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Assessment of VAV2 expression refines prognostic prediction in adrenocortical carcinoma
Silviu Sbiera, Iuliu Sbiera, Carmen Ruggiero, Mabrouka Doghman-Bouguerra, Esther Korpershoek, Ronald R. de Krijger, Hester Ettaieb, Harm Haak, Marco Volante, Mauro Papotti, Giuseppe Reimondo, Massimo Terzolo, Michaela Luconi, Gabriella Nesi, Massimo Mannelli, Rossella Libé, Bruno Ragazzon, Guillaume Assié, Jérôme Bertherat, Barbara Altieri, Guido Fadda, Natalie Rogowski-Lehmann, Martin Reincke, Felix Beuschlein, Martin Fassnacht, Enzo Lalli
The Journal of Clinical Endocrinology & Metabolism Endocrine Society
Submitted: April 26, 2017
Accepted: June 28, 2017 First Online: July 03, 2017
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THE JOURNAL OF CLINICAL ENDOCRINOLOGY & METABOLISM
VAV2 in adrenocortical carcinoma
Assessment of VAV2 expression refines prognostic prediction in adrenocortical carcinoma
Silviu Sbiera1 , Iuliu Sbiera1, Carmen Ruggiero2-5, Mabrouka Doghman-Bouguerra2-5, Esther Korpershoek6, Ronald R. de Krijger6,7, Hester Ettaieb8, Harm Haak8-10, Marco Volante11, Mauro Papotti11, Giuseppe Reimondo12, Massimo Terzolo12, Michaela Luconi13, Gabriella Nesi13, Massimo Mannelli13, Rossella Libé14-16, Bruno Ragazzon14-16, Guillaume Assié14-16, Jérôme Bertherat14-16, Barbara Altieri1,17, Guido Fadda18, Natalie Rogowski-Lehmann19, Martin Reincke19, Felix Beuschlein19,20, Martin Fassnacht21, Enzo Lalli2-5”
1Department of Internal Medicine I - Division of Endocrinology and Diabetes, University Hospital, University of Würzburg, 97080 Würzburg, Germany; 2 Université Côte d’Azur, Sophia Antipolis, 06560 Valbonne, France; 3 CNRS UMR7275, Sophia Antipolis, 06560 Valbonne, France; 4NEOGENEX CNRS International Associated Laboratory, Sophia Antipolis, 06560 Valbonne, France; 3Institut de Pharmacologie Moléculaire et Cellulaire, Sophia Antipolis, 06560 Valbonne, France; ‘Department of Pathology, Erasmus MC Cancer Institute, University Medical Center, 3000 CA Rotterdam, The Netherlands; 7Department of Pathology, Reinier de Graaf Hospital, 2625 AD Delft, The Netherlands; 8Department of Internal Medicine, Máxima Medical Centre, Eindhoven/Veldhoven, The Netherlands; 9 Department of Internal Medicine, Division of General Internal Medicine, Maastricht University Medical Centre+, Maastricht, The Netherlands; 10Maastricht University, CAPHRI School for Public Health and Primary Care, Ageing and Long-Term Care, Maastricht, The Netherlands; 11 Department of Oncology and 12Department of Clinical and Biological Sciences, University of Turin at San Luigi Hospital, 10043 Orbassano, Italy; 13Department of Experimental and Clinical Biomedical Sciences “Mario Serio”, University of Florence, 50139 Florence, Italy; 4Inserm U1016, Institut Cochin, 75014 Paris, France; 15 CNRS UMR8104, 75014 Paris, France; 16 Université Paris Descartes, Sorbonne Paris Cité, 75014 Paris, France; Division of Endocrinology and Metabolic Diseases and 8Division of Anatomic Pathology and Histology, Catholic University of the Sacred Heart, 00168 Rome, Italy;
9 Medizinische Klinik and Poliklinik IV, Ludwig-Maximilians-Universität 80336 Munchen, Germany; 20 Klinik für Endokrinologie, Diabetologie und Klinische Ernährung, Universitätsspital Zürich, 8091 Zürich, Switzerland; 21 Comprehensive Cancer Center Mainfranken, University of Würzburg, 97080 Würzburg, Germany.
Received 26 April 2017. Accepted 28 June 2017.
Background: Adrenocortical carcinoma (ACC) is a rare endocrine malignancy with overall poor prognosis. The Ki67 labeling index (LI) has a major prognostic role in localized ACC after complete resection but its estimates may suffer from considerable intra- and interobserver variability. VAV2 overexpression induced by increased SF-1 dosage is an essential factor driving ACC tumor cell invasion.
