Background:Renal cell carcinoma (RCC) is a highly vascularised tumour, where anti-angiogenic treatment with multi-tyrosine-kinase-inhibitor, is used for first-line treatment of metastatic disease. Variations in vascular characteristics are likely to contribute to variations in intrinsic aggressiveness of the disease. Emerging studies are identifying perivascular status, including perivascular PDGFR-β, as a determinant of prognosis in other tumour types.Methods:This work explored the impact on prognosis of vascular characteristics in RCC through analyses of a population-based collection of tumours from surgery-alone-treated patients. The quantitative data from a panel of vascular metrics were obtained through computerised image analysis of sections double-stained for expression of the endothelial cell marker CD34 together with perivascular markers α-SMA or PDGFR-β.Results:Perivascular expression of PDGFR-β and α-SMA were positively correlated to each other, and negatively correlated to vessel density. High expression of PDGFR-β and α-SMA as well as low vessel density was significantly associated with short survival in uni- and multivariate analyses. Subgroup analyses demonstrated that the prognostic impact of the perivascular markers was particularly prominent in the T4-subgroup. A novel metric, related to PDGFR-β perivascular heterogeneity, was also associated with prognosis in uni-and multi-variate analyses. This novel metric also acted as a prognosis marker in ovarian cancer.Conclusions:The study demonstrates previously unrecognised associations between RCC survival and the absolute levels, and variability, of perivascular PDGFR-β. This marker should be further explored in other RCC cohorts. Findings also suggest mechanistic analyses and studies on the relationship between perivascular status and efficacy of multi-tyrosine-kinase-inhibitors.

Background The role of B-lymphocytes in solid tumours is unclear. Tumour biology studies have implied both anti- and pro-tumoural effects and prognostic studies have mainly linked B-cells to increased survival. This study aimed to analyse the clinical relevance of B-lymphocytes in renal cell cancer (RCC), where information on the prognostic impact is lacking. Methods Following immunohistochemistry (IHC) stainings with a CD20 antibody, density of CD20+ B-cells was quantified in an RCC discovery- and validation cohort. Associations of B-cell infiltration, determined by CD20 expression or a B-cell gene-signature, and survival was also analysed in 14 publicly available gene expression datasets of cancer, including the kidney clear cell carcinoma (KIRC) dataset. Results IHC analyses of the discovery cohort identified a previously unrecognised subgroup of RCC patients with high infiltration of CD20+ B-cells. The B-cell-high subgroup displayed significantly shorter survival according to uni- and multi-variable analyses. The association between poor prognosis and high density of CD20+ B-cells was confirmed in the validation cohort. Analyses of the KIRC gene expression dataset using the B-cell signature confirmed findings from IHC analyses. Analyses of other gene expression datasets, representing 13 different tumour types, indicated that the poor survival-association of B-cells occurred selectively in RCC. Conclusion This exploratory study identifies a previously unrecognised poor-prognosis subset of RCC with high density of CD20-defined B-cells.

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Following traumatic brain injuries (TBI), insulin-like growth factor (IGF) is cortically widely upregulated. This upregulation has a potential role in the recovery of neuronal tissue, plasticity, and neurotrophic activity, though the molecular mechanisms involved in IGF regulation and the exact role of IGF after TBI remain unclear. Vitronectin (VN), an extracellular matrix (ECM) molecule, has recently been shown to be of importance for IGF-mediated cellular growth and migration. Since VN is downregulated after TBI, we hypothesized that insufficient VN levels after TBI impairs the potential beneficial activity of IGF. To test if vitronectin and IGF-1/IGFBP-2 could contribute to neurite growth, we cultured hippocampal neurons on ± vitronectin-coated coverslips and them treated with ± IGF-1/IGF binding protein 2 (IGFBP-2). Under same conditions, cell cultures were also subjected to in vitro trauma to investigate differences in the posttraumatic regenerative capacity with ± vitronectin-coated coverslips and with ± IGF-1/IGFBP-2 treatment. In both the control and trauma situations, hippocampal neurons showed a stronger growth pattern on vitronectin than on the control substrate. Surprisingly, the addition of IGF-1/IGFBP-2 showed a decrease in neurite growth. Since neurite growth was measured as the number of neurites per area, we hypothesized that IGF-1/IGFBP-2 contributes to the polarization of neurons and thus induced a less dense neurite network after IGF-1/IGFBP-2 treatment. This hypothesis could not be confirmed and we therefore conclude that vitronectin has a positive effect on neurite growth in vitro both under normal conditions and after trauma, but that addition of IGF-1/IGFBP-2 does not have a positive additive effect.

