Intratumoral Microvessel Density Research Articles

Prognostic Significance of Microvessel Density and Hypoxic Markers in Canine Osteosarcoma: Insights into Angiogenesis and Tumor Aggressiveness.

Canine osteosarcoma (OSA) is an aggressive and highly malignant tumor of bone with a poor prognosis and it mirrors the disease in humans. Angiogenesis, the formation of new blood vessels, is driven by hypoxia-induced factors such as HIF-1α and VEGF, both of which play a crucial role in tumor growth and metastasis. However, the role of angiogenesis in OSA remains a topic of ongoing debate. This study aimed to investigate the relationship between angiogenesis, measured by intratumoral microvessel density (MVD), hypoxic markers, and clinical outcomes in 28 dogs diagnosed with appendicular OSA. Clinicopathological data such as age, breed distribution, tumor localization, histopathological subtypes, and metastatic behavior were consistent with reported epidemiologic characteristics of canine OSA, though no significant correlation was found among these variables. The results indicated a significant association between higher MVD and high-grade OSA (p = 0.029), suggesting that increased tumor vascularization is linked to more aggressive tumor behavior. Additionally, elevated VEGF expression was strongly correlated with disease-free interval DFI), with a p-value of 0.045. Although HIF-1α positivity showed a trend towards poorer survival, the results did not reach statistical significance (p = 0.07). These findings highlight the potential role of VEGF as a valuable prognostic marker in canine OSA, which could have potentially important implications for therapeutic targeting and clinical management of the disease. This study advances the understanding of angiogenesis in canine OSA, while emphasizing the need for continued research into the complex mechanisms regulating the interplay between hypoxia, angiogenesis and tumor progression.

Prognostic Significance of Angiogenesis in invasive ductal carcinoma

Background: Angiogenesis is essential for tumor growth and metastasis. Axillary lymphnode status has been the most important prognostic factor in operable breast carcinoma, but it does not fully account for the varied disease outcome. More accurate prognostic indicators would help in selection of patients at high risk for disease recurrence and death who are candidates for systemic adjuvant therapy. Microvessel density in invasive ductal carcinoma (measures of tumor angiogenesis) is associated with metastasis and thus may be a prognostic indicator. Objective: To correlate intratumoral and peritumoral angiogenic microvessel density with lymphnode metastasis in invasive ductal carcinoma. Material and Methods: It is a cross sectional observational study, carried out at the department of Pathology, BSMMU from January 2016 to December 2017. A total 48 mastectomy samples with axillary nodes from histologically confirmed invasive ductal carcinoma were included in this study. Weidner method was used for calculating micro vessels density. Sections examined to evaluate the density of angiogenic vessels by immuohistological stain with vWF expression in invasive breast cancer. Correlation between angiogenic vessels density with or without lymphnode metastasis was taken. Results: In this study angiogenic vessel count is more in the intratumoral area than peritumoural area. There was a positive significant correlation between lymphnode metastasis with micro vessel density in both peritumoral and intratumoral areas in Weidner method in vWF stain. J Shaheed Suhrawardy Med Coll 2023; 15(1): 59-65

Characteristics of mast cell infiltration in lung adenocarcinoma and its impact on prognosis

BackgroundThe role of mast cells in malignancies remains unclear, and there is no clear correlation between mast cells and tumor microvessels, tumor growth, or lung adenocarcinoma (LUAD) prognosis. This study aims to explore the association between mast cell density (MCD) and intratumoral microvessel density (MVD), clinicopathological parameters, and prognosis in LUAD, by evaluating mast cell infiltration characteristics and their prognostic significance.MethodsThis retrospective investigation involved 238 patients with LUAD undergoing complete resection. Tumor and normal lung tissue sections outside the tumor were immunohistochemically stained for MCD in the intratumoral and outside regions, respectively. CD34 polyclonal antibody was used to measure intratumoral MVD.ResultsIntratumoral regions of LUAD had a higher MCD (P < 0.001) than normal lung tissue. In the intratumoral region, MCD and CD34-MVD were positively correlated (r = 0.411, P < 0.001). Intratumoral MCD correlated with sex, smoking history, tumor differentiation, pathological subtype, and tumor size. Female sex (P = 0.012), no smoking history (P = 0.002), acinar predominant type (P = 0.012), and tumor size ≤ 3 cm (P = 0.009) were associated with a higher MCD, whereas poorly differentiated (P = 0.039) and solid/micropapillary predominant types (P = 0.001) were associated with a lower MCD. Higher intratumoral MCD exhibited a marginally improved overall survival, and individuals with higher MCD infiltration ratios (intratumoral MCD/outside the MCD) had higher disease-free and overall survival rates (log-rank P < 0.001). A high MCD infiltration ratio was associated with decreased risk of tumor progression and death following complete resection.ConclusionThe tumor microenvironment controls mast cell infiltration in LUAD, and patients with increased intratumoral mast cell infiltration have better prognosis.

