by hijacking pre-existing blood vessels of the host organ to secure their blood and nutrient supply. Although VCO has been described in the context of AAT resistance, the molecular mechanisms underlying this process and the profile and characteristics of co-opted vascular cell types (endothelial cells (ECs) and pericytes) remain poorly understood, resulting in the lack of therapeutic strategies to inhibit VCO (and to overcome AAT resistance). In the past few years, novel next-generation technologies (such as single-cell RNA sequencing) have emerged and revolutionized the way of analyzing and understanding cancer biology. While most studies utilizing single-cell RNA sequencing with focus on cancer vascularization have centered around ECs during sprouting angiogenesis, we propose that this and other novel technologies can be used in future investigations to shed light on tumor EC biology during VCO. In this review, we summarize the molecular mechanisms driving VCO known to date and introduce the models used to study this phenomenon to date. We highlight VCO studies that recently emerged using sequencing approaches and propose how these and other novel state-of-the-art methods can be used in the future to further explore ECs and other cell types in the VCO process and to identify potential vulnerabilities in tumors relying on VCO. A better understanding of VCO by using novel approaches could provide new answers to the many open questions, and thus pave the way to develop new strategies to control and target tumor vascularization.

Abstract 1580: Cancer cells promote phenotypic alterations in adjacent hepatocytes to facilitate vessel co-option in colorectal cancer liver metastasis

Abstract Vessel co-option is associated with resistance against anti-angiogenic therapy in colorectal cancer liver metastases (CRCLM). Vessel co-opted lesions are characterized by highly motile cancer cells that move toward and along the pre-existing vessels in the surrounding non-malignant tissue and hijack them to gain access to nutrient. In order to access the sinusoidal vessels, the cancer cells must displace the hepatocytes and occupy their space. However, the mechanisms underlying this displacement are unknown. Herein, we examined the involvement of apoptosis, motility, epithelial-mesenchymal transition (EMT) and autophagy pathways in hepatocyte’s displacement by cancer cells. We demonstrated that cancer cells induce the expression of the proteins that associated with upregulation of apoptosis, motility and EMT in adjacent hepatocytes in vitro and in vivo. Accordingly, we observed upregulation of cleaved caspase-3, cleaved poly (ADP-ribose) polymerase-1 (PARP-1) and Actin Related Protein 2/3 (ARP2/3) in adjacent hepatocytes to cancer cells, while we noticed lower expression levels of E-cadherin. Importantly, Runt-related transcription factor 1 (RUNX1) knockdown in the cancer cells has impaired their function towards adjacent hepatocytes in vitro and in vivo. Altogether, our data indicating that cancer cells may exploit various mechanisms to displace the hepatocytes and access to the sinusoidal vessels to establish vessel co-option. Citation Format: Miran Rada, Anthoula Lazaris, Peter Metrakos. Cancer cells promote phenotypic alterations in adjacent hepatocytes to facilitate vessel co-option in colorectal cancer liver metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1580.

Angiopoietin-1 Upregulates Cancer Cell Motility in Colorectal Cancer Liver Metastases through Actin-Related Protein 2/3.