Objective: To assess the prognostic role of VAV2 expression in ACC by investigation of a large cohort of patients.
Design, Setting and Participants: 171 ACC cases (157 primary tumors, 6 local recurrences, 8 metastases) from seven ENS@T centers were studied.
Outcome Measurements: H-scores were generated quantifying VAV2 expression. VAV2 expression was divided into two categories, low (H-score <2) and high (H-score ≥2). Ki67 LI retrieved from patients’ pathological records was also categorized into low (<20%) and high (≥20%). Clinical and immunohistochemical markers were correlated with progression-free (PFS) and overall survival (OS).
Results: VAV2 expression and Ki67 LI were significantly correlated with each other and with PFS and OS. Heterogeneity of VAV2 expression inside the same tumor was very low. Combined
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assessment of VAV2 expression and Ki67 LI allowed to improve patient stratification to low- risk and high-risk groups.
Conclusion: Combined assessment of Ki67 LI and VAV2 expression improves prognostic prediction in ACC.
We studied VAV2 expression in a large multicentric cohort of adrenocortical carcinoma cases and validated its role as a prognostic marker.
Introduction
Adrenocortical carcinoma (ACC) is a rare endocrine malignancy with overall poor prognosis, limited treatment options when progressed into metastatic stage and unsatisfactory response to polychemotherapeutic cytotoxic regimens (1, 2). Hence the most efficient method to eradicate the disease consists in complete surgical resection of the primary tumor. However, risk of recurrence is high even in this condition. Molecular studies have identified two subclasses of ACCs with aggressive (C1A) or indolent (C1B) clinical behavior, respectively (3-6). However, since molecular markers identified by those studies have not yet found entrance into clinical practice, it would be of particular importance to stratify patients with ACC into low-or high-risk groups to adequately monitor disease recurrence and assign them to appropriate therapeutic interventions. The histological Weiss score, which is commonly used as an established morphometric criterion for the differential diagnosis in adrenocortical tumors, has limited value as a prognostic indicator, especially in cases with borderline features (7, 8). Conversely, it was shown that a number of immunohistochemical markers have a prognostic value in ACC (9-18). Among those, the most widely used in clinical pathology reports is the Ki67 labeling index (LI), which is directly related to the proliferative activity of a given tissue (14-18). A study recently completed by the European Network for the Study of Adrenal Tumors (ENS@T) could indeed demonstrate that Ki67 LI has a major prognostic role in localized ACC after complete resection (18). However, Ki67 LI estimates suffer from considerable intra- and interobserver variability, as highlighted in a recent study (19). New prognostic markers are therefore needed to further refine prognostic classification of patients with ACC as part of a multiparametric analysis.
The transcription factor Steroidogenic Factor-1 has a pivotal role in regulating adrenocortical cell proliferation and differentiation (20). Its overexpression is associated to adrenocortical tumorigenesis through regulation of a specific set of SF-1 dosage-dependent target genes (21, 22). One of these genes encodes VAV2, a guanine nucleotide exchange factor (GEF) for small GTPases of the Rho family (23). We have recently shown that VAV2 overexpression induced by an increased SF-1 dosage in ACC is an essential factor driving tumor cell invasion (24). Herein, we present the results of a large study involving ACC cases provided by seven European institutions aimed to assess the prognostic value of VAV2 expression in ACC and to compare and integrate it with the Ki67 LI.