Emerging Treatments Targeting the Tumor Microenvironment for Advanced Chondrosarcoma.

Chondrosarcoma (ChS), a malignant cartilage-producing tumor, is the second most frequently diagnosed osseous sarcoma after osteosarcoma. It represents a very heterogeneous group of malignant chemo- and radiation-resistant neoplasms, accounting for approximately 20% of all bone sarcomas. The majority of ChS patients have a good prognosis after a complete surgical resection, as these tumors grow slowly and rarely metastasize. Conversely, patients with inoperable disease, due to the tumor location, size, or metastases, represent a great clinical challenge. Despite several genetic and epigenetic alterations that have been described in distinct ChS subtypes, very few therapeutic options are currently available for ChS patients. Therefore, new prognostic factors for tumor progression as well as new treatment options have to be explored, especially for patients with unresectable or metastatic disease. Recent studies have shown that a correlation between immune infiltrate composition, tumor aggressiveness, and survival does exist in ChS patients. In addition, the intra-tumor microvessel density has been proven to be associated with aggressive clinical behavior and a high metastatic potential in ChS. This review will provide an insight into the ChS microenvironment, since immunotherapy and antiangiogenic agents are emerging as interesting therapeutic options for ChS patients.

Super-Resolution Ultrasound Imaging for Monitoring the Therapeutic Efficacy of a Vascular Disrupting Agent in an Animal Model of Breast Cancer.

Evaluate the use of super-resolution ultrasound (SRUS) imaging for the early detection of tumor response to treatment using a vascular-disrupting agent (VDA). A population of 28 female nude athymic mice (Charles River Laboratories) were implanted with human breast cancer cells (MDA-MB-231, ATCC) in the mammary fat pad and allowed to grow. Ultrasound imaging was performed using a Vevo 3100 scanner (FUJIFILM VisualSonics Inc) equipped with the MX250 linear array transducer immediately before and after receiving bolus injections of a microbubble (MB) contrast agent (Definity, Lantheus Medical Imaging) via the tail vein. Following baseline ultrasound imaging, VDA drug (combretastatin A4 phosphate, CA4P, Sigma Aldrich) or control saline was injected via the placed catheter. After 4 or 24 hours, repeat ultrasound imaging along the same tumor cross-section occurred. Direct intratumoral pressure measurements were obtained using a calibrated sensor. All raw ultrasound data were saved for offline processing and SRUS image reconstruction using custom MATLAB software (MathWorks Inc). From a region encompassing the tumor space and the entire postprocessed ultrasound image sequence, time MB count (TMC) curves were generated in addition to traditional SRUS maps reflecting MB enumeration at each pixel location. Peak enhancement (PE) and wash-in rate (WIR) were extracted from these TMC curves. At termination, intratumoral microvessel density (MVD) was quantified using tomato lectin labeling of patent blood vessels. SRUS images exhibited a clear difference between control and treated tumors. While there was no difference in any group parameters at baseline (0 hour, P > .09), both SRUS-derived PE and WIR measurements in tumors treated with VDA exhibited significant decreases by 4 (P = .03 and P = .05, respectively) and 24 hours (P = .02 and P = .01, respectively), but not in control group tumors (P > .22). Similarly, SRUS derived microvascular maps were not different at baseline (P = .81), but measures of vessel density were lower in treated tumors at both 4 and 24 hours (P < .04). An inverse relationship between intratumoral pressure and both PE and WIR parameters were found in control tumors (R2 > .09, P < .03). SRUS imaging is a new modality for assessing tumor response to treatment using a VDA.