Simple SummaryVessel co-option has been recognized as a mechanism associated with resistance to anti-angiogenic treatment in colorectal cancer liver metastases (CRCLMs). Recently, we reported that Angiopoietin-1 (Ang1) stimulates vessel co-option in CRCLM through an unknown mechanism. In this manuscript, we found the molecular pathways that mediate the function of Ang1 in CRCLM. We showed that Ang1 induces the expression levels of actin-related protein 2/3 (ARP2/3) in the cancer cells via various mechanisms. Importantly, different studies have shown that high levels of ARP2/3 in cancer cells are essential for the formation of vessel co-opting CRCLM tumours. Highlighting these pathways is an important step to identify therapeutic strategies to overcome vessel co-option and resistance to anti-angiogenic therapy in CRCLM.Resistance to anti-angiogenic therapy is a major challenge in the treatment of colorectal cancer liver metastases (CRCLMs). Vessel co-option has been identified as a key contributor to anti-angiogenic therapy resistance in CRCLMs. Recently, we identified a positive correlation between the expression of Angiopoietin1 (Ang1) in the liver and the development of vessel co-opting CRCLM lesions in vivo. However, the mechanisms underlying its stimulation of vessel co-option are unclear. Herein, we demonstrated Ang1 as a positive regulator of actin-related protein 2/3 (ARP2/3) expression in cancer cells, in vitro and in vivo, which is known to be essential for the formation of vessel co-option in CRCLM. Significantly, Ang1-dependent ARP2/3 expression was impaired in the cancer cells upon Tie2 or PI3K/AKT inhibition in vitro. Taken together, our results suggest novel mechanisms by which Ang1 confers the development of vessel co-option in CRCLM, which, targeting this pathway, may serve as promising therapeutic targets to overcome the development of vessel co-option in CRCLM.

Cancer Cells Promote Phenotypic Alterations in Hepatocytes at the Edge of Cancer Cell Nests to Facilitate Vessel Co-Option Establishment in Colorectal Cancer Liver Metastases.

Simple SummaryTumour cells in colorectal cancer liver metastases (CRCLM) obtain their blood supply via two major mechanisms: (i) sprouting angiogenesis, through the generation of new vessels; (ii) vessel co-option, where the cancer cells hijack the pre-existing vasculature. The current treatment for CRCLM targets angiogenesis; however, these treatments are ineffective on cancer cells utilizing vessel co-option to gain their blood supply. Our study suggests that cancer cells stimulate phenotypic alterations in the cells of surrounding liver tissue (hepatocytes) in vessel co-opting lesions. These modifications facilitate cancer cells to infiltrate through the liver tissue and hijack the pre-existing vasculature to obtain oxygen and nutrients.Vessel co-option is correlated with resistance against anti-angiogenic therapy in colorectal cancer liver metastases (CRCLM). Vessel co-opting lesions are characterized by highly motile cancer cells that move toward and along the pre-existing vessels in the surrounding nonmalignant tissue and co-opt them to gain access to nutrients. To access the sinusoidal vessels, the cancer cells in vessel co-opting lesions must displace the hepatocytes and occupy their space. However, the mechanisms underlying this displacement are unknown. Herein, we examined the involvement of apoptosis, autophagy, motility, and epithelial–mesenchymal transition (EMT) pathways in hepatocyte displacement by cancer cells. We demonstrate that cancer cells induce the expression of the proteins that are associated with the upregulation of apoptosis, motility, and EMT in adjacent hepatocytes in vitro and in vivo. Accordingly, we observe the upregulation of cleaved caspase-3, cleaved poly (ADP-ribose) polymerase-1 (PARP-1) and actin-related protein 2/3 (ARP2/3) in adjacent hepatocytes to cancer cell nests, while we notice a downregulation of E-cadherin. Importantly, the knockdown of runt-related transcription factor 1 (RUNX1) in cancer cells attenuates the function of cancer cells in hepatocytes alterations in vitro and in vivo. Altogether, our data suggest that cancer cells exploit various mechanisms to displace hepatocytes and access the sinusoidal vessels to establish vessel co-option.

Vessel co-option and angiotropic extravascular migratory metastasis: a continuum of tumour growth and spread?