Materials and Methods
Immunostaining on formalin-fixed, paraffin embedded ACC samples
We analyzed a total of 171 adrenocortical tumor tissues from patients with ACC provided by seven ENS@T centers (Italy 103, The Netherlands 42, France 20, Germany 6 samples). 145 samples were previously assembled in 7 tissue microarrays (TMA) with 2 or 3 cores per sample, interspersed with normal human liver, kidney and placenta tissues, and 26 samples were available as full slides. Among the ACC samples, 157 samples derived from primary tumors (male/female 59/98, average age±SD 48.7±15.2 years, average tumor size±SD 11.2+5.4 cm; for
patients’ characteristics see Table S1), 6 from local recurrences and 8 from distant metastases (liver and lung). The diagnosis of ACC was made by established criteria based on clinical, biochemical and morphological data (25). All clinical data were collected through the ENS@T database (registry.ensat.org). All patients gave informed consent and the study was approved by ethical committees from all participating institutions. Immunohistochemical detection was performed in all samples using an indirect immunoperoxidase technique after high temperature antigen retrieval in 0.01 M citrate buffer (pH 6.5) in a pressure-cooker for 13 minutes. The primary antibody was a rabbit monoclonal antibody against the VAV2 protein (clone EP1067Y, ab52640 Abcam) diluted 1:250 in 25% AB serum in PBS and incubated 1 h at RT. Signal detection was performed with the Advance HRP detection system (Dako) and DAB chromogen according to the manufacturer’s instructions. Nuclei were counterstained with Mayer’s hematoxylin for 3 minutes. As negative control, universal rabbit negative control (Dako) was used. Immunostaining results were analyzed using a light microscope at high magnification. VAV2 staining intensity was evaluated independently by two investigators blinded to the clinical data (S.S. and I.S.). Cytoplasmic staining intensity was evaluated with a grading score of 0, 1, 2 or 3, corresponding to negative, weak, moderate and strong intensity, respectively. The proportion of positive tumor cells was calculated for each specimen and set up to be scored 0, 0.1, 0.5 or 1, if 0%, 1-9%, 10-49% or >50% of the tumor cells were positive for VAV2, respectively. A semi-quantitative H-score was then calculated by multiplying the staining intensity grade by the proportion score (12, 24). In all cases analyzed, the proportion of VAV2 positive cells was always >50%, so all intensity values were multiplied by a factor equal to 1 to yield the H-score. The cut-off point to separate samples in high or low VAV2 expression was between H-scores <2 and >2. Ki67 LI data assessed by the local pathologists in each expert center were retrieved from the ENS@T database. The Ki67 LI cut-off value used in this study to separate low LI and high LI groups was 20%.
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Statistical analysis
Correlation analyses were performed using a x2 test for categorical variables. The inter-observer agreement for the scoring system was evaluated using Cohen’s kappa-coefficient and confirmed using Pearson’s correlation coefficient. As cutoff for strong agreement 0.81 was chosen for the kappa-coefficient and 0.75 for Pearson’s coefficient (26). The comparison of clinical and histopathological characteristics was performed on GraphPad Prism 6.0 software using non- parametric Mann-Whitney test (for two groups) and Kruskal-Wallis test with Dunn’s correction for multiple testing (for more than two groups), as appropriate. A p value <0.05 was considered to be statistically significant. Survival analysis for ACC patients was calculated as described (24) using the Kaplan-Meier method and differences between groups were assessed with log-rank and Cox proportional hazards statistics, using the SPSS software package (version 23.0.0 for Mac), after adjustment for sex, age and tumor stage. Progression-free survival (PFS) was defined as time elapsed from primary resection of ACC to the first recurrence, loco-regional or systemic. Overall survival (OS) was defined as time elapsed from primary resection of ACC to disease- related death or last follow-up visit. In the group of patients with RO resection, OS data were available for 100 (VAV2) and 105 (Ki67 LI) patients, respectively. 92 of those patients had both VAV2 and Ki67 LI OS data available. Viable cell data after VAV2 knockdown were analyzed by 1-way ANOVA with Dunnett’s correction for multiple comparisons.
Results
VAV2 expression is a strong predictor of PFS and OS in ACC patients
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Examples of different VAV2 expression patterns in ACC are shown in Fig. 1. A H-score was assigned to each sample, which took in consideration both staining intensity and the percentage of cells stained by the anti-VAV2 antibody. The inter-observer agreement was very good with Cohen’s kappa coefficient equal to 0.85 (95%CI: 0.72-0.89) and Pearson coefficient r=0.90 (95%CI:0.86-0.93), p<0.001. In contrast to Ki67 staining, which is usually heterogeneous throughout a tumor, VAV2 expression was fairly equally distributed within a given tumor, with all samples presenting a percentage of stained cells >50%. H-score heterogeneity among different TMA tissue cores belonging to the same tumors was limited, with a residual standard deviation o=0.14 and an intra-class correlation coefficient a=0.95 (95%CI: 0.92-0.97) (Fig. S1). The same homogenous distribution was also observed when whole tumor slides were analyzed (Fig. 1). VAV2 expression in the tumor was strongly correlated to both PFS (Fig. 2A) and OS (Fig. 2B), confirming the results of our previous study performed on an independent smaller cohort of ACC patients (24). Patients with strong VAV2 expression had a 2.8-fold higher risk to experience a recurrence and 1.6-fold increased risk to die. No statistically significant difference existed for VAV2 expression in primary tumors and metastatic sites from the same patients (p=0.67). The Ki67 LI was also a strong predictor of PFS (Fig. 2C) and OS (Fig. 2D), as reported in previous studies (14-18). Both VAV2 expression and Ki67 LI were strongly correlated with OS even in patients with RO resection (Fig. S2). VAV2 expression and Ki67 LI had a similar strong prognostic value for PFS and OS both in univariate and in multivariate analysis, taking into account patients’ age, sex and tumor stage (Table 1).