N-myc downstream–regulated gene 1 can promote vasculogenic mimicry and angiogenesis in urothelial carcinoma

Urothelial carcinoma (UC) of the bladder is a common cause of cancer-related death worldwide. Vasculogenic mimicry (VM) is a process by which the malignant cells can generate vascular-like structures formed of periodic acid–Schiff (PAS) positive/CD31 negative extracellular matrix independent of angiogenesis and thus promotes tumor progression. N-myc downstream–regulated gene 1 (NDRG1) is a protein that can modulate tumor angiogenesis; however, its role in regulating tumor angiogenesis and VM formation has not been previously investigated in UC. This study aims to evaluate the role of intra-tumor microvessel density (MVD) (as a surrogate measure of angiogenesis), VM, and NDRG1 in UC and their correlation with different clinicopathologic features, then assess the correlation between them in UC. Sixty specimens of UC of the bladder were included. PAS-CD31 immunohistochemical double staining method was used to evaluate the intra-tumor MVD and VM. Immunohistochemical expression of NDRG1 was also examined. VM and NDRG1 expression were detected in 41.7% and 83.3% of UC specimens respectively. The mean of intra-tumor MVD, VM area, and NDRG1 was significantly higher in tumors with higher grade, lymphovascular invasion, and higher T stage. NDRG1 expression was positively correlated with MVD and VM. We can suggest that MVD, VM, and NDRG1 may serve as poor prognostic markers for UC. The positive correlation between NDRG1 and both MVD and VM may provide the first evidence that NDRG1 can induce tumor angiogenesis and VM in UC which may offer a novel pathway for further therapeutic strategies.

Ultrasound Microvessel Visualization in Cervical Cancer: Association Between Novel Ultrasound Techniques and Histologic Microvessel Densities.

The aim of the work described here was to evaluate the feasibility of superb microvascular imaging (SMI) and vascular endothelial growth factor receptor 2 (VEGFR2)-targeted microbubble (MBVEGFR2)-based ultrasound molecular imaging (USMI) for visualizing microvessels in cervical cancer. Hela cells were used to establish subcutaneous cervical cancer models. SMI and MBVEGFR2-based USMI were performed, and the results were compared with intratumoral microvessel density (MVD) in four groups based on tumor diameter (<3 mm, 3-5 mm, 5-7 mm and ≥7 mm). The vascularization index (VI, %) was evaluated for SMI, and the normalized intensity difference (NID) for USMI. Tumors with diameters ranging from 3 to 5 mm had the highest VI (39.07 ± 1.58) in SMI, and VI significantly decreased with increasing tumor size (all p values <0.001). The strongest signal intensity was observed in very early tumors (d < 3 mm: 43.80 ± 3.58%) after MBVEGFR2 administration; the NID gradually decreased with increasing diameter of tumors (all p values=0.007). However, no significant differences were observed in NID after administration of non-targeted (control) microbubbles (MBCon) (all p values=0.125). MBVEGFR2-based USMI had the strongest correlation with MVD in displaying microvessels of cervical cancer compared with SMI and MBCon (R2=0.78 vs. R2=0.40 and R2=0.38). These findings validate the superiority and accuracy of MBVEGFR2-based USMI for microvessel imaging and monitoring of angiogenesis in cervical cancer compared with SMI and MBCon. Nonetheless, SMI remains an alternative to microvessel imaging when ultrasonic contrast agent use is contraindicated.

Immunohistochemical Assessment of Microvessel Density in OSCC: Spatial Heterogeneity of Angiogenesis and Its Impact on Survival.

(1) Background Oral squamous cell carcinomas (OSCC) are a common malignancy of the oral cavity and are often diagnosed when they have already spread to the regional lymph nodes. Advanced stages of cancer are characterized by the development of distant metastases. Angiogenesis, a hallmark of cancer, is known to contribute to cancer progression and metastasis. High microvessel density (MVD) has been linked to poor clinical outcomes in various types of cancer. (2) Methods: In this study, we aimed to investigate the spatial heterogeneity of blood vessels by comparing the tumor center and invasion front and to evaluate its prognostic value in OSCC. A total of 71 OSCC patient specimens were collected. The tissue was immunohistochemically stained using CD31 antibody to assess the MVD in the tumor center and the invasion front. Furthermore, the associations between the histopathological parameters, including MVD, disease-free survival (DFS), and overall survival (OS) were computed. (3) Results: In our study, we found a significantly higher presence of blood vessels at the invasion front of OSCCs compared to the tumor center. However, we did not observe any significant differences in MVD between different tumor stages. High intratumoral MVD was shown to be a positive prognostic factor for DFS (p = 0.047). (4) Conclusions: To the best of our knowledge, we were the first to analyze MVD as a prognostic factor by considering its spatial heterogeneity in OSCC. However, further studies are warranted to further elucidate the complexity of microvascular spatial heterogeneity and its influence on prognosis.