Two fields of cancer research have emerged dealing with the biology of tumour cells localised to the abluminal vascular surface: vessel co-option (VCo), a non-angiogenic mode of tumour growth and angiotropic extravascular migratory metastasis (EVMM), a non-hematogenous mode of tumour migration and metastasis. VCo is a mechanism by which tumour cells gain access to a blood supply by spreading along existing blood vessels in order to grow locally. Angiotropic EVMM involves "pericytic mimicry" (PM), which is characterised by tumour cells continuously migrating in the place of pericytes distantly along abluminal vascular surfaces. When cancer cells are engaged in PM and EVMM, they migrate along blood vessels beyond the advancing front of the tumour to secondary sites with the formation of regional and distant metastases. In the present perspective, the authors review the current scientific literature, emphasising the analogies between embryogenesis and cancer progression, the re-activation of embryonic signals by "cancer stem cells", and the important role of laminins and epithelial-mesenchymal-transition. This perspective maintains that VCo and angiotropic EVMM constitute complementary processes and represent a continuum of cancer progression from the primary tumour to metastases and of tumour growth to EVMM, analogous to the embryonic development program.

A novel second near-infrared theranostic agent: a win–win strategy of tracing and blocking tumor-associated vessels for oral squamous cell carcinoma

Tumor vessel co-option is a crucial predictor for tumor invasiveness but is easy to ignore in tumor vascular targeting therapy. A high density of tumor vessel co-option located alongside the tumor frontier predicted a high tendency of tumor invasion and regional metastasis. Herein, a novel second near-infrared (NIR-II) nanoagent (Coi8DFIC-sorafenib nanoparticles, CS NPs [CS NPs]) was constructed with an organic Coi8DFIC dye and sorafenib, which demonstrated high tissue penetration and low tissue autofluorescence, enabling imaging of vessel co-option and tumor micrometastasis. This nanotherapeutic agent displayed considerably improved quantum yield of fluorescence (0.89%), high reactive oxygen species (ROS) generation, and fairly good biosafety in vivo. Compared with indocyanine green (ICG), CS NPs exhibited better photostability and photothermal conversion efficiency. By tracking tumor-associated vessels and real-time tumor imaging, CS NPs could open/stop vascular targeting therapy by laser on/off. The combination of vessel disruption and imaging-guided photothermal therapy/photodynamic therapy provided a win–win strategy for oral squamous cell carcinoma.

The Presence of Vessel Co-option Determines Overall Survival in Patients with Colorectal Cancer Liver Metastasis

Introduction: Colorectal cancer liver metastasis (CRCLM) remains a lethal diagnosis with an overall 5-year survival rate of 5-10%. Two distinct histopathological growth patterns (HGPs) of CRCLM are known to have significantly different rates of patient survival, and response to treatment. We set out to review the results of 275 patients who underwent liver resection for CRCLM at the McGill University Health Center (MUHC) and analyze their clinical outcome, mutational burden, and pattern of cancer progression in light of their HGP’s, and to consider their potential effect on surgical decision making. Methods: We performed a retrospective multivariant analysis on clinical data from patients with CRCLM (n=275) who underwent liver resection at the McGill University Health Center (MUHC). All tumors were scored using international consensus guidelines by pathologists trained in HGP scoring. Results: 107 patients (43.7%) were angiogenic and 138 patients (56.3%) were vessel co-opting. The 3 and 5-year survival rates for angiogenic patients compared to vessel co-opting patients were 54% and 34.6% vs 47.1% and 13% respectively (p<0.0001). Multivariate analysis showed patients with vessel co-opting CRCLM had a higher incidence of extrahepatic metastatic disease (p=0.0215) compared to angiogenic CRCLM. Additionally, KRAS mutation status was a marker of an increased likelihood of disease recurrence (p=0.0434) as was increased number of liver tumors (p=0.0071) and multiple sites of extrahepatic metastatic disease (p<0.0001). Conclusions: Multivariate analysis identified key clinical features correlating to the two HGPs. Determining liver tumor HGPs is essential for patient prognostication and treatment optimization.

Matrix Hyaluronic Acid and Hypoxia Influence a CD133+ Subset of Patient-Derived Glioblastoma Cells.