A novel set of integrated procedures for quantification of fibroblast‐rich stroma and vascular characteristics has recently been presented allowing discovery of novel perivascular and stromal biomarkers in colorectal, renal cell, and ovarian cancer. In the present study, data obtained through these procedures from clinically well‐annotated collections of these three tumour types have been used to address two novel questions. First, data have been used to investigate if the three tumour types demonstrate significant differences regarding features such as vessel diameter, vessel density, and perivascular marker expression. Second, analyses of the cohorts have been used to explore the prognostic significance of a novel vascular metric, ‘vessel distance inter‐quartile range (IQR)’ that describes intra‐case heterogeneity regarding vessel distribution. The comparisons between the three tumour types demonstrated a set of significant differences. Vessel density of renal cell cancer was statistically significantly higher than in colorectal and ovarian cancer. Vessel diameter was statistically significantly higher in ovarian cancer. Concerning perivascular status, colorectal cancer displayed significantly higher levels of perivascular PDGFR‐β expression than the other two tumour types. Intra‐case heterogeneity of perivascular PDGFR‐β expression was also higher in colorectal cancer. Notably, these fibroblast‐dominated stroma phenotypes matched previously described experimental tumour stroma characteristics, which have been linked to differential sensitivity to anti‐VEGF drugs. High ‘vessel distance IQR’ was significantly associated with poor survival in both renal cell cancer and colorectal cancer. In renal cell cancer, this characteristic also acted as an independent prognostic marker according to multivariate analyses including standard clinico‐pathological characteristics. Explorative subset analyses indicated particularly strong prognostic significance of ‘vessel distance IQR’ in T stage 4 of this cancer type. Together, these analyses identified tumour‐type‐specific vascular‐stroma phenotypes of possible functional significance, and suggest ‘vessel distance IQR’ as a novel prognostic biomarker.

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Background : Studies in experimental models have implied perivascular cells (PC) as regulators of multiple aspects of tumor biology, including tumor growth, metastasis, immune cell activity and response to treatment. However, the extent of inter- and intra-case variability of perivascular cells in clinical samples, and potential of perivascular cells as biomarkers, remain poorly characterized. Methods: We have developed an integrated analytical pipeline to investigate vascular and PC status in clinical samples. The analytical procedure uses double-staining with one endothelial cell marker (CD34) and one of four PC markers; platelet-derived growth factor receptor-alpha and -beta (PDGFR-α, PDGFR-β;), smooth muscle α actin (ASMA) and desmin. Digital image-analyses are subsequently used to quantitate PC-related metrics including intensity of marker expression in perivascular areas, and fraction of marker-covered vessels. The approach has been applied to multiple cohorts of clinically well-annotated tumor collections from three cancer types; colorectal cancer (CRC), ovarian cancer (OC) and renal cell cancer (RCC). Results: Analyses of PC status uncovered previously un- recognized independent and heterogeneous expression of the analyzed perivascular markers on different levels (perivascular cells, vessels and tumors). Notably, expression of perivascular markers was largely independent of vessel size and density in all three cancer types. Comparative analyses across tumor types identified differences including e.g. higher abundance of PDGFR-β; positive perivascular cells in CRC. A comparison of perivascular status in matched primary tumors and distant metastases uncovered a large degree of dis-concordance of most vascular characteristics except perivascular PDGFR-β; (CRC and OC cohorts). Importantly, this finding implies that vascular status in metastases cannot, in general, be predicted by analyses of primary tumors. Efforts to uncover the basis for inter-case variations in PC status of CRC revealed strong associations between tumor PDGFR-β; PC status and both PC status in adjacent normal tissue and BRAF status, but not RAS mutations. These findings suggest both host features and cancer mutations as determinants of PC characteristics. Concerning relationships between PC features and survival, we observed significant associations between high perivascular PDGFR-β; status and shorter survival in OC and RCC. These associations remain significant in multivariate analyses including standard clinicopathological characteristics. Impact on response to chemotherapy (CT) treatment was analyzed in both early stage and metastatic CRC. Ongoing analyses indicate that low perivascular PDGFR-βdefines a sub-group with shorter survival in the CT-treated group. Explorative analyses of a small set of bevacizumab-treated metastatic CRC also demonstrated significantly shorter progression free survival in the subset with low perivascular PDGFR-β expression. Conclusion: Digital-image-analyses-supported quantifications of PC features in three different tumor types have revealed heterogeneous expression of PC markers in a manner independent of vessel density and vessel size. Host characteristics and oncogenic status have been identified as candidate determinants of perivascular PDGFR-β; status in CRC. Notably, perivascular PDGFR-β; was identified as a novel independent predictor of survival in OC, RCC and CRC. Treatment-stratified analyses in CRC also implied perivascular PDGFR-β; as a determinant of benefit of chemotherapy and VEGF-directed anti-angiogenic drugs. Citation Format: Sara corvigno, artur mezheyeuski, Magnus Frodin, Maja Bradic Lindh, Tormod Kyrre Guren, Per Pfeiffer, H. Elin Kure, Tone Ikdahl, Eva Skovlund, Kjell Magne Tveit, Kristian Pietras, Anna Portyanko, Ina Hrynchyk, David Edler, Anca Dragomir, friedrik Ponten, Jan Mulder, Camilla Qvortrup, Maria Hallström, Katarina Öhrling, G. Bea A. Wisman, Elisabeth Åvall-Lundqvist, Martin Johansson, Ulrika Harmenberg, Hans Nijman, Per Sandström, Hanna Dahlstrand, Halfdan Sorbye, Bengt Gimelius, Ate Van Der Zee, Lars Egevad, Arne Östman. Marker-defined perivascular cells predict prognosis and response to treatment. [abstract]. In: Proceedings of the AACR Special Conference: Function of Tumor Microenvironment in Cancer Progression; 2016 Jan 7–10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2016;76(15 Suppl):Abstract nr C38.