Podophyllotoxin-loaded PEGylated E-selectin peptide conjugate targeted cancer site to enhance tumor inhibition and reduce side effect

E-selectin, which is highly expressed in vascular endothelial cells near tumor and get involved in the all tumor growth steps: occurrence, proliferation and metastasis, is considered as a promise targeted protein for antitumor drug discovery. Herein, we would like to report the design, preparation and the anticancer evaluation of the peptide-PEG-podophyllotoxin conjugate(PEG-Pep-PODO), in which the short peptide (CIELLQAR) was used as the E-selectin ligand for the targeting purpose and the PEG portion the molecule got the conjugate self-assembled to form a water soluble nanoparticle. In vitro release study showed that the conjugated and entrapped PODO could be released simultaneously in the presence of GSH (highly expressed in tumor environmental conditions) and the GSH would catalyze the break of the disufur bond which linked of the PODO and the peptide-PEG portion of the conjugate. Cell adhesion test of the PEG-Pep-PODO indicated that E-selectin ligand peptide CIELLQAR could get specifically and efficiently binding to the E-selectin expressing human umbilical vein endothelial cells (HUVEC). In vitro cytotoxicity assay further revealed that PEG-Pep-PODO significantly improved the selectivity of PEG-Pep-PODO for killing the tumor cells and normal cells compared with PODO solution formulation. More importantly, the in vivo experiment demonstrated that the conjugate would accumulate of the PODO payload in tumor through targeting endothelial cells in the tumor microenvironment, which resulted in the much improved in vivo inhibition of tumor growth, intratumoral microvessel density, and decreased systemic toxicity of this nanoparticle over the free PODO. Furthermore, this water soluble conjugate greatly improved the pharmacokinetic properties of the mother molecule.

Targeted Inhibition of the HGF/c-Met Pathway by Merestinib Augments the Effects of Albumin-Bound Paclitaxel in Gastric Cancer

Background and Hypothesis: Combination chemotherapy regimens are commonly used to treat gastric adenocarcinoma (GAC), but the median survival time remains less than one year. Nab-paclitaxel has demonstrated high antitumor activity in previous GAC studies. Many growth factors and their receptors are overexpressed in GAC and have been implicated in its pathophysiology. We hypothesize that merestinib, a small-molecule inhibitor targeting c-Met, Axl, and DDR1/2 pathways, will have significant antitumor effects and will enhance the response to nab-paclitaxel in GAC preclinical models.&#x0D; Project Methods: In vitro proliferation and protein expression were assessed using WST-1 and immunoblot assays. Subcutaneous xenografts of MKN-45 and SNU-1 cell lines were implanted in mice to study tumor growth inhibition. Immunohistochemistry was performed to examine intratumor proliferation and microvessel density.&#x0D; Results: In vitro assays showed that nab-paclitaxel and merestinib decreased cell proliferation in all three cell lines, with an additive effect in combination. Reduction in cell proliferation at low doses of nab-paclitaxel (10 nM), merestinib (100 nM), and their combination was 87%, 82%, and 94% (MKN-45 cell line, high phospho-c-Met expression), 59%, 50%, and 82% (SNU-1 cell line, low phospho-c-Met expression), and 53%, 19%, and 66% in gastric fibroblasts. Immunoblot analysis of merestinib treated MKN-45 cells revealed increased expression of apoptotic proteins and decreased expression of phospho-c-Met, phospho-EGFR, phospho-IGF-1R, phospho-ERK, and phospho-AKT. In gastric fibroblasts, merestinib decreased phospho-ERK and increased apoptotic protein expression. Phospho-c-Met and phospho-EGFR were not detected in SNU-1 immunoblots; however, phospho-ERK, phospho-VEGFR, and apoptotic protein expression increased after treatment. In MKN-45 xenografts, net tumor growth in control, nab-paclitaxel, merestinib, and combination groups was 503 mm3, 115 mm3, 91 mm3, and -9.7 mm3. Immunohistochemistry analysis of tumor cell proliferation and microvessel density corroborated tumor growth study results.&#x0D; Conclusion: The data suggest that merestinib in combination with nab-paclitaxel carry a promising potential for improving clinical GAC therapy.