Glioblastoma (GBM) displays diffusive invasion throughout the brain microenvironment, which is partially responsible for its short median survival rate (<15 months). Stem-like subpopulations (GBM stem-like cells, GSCs) are believed to play a central role in therapeutic resistance and poor patient prognosis. Given the extensive tissue remodeling and processes such as vessel co-option and regression that occur in the tumor microenvironment, it is essential to understand the role of metabolic constraint such as hypoxia on GBM cell populations. This work describes the use of a multidimensional gelatin hydrogel to culture patient-derived GBM cells, to evaluate the influence of hypoxia and the inclusion brain-mimetic hyaluronic acid on the relative activity of GSCs versus overall GBM cells. Notably, CD133+ GBM cell fraction is crucial for robust formation of tumor spheroids in multidimensional cultures. In addition, while the relative size of the CD133+ GBM subpopulation increased in response to both hypoxia and matrix-bound hyaluronan, we did not observe cell subtype-specific changes in invasion signaling pathway activation. Taken together, this study highlights the potential of biomimetic culture systems for resolving changes in the population dynamics and behavior of subsets of GBM specimens for the future development of precision medicine applications. Impact Statement This study describes a gelatin hydrogel platform to investigate the role of extracellular hyaluronic acid and hypoxia on the behavior of a CD133+ subset of cells within patient-derived glioblastoma (GBM) specimens. We report that the relative expansion of the CD133+ GBM stem cell-like population is strongly responsive to extracellular cues, highlighting the significance of biomimetic hydrogel models of the tumor microenvironment to investigate invasion and therapeutic response.

The emerging roles of circular RNAs in vessel co-option and vasculogenic mimicry: clinical insights for anti-angiogenic therapy in cancers.

Unexpected resistance to anti-angiogenic treatment prompted the investigation of non-angiogenic tumor processes. Vessel co-option (VC) and vasculogenic mimicry (VM) are recognized as primary non-angiogenic mechanisms. In VC, cancer cells utilize pre-existing blood vessels for support, whereas in VM, cancer cells channel and provide blood flow to rapidly growing tumors. Both processes have been implicated in the development of tumor and resistance to anti-angiogenic drugs in many tumor types. The morphology, but rare molecular alterations have been investigated in VC and VM. There is a pressing need to better understand the underlying cellular and molecular mechanisms. Here, we review the emerging circular RNA (circRNA)-mediated regulation of non-angiogenic processes, VC and VM.

P14.32 Patterns of invasion in Glioblastoma have unique radiological features, varied response to radiotherapy and long term outcomes

Abstract BACKGROUND To assess the correlation of the three patterns of invasion of Glioblastoma (GB) with the respective MRI features, response to radiotherapy and disease free survival MATERIAL AND METHODS Histopathology of 62 patients with Glioblastoma who had undergone maximal safe resection (MSR)/ stereotactic biopsy (STB) &amp; referred for adjuvant therapy was reviewed to assess the pattern of invasion. Three patterns were observed- single cell infiltration (pattern 1), perineuronal satellitosis (pattern 2) and vessel cooption (or perivascular spread, pattern 3), majority also had diffuse infiltration pattern in the background. The pre and post operative MRI scans were utilized for RT planning of 3DCRT/ IMRT/ VMAT, the modality decided by the treating oncologist. RT volumes were as per ESTRO-ACROP guidelines. Duration of radiotherapy- the Stupp regimen or a short course of hypofractionated radiotherapy (HFRT) (35Gy/10Fr- planned gap- completion of total dose) was based on the extent of resection (EOR) and KPS at the time of RT. Correlation of the pattern of invasion, MRI characteristics of each on the response to radiotherapy as assessed by RANO criteria and survival was analysed with the help of SPSS V21. RESULTS MRI findings after stratification of 62 patients, median age 43 years(18–70), M:F::42:20) into type 1:17, type 2:21 and type 3:24- patterns of invasion, were cortical tumours (subtle enhancement) close to SVZ, solid (+/- cystic) lesion in the eloquent cortex or large mass effect crossing the barriers respectively. OS of entire cohort at 1 year was 75%, with significant differences among groups (82% vs. 85% vs. 50%). Among pattern 1, those patients with IDH mutant (20%) &amp; mean dose to C/L SVZ of 30Gy or more had longer DFS at 1 year (89% Vs. 38% &amp; 85% vs. 0%, p&amp;lt;0.05). In pattern 2, EOR (gross total or near total resection Vs. only STB) showed a better DFS at 1 year (90% vs. 40%, p=0.095). In patients with pattern 3, 40% had post op KPS of &amp;lt;80 and 20% of them underwent HFRT who showed higher CR/PR rates compared to the conventional RT. HFRT in low KPS with m-MGMT showed higher local control at 6 months (100% Vs. 66%, p= 0.02). CONCLUSION Glioblastoma has three patterns of invasion with unique MRI features and aggressiveness for each. Identification of this may guide choice of adjuvant therapy, patterns of failure and therapy resistance