The Vessels That Encapsulate Tumor Clusters (VETC) Pattern Is a Poor Prognosis Factor in Patients with Hepatocellular Carcinoma: An Analysis of Microvessel Density.

The outcomes of patients with hepatocellular carcinoma (HCC) are unsatisfactory because of its high recurrence rate. The Vessels that encapsulate tumor clusters (VETC) pattern is a unique vascular structure. In this study, we investigated the clinical-pathological features of HCC patients with the VETC pattern. We retrospectively reviewed patients with HCC who underwent curative hepatectomy at Chang Gung Memorial Hospital between 2007 and 2013. The form of the VETC pattern was established using an anti-CD31 stain. The results were classified into positive (VETC+) and negative (VETC-) patterns. We investigated and compared demographic data between these two groups. Overall, 174 patients were classified into either the VETC+ or VETC- groups. The median followed-up period was 80.5 months. There were significant differences in the number of hepatitis B carriers, the occurrence of vascular invasion, tumor size, TNM staging, microvessel density, and recurrence (all p &lt; 0.05). Regarding the prediction of disease-free survival, after COX regression multivariate analysis, VETC+ remained independently associated with recurrent episodes (p = 0.003). The intra-tumoral microvessel density, demonstrated by CD-31, was the only clinical-pathological feature independently associated with VETC+. Our study demonstrated that the VETC pattern is an independent factor of poor prognosis for DFS. Higher intra-tumoral microvessel density was significantly associated with the VETC pattern. Further studies are needed to validate our findings.

Expression of GLUT4 and FAP in urothelial bladder carcinoma: correlation with angiogenesis and clinicopathological characteristics

BackgroundUrothelial carcinoma (UC) is the most common type of bladder cancer. Glucose transporter 4 (GLUT4) is one of glucose transporter proteins’ family which facilitates glucose transport inside the cells. It was found to be overexpressed in several malignant tumors. Cancer-associated fibroblasts (CAFs) are heterogeneous stromal cells located adjacent to cancer cells and are considered one of the most important tumor stromal cells. They have been associated with enhancing tumor growth and invasion. GLUT4 expression in malignant epithelial cells and fibroblast activation protein (FAP) expression in CAFs of UC in relation to angiogenesis and clinicopathological characteristics are studied in this work.Materials and methodsThe study was carried out on 72 paraffin blocks of UC (27 radical cystectomies and 45 transurethral resections). Immunohistochemical staining was performed with GLUT4, FAP, and CD34 antibodies. Expression of GLUT4 and FAP was classified according to the staining intensities and percentages into low and high groups. CD34-stained microvessels’ mean count in five microscopic fields (×200) was taken as the microvessel density (MVD).ResultsGLUT4 overexpression was detected in 32 UC. It was significantly associated with high-grade tumors, advanced primary tumor (pT) stage, lymphovascular invasion (LVI), and regional lymph node invasion. High FAP expression was appreciated in 27 UC and was significantly linked to LVI and advanced TNM staging. Intratumor MVD significantly increased in UC with muscle invasion, LVI, and regional lymph node and/or distant metastasis. A significant positive correlation between GLUT4, FAP expression, and MVD was found.ConclusionGLUT4 and FAP expression was significantly associated with increased intratumor MVD and adverse clinicopathological factors.

Complex genetic and histopathological study of 15 patient-derived xenografts of aggressive lymphomas

AbstractNon-Hodgkin lymphomas (NHL) represent the most common hematologic malignancies. Patient-derived xenografts (PDXs) are used for various aspects of translational research including preclinical in vivo validation of experimental treatment approaches. While it was repeatedly demonstrated that PDXs keep majority of somatic mutations with the primary lymphoma samples, from which they were derived, the composition of PDX tumor microenvironment (TME) has not been extensively studied. We carried out a comparative genetic and histopathological study of 15 PDX models derived from patients with various types of NHL including diffuse large B-cell lymphoma (DLBCL; n = 7), Burkitt lymphoma (BL; n = 1), mantle cell lymphoma (MCL; n = 2), and peripheral T-cell lymphomas (PTCL; n = 5). Whole exome sequencing (WES) of the PDXs and primary lymphoma cells was implemented in 13 out of 15 cases with available DNA samples. Standard immunohistochemistry (IHC) was used to analyze the composition of PDX TME. WES data confirmed that PDXs maintained the genetic heterogeneity with the original primary lymphoma cells. In contrast, IHC analysis revealed the following recurrently observed alterations in the composition of PDX tumors: more blastoid lymphoma cell morphology, increased proliferation rate, lack of non-malignant cellular components including T cells and (human or murine) macrophages, and significantly lower intratumoral microvessel density and microvessel area composed of murine vessels. In addition, PDX tumors derived from T-NHL displayed additional differences compared to the primary lymphoma samples including markedly lower desmoplasia (i.e., the extent of both reticular and collagen fibrosis), loss of expression of cytotoxic granules (i.e., perforin, TIA, granzyme B), or loss of expression of T-cell specific antigens (i.e., CD3, CD4, CD8). Our data suggest that despite keeping the same genetic profiles, PDX models of aggressive NHL do not recapitulate the microenvironmental heterogeneity of the original lymphomas. These findings have implications on the relevance of PDX models in the context of preclinical research.

Different predictive values of microvessel density for biochemical recurrence among different PCa populations: A systematic review and meta-analysis.

Several studies have explored the relationship between intratumoral microvessel density (MVD) and the risk of postoperative biochemical recurrence (BCR) in prostate cancer (PCa), although the results are contradictory. Therefore, we conducted a meta-analysis to investigate the effect of MVD on BCR in PCa. We searched PubMed, MEDLINE, Science Direct/Elsevier, the Cochrane Library, CNKI, and EMBase databases from inception through January 2022, with no year or language restrictions, and used NOS guidelines to evaluate the quality of the 19 eligible studies. The derived hazard ratio (HR) and 95% confidence interval (95%CI) were used to assess each endpoint. Data synthesis was performed with RevMan to assess the prognostic value of MVD in PCa and its heterogeneity, while the publication bias was examined using STATA 16.0. Our meta-analysis included 19 articles (4 for T1-2, 6 for T1-3, and 9 for T1-4) on postoperative biochemical recurrence of PCa, among which, 3933 patients were pooled. The predictive ability of intratumoral MVD for different stages of PCa on BCR was T1-2 (HR, 2.46; 95% CI, 1.08-5.58; p= 0.03; I2 = 83%), T1-3 (HR, 2.38, 95% CI, 1.41-4.01; p= 0.001; I2 = 82%), T1-4 (HR, 1.61; 95% CI, 1.19-2.19; p= 0.002; I2 = 61%).The subgroup analyses based on European and immunohistochemical antibody none-factor VII were consistent with primary one. Sensitivity analysis excluding those studies judged to be at high risk of bias in T1-2 showed a HR of 2.99[1.70,5.27] (I2 = 38%, p= 0.0001), demonstrating the robustness of risk estimates of MVD for the assessment of biochemical recurrence. Microvessel density is a predictor of BCR among patients with PCa, and earlier T stage PCa with a stronger MVD is associated with BCR. Further studies are needed to investigate neoangiogenesis in different T stages of PCa and whether MVD will be of benefit to the EAU-recommended tool for biochemical recurrence risk assessment.

The Value of Intratumoral and Extratumoral Microvessel Density for the Tumor-dominated Area in the Endometrial Carcinoma.