Runt related transcription factor-1 plays a central role in vessel co-option of colorectal cancer liver metastases

Colorectal cancer liver metastasis (CRCLM) has two major histopathological growth patterns: angiogenic desmoplastic and non-angiogenic replacement. The replacement lesions obtain their blood supply through vessel co-option, wherein the cancer cells hijack pre-existing blood vessels of the surrounding liver tissue. Consequentially, anti-angiogenic therapies are less efficacious in CRCLM patients with replacement lesions. However, the mechanisms which drive vessel co-option in the replacement lesions are unknown. Here, we show that Runt Related Transcription Factor-1 (RUNX1) overexpression in the cancer cells of the replacement lesions drives cancer cell motility via ARP2/3 to achieve vessel co-option. Furthermore, overexpression of RUNX1 in the cancer cells is mediated by Transforming Growth Factor Beta-1 (TGFβ1) and thrombospondin 1 (TSP1). Importantly, RUNX1 knockdown impaired the metastatic capability of colorectal cancer cells in vivo and induced the development of angiogenic lesions in liver. Our results confirm that RUNX1 may be a potential target to overcome vessel co-option in CRCLM.

Diversity in cell differentiation, histology, phenotype and vasculature of mass-forming intrahepatic cholangiocarcinomas.

Mass-forming intrahepatic cholangiocarcinomas (MF-iCCAs), involving small bile ducts, bile ductules or canals of Hering, remain treated as a single entity. We aimed to examine the diversity in histology, phenotype and tumour vasculature of MF-iCCAs. Based on morphology and immunophenotype, we classified MF-iCCAs into small bile duct (SBD), cholangiolocarcinoma (CLC), ductal plate malformation (DPM) and hepatocellular carcinoma (HCC)-like subtypes. Genetic correlations among the histological subtypes were examined by multi-region tumour sequencing. Vasculatures and other clinicopathological features were compared among tumour groups with various proportions of the histological subtypes in 62 MF-iCCAs. Cases of pure SBD, CLC, DPM and HCC-like subtypes numbered 18 (29%), seven (11.3%), none (0%) and two (3%), respectively; the remaining 35 (56.4%) cases comprised several components. Genetic alterations, isocitrate dehydrogenase (IDH)1/2, KRAS, TP53, polybromo-1 (PBRM1) and BRCA1-associated protein 1 (BAP1), were shared among SBD, CLC, DPM and hepatoid components within a tumour. We uncovered distinct vascularisation mechanisms among SBD, CLC and DPM subtypes with a prominent vessel co-option in CLC tumours. iCCA with a DPM pattern had the highest vascular densities (mean microvascular density,140/mm2 ; arterial vessel density, 18.3/mm2 ). Increased CLC component was correlated with longer overall survival time (r=0.44, P=0.006). Pure SBD tumours had a lower 5-year overall survival rate compared with MF-iCCA with CLC pattern (30.5 versus 72.4%, P=0.011). MF-iCCAs comprise four histological subtypes. Given their sharing some driver gene alterations, indicating they can have a common cell origin, SBD, CLC and DPM subtypes, however, differ in cell differentiation, histology, phenotype or tumour vasculature.