Microvessel density (MVD) measuring angiogenesis can influence clinicopathologic variables in endometrial carcinoma (EC). MVD is usually assessed in the densest vascular area the tumor, but the distinction between intratumoral and extratumoral MVD is not reported, and tumor-dominated area is not examined for MVD in the EC in the literature. A total of 151 cases with EC, which had hysterectomy from 2005 to 2020, were included. All histopathologic parameters were re-evaluated blindly. MVD was counted in the intratumoral (densest tumoral and vascular area) and extratumoral (periphery of the tumor, tumor invasion tip, densest tumoral, and vascular area) areas using immunohistochemical CD31 expression. Large tumor size, deep myometrial invasion, high grade, nonendometrioid tumor type, cervix invasion, lymph node metastasis, human epidermal growth factor receptor 2 positivity, stage III to IV, substantial lymphovascular invasion, and overall survival had significant relations with intratumoral and extratumoral MVD ( P <0.05). Age and lymphoplasmacytic inflammation showed marginal significance for extratumoral MVD. Extratumoral and intratumoral MVD had high (near-perfect) agreement (κ=0.870, P =0.001). Cervix invasion, stage III to IV, high grade, intratumoral, and extratumoral MVD were correlated with lower overall survival in the multivariate analyses. High MVD had an impact on the behavior and prognosis of EC and may be a potential indicator for antiangiogenic treatments for aggressive tumors. Although intratumoral and extratumoral MVD in the tumor-dominated area have mostly similar effects in our study, this situation can be cleared more by further investigations.

Abstract 5136: Development of a novel gene expression signature that correlates with intratumor microvessel density and antitumor activity of lenvatinib

Abstract Lenvatinib is a multiple receptor tyrosine kinase inhibitor and an angiogenesis inhibitor targeting mainly vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF) receptors. In our previous study, we reported that human xenograft tumors sensitive to lenvatinib showed higher microvessel density (“MVD (IHC)”) than relatively resistant tumors in immunohistochemistry (IHC). Although MVD (IHC) is a potential indicator to predict antitumor activity of lenvatinib, available IHC data are limited. Here, we investigated a novel gene expression signature (MVD gene score) calculated from RNA-sequencing (RNA-seq) data that reflects MVD (IHC) and antitumor effect of lenvatinib. Tumor tissues were collected from 12 mouse syngeneic tumor models followed by RNA-seq. We also stained CD31 in the same tumors and calculated MVD (IHC) score by dividing the number of CD31 positive blood vessels by tumor area. Genes that met the following criteria were selected for MVD gene score calculation: (1) genes that are specifically expressed on endothelial cell cluster in in-house single cell RNA-seq data using Hepa 1-6 mouse hepatocellular carcinoma tumor tissues; (2) well-known angiogenesis markers from literature or genes whose expression show significant correlation with MVD (IHC) in mouse syngeneic tumor models. MVD gene score was calculated as the average of logarithm transformed gene expression values. In order to examine applicability of MVD gene score, we compared MVD (IHC) and MVD gene score to lenvatinib antitumor activities on mouse syngeneic tumors. We measured MVD (IHC) and computed MVD gene score from RNA-seq data using matched tumor tissues of 12 mouse syngeneic models. MVD gene score was correlated with MVD (IHC). As reported, lenvatinib sensitive tumor models showed higher MVD (IHC) score. They also showed higher MVD gene score calculated from RNA-seq data. Our newly developed MVD gene score has a potential to predict the antitumor activity of angiogenic inhibitors such as lenvatinib using RNA-seq data in tumor samples. Further validation study is ongoing using various human tumor samples. Citation Format: Megumi Kuronishi, Yoichi Ozawa, Takayuki Kimura, Yu Kato. Development of a novel gene expression signature that correlates with intratumor microvessel density and antitumor activity of lenvatinib [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 5136.

Expression of endoglin, CD105, in conjunctival melanocytic nevi: Is it suspicious like in thyroidology? Oculi plus vident quam oculus?

The aim of this study was to evaluate the expression of endoglin and its correlation with histopathological and clinical findings in conjunctival nevi. The study included archival formalin-fixed, paraffin-embedded tissue sections of 44 patients with conjunctival nevi. Immunohistochemical staining for CD105 had been performed with monoclonal mouse antihuman CD105 antibodies. The intratumoral microvessel density for quantification of tumoral vascularization had been determined by this marker. The expression of CD105 was positive in 30 (68.2%) cases. There was a statistically significant difference in the level of CD105 expression regarding the histological type of nevus (p=0.03) and intralesional cysts status (p=0.02). Spearman's rho (ρ -0.316) revealed a significant negative correlation between the expression of endoglin and the histological type of nevus (p=0.03) and between the expression of endoglin and the presence of intralesional cysts (ρ -0.380, p=0.01). This study suggests that endoglin could be a useful diagnostic and prognostic marker in differentiating between benign and malignant melanocytic ocular lesions.

Histopathological Investigation of Dura-like Membrane in Vestibular Schwannomas.

The dura-like membrane (DLM) is an outermost membranous structure arising from the dura mater adjacent to the internal auditory meatus (IAM) that envelops some vestibular schwannomas (VSs). Its recognition is important for the preservation of the facial and cochlear nerves during tumor resection. This study analyzes the histopathological characteristics of the DLM. The expression of CD34 and αSMA was histopathologically analyzed in tumor and DLM tissue of 10 primary VSs with and without a DLM. Tumor volume, resection volume percentage, microvessel density (MVD), and vessel diameter were analyzed. Volumetric analysis revealed that the presence of a DLM was significantly associated with lower tumor resection volume (p < 0.05). Intratumoral vessel diameter was significantly larger in the DLM group than the non-DLM group (p < 0.01). Larger VSs showed a higher intratumoral MVD in the DLM group (p < 0.05). Multilayered αSMA-positive vessels were identified in the DLM, tumor, and border; there tended to be more of these vessels within the tumor in the DLM group compared to the non-DLM group (p = 0.08). These arteriogenic characteristics suggest that the DLM is formed as the tumor induces feeding vessels from the dura mater around the IAM.

Leukocytes infiltration correlates intratumoral microvessel density and influence overall and late-phase disease-free survival in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a severe type of primary liver cancer with high postoperative recurrence. The prognosis predictability of tumor-infiltrating leukocytes (TILs) for patients who underwent HCC resection has been widely reported. However, limited information is available about TIL trafficking, which is also crucial for HCC patients.We included tumor tissue samples and clinical data from 89 HCC patients in this study and performed immunohistochemistry for CD3, CD8, FoxP3, and CD31. TILs were measured using an algorithm for quantification of tumor immune stroma (QTiS). Intratumoral microvessels were counted using Weidner's method. We first examined correlations among them and analyzed their relationships with clinical and survival data.Intratumoral microvessel density (iMVD) was significantly correlated with infiltration of CD3+ (r = 0.338, P = .001) and CD8+ (r = 0.320, P = .002) cells, but not FoxP3+ (r = 0.153, P = .152) cells. After multivariate analysis, higher infiltration of CD3+ (P = .038) independently showed significant predictability on better overall survival after resection of HCC. Although no influence of CD3+ (P = .386) and CD8+ (P = .648) cells were found on general disease-free survival, infiltration of CD3+ (P = .012), tumor size (P = .032) and albumin (P = .007) cells independently predicted late-phase disease-free survival. No significant relationships regarding iMVD, and infiltration of FoxP3+ cells with overall and disease-free survival were found.Our data suggest that increased iMVD could enrich tumor-infiltrating CD3+ cells. Infiltrated CD3+ cells could help to better predict both the overall and late-phase disease-free survival after resection of HCC.

Canine mammary cancer in overweight or obese female dogs is associated with intratumoral microvessel density and macrophage counts.

Obesity is a major health condition owing to its effects on chronic diseases and cancers in humans, but little information is available regarding the role of obesity in canine mammary cancer (CMC). In the present study, we performed immunohistochemistry to investigate the effect of obesity on CMC by analyzing the number of tumor-associated macrophages, intratumoral microvessel density (iMVD), and the expression of prognostic factors including epidermal growth factor receptor (EGFR), cyclooxygenase 2 (COX-2), and Ki67 in CMC specimens. These data were compared in CMC specimens from lean or ideal body weight (Group 1) versus overweight or obese (Group 2) female dogs (n = 60 for each group). Associations between obesity status and histologic characteristics, such as histologic subtype, grading, and lymphatic invasion, were also investigated. Compared with lean or ideal body weight dogs, TAM (tumor-associated macrophage) counts (P < .005) and iMVD (P < .001) were significantly higher in overweight or obese dogs. CMC specimens of dogs in the overweight or obese group also showed higher histologic grade (P < .001). In addition, although no association was found between obesity status and either COX-2 or EGFR expression, Ki67 expression was greater in CMC specimens of overweight or obese dogs (P < .005). The results of this study suggest that obesity may influence CMC development and progression, being associated with higher histologic grade, greater infiltration of TAMs, and increased tumor angiogenesis.