Combined assessment of VAV2 expression and Ki67 LI improves prognostic power
In general, a significant correlation existed between Ki67 LI and VAV2 expression in our ACC cohort (Fig. S3). A strong correlation also existed when Ki67 LI and VAV2 expression were considered as categorical (low vs. high) variables (x2 = 6.18, p=0.01). However, in several cases these two parameters were dissociated with one value being elevated and the other low in the same tumor. Remarkably, in those patients PFS and OS were intermediate between the high- risk (high VAV2 expression-high Ki67 LI) and the low-risk groups (low VAV2 expression-low Ki67 LI) (Fig. 3A, B). Merging the groups with high VAV2-low Ki67 LI and low VAV2-high Ki67 LI and comparing them to the high VAV2-high Ki67 LI and low VAV2-low Ki67 LI groups identified three classes of patients with very different RFS (159.7±23.2, 90.3±15.7 and 20.8±5.8 months, respectively) and OS (203.7±29.6, 130.3±29.6 and 41.6±5.1 months, respectively) (Fig. 3C, D). This type of stratification maintained a strong prognostic value even in RO patients (Fig. S4). Remarkably, when considering the high-risk group apart from all other patients with ACC, a very strong correlation existed with OS in the whole cohort (Fig. 4A) and with both PFS and OS in RO patients (Fig. 4B, C). Furthermore, isolated high VAV2 expression or high Ki67 LI showed a prediction value for worse PFS and OS that was slightly lower compared with the combination of both high VAV2 expression + high Ki67 LI [PFS: 22 months, HR=0.67 (VAV2) and 28 months, HR=0.66 (Ki67 LI) vs. 9 months for the combination; OS: 66 months, HR=0.73 (VAV2) and 40 months, HR=0.82 (Ki67 LI) vs. 33 months for the combination].
Discussion
The prognosis of ACC patients is variable and poorly predictable. A recent large multicentric ENS@T study has shown that the KI67 LI is the most powerful parameter predicting disease recurrence and survival in ACC patients after complete tumor resection (18). The Ki67 LI has been integrated with the combined evaluation of morphological parameters (number of
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mitoses/presence of necrosis) in the newly introduced Helsinki score, which reportedly is able to more accurately predict recurrence in ACC (8, 27). However, even if Ki67 LI assessment is routinely performed in diagnostic pathology laboratories for a large number of neoplastic disorders, its standardization and reproducibility have been questioned for many tumor types, including ACC (19). It is therefore important to identify other molecular markers that can complement the Ki67 LI to obtain a more accurate stratification of the risk of recurrence in patients with ACC. In this perspective, molecular prognostic indicators derived from genomic studies are very promising (3, 28, 29), but for routine implementation they suffer from the important drawback that, at least at the present state of technology, frozen tumoral material is required. On the other hand, prognostic value of circulating markers of malignancy awaits validation in large cohorts of ACC patients (30-33).
We have recently shown that VAV2 overexpression is an essential driver of cell invasion in conditions of increased SF-1 dosage through its GEF activity for the small GTPases Rac1 and Cdc42 (24). Those data directly link VAV2 with the potential mechanism of malignancy consisting in increased cellular invasiveness. In the present study we extended the previous study to a large European cohort of patients with ACC and show that the tumor VAV2 H-score is significantly correlated to PFS and OS. The combined assessment of VAV2 expression and Ki67 LI improves patient risk stratification, with cases presenting high Ki67 LI but low VAV2 expression having significantly longer PFS and OS compared to patients with concordant high- risk parameters. In our study, VAV2 H-score assessment, which was mainly performed on TMA tissue cores, was associated to an excellent intratumoral reproducibility and is then in principle less prone to intra- and interobserver variability, although further work is needed to specifically address this question on an even larger number of cases. These results show that immunohistochemical assessment of VAV2 expression may usefully complement the measurement of the Ki67 LI for prognostic stratification of patients with ACC.
Acknowledgments
This work was supported by the Else Kroner-Fresenius-Stiftung Grant 2016_A96 and by a fellowship from the “Novartis-Stiftung für theraputische Forschung” to S.S; Italian Association for Cancer Research (AIRC) grants no. IG/14820/2013 to M.P., 14411 to M.T. and IG/17691/2015 to M.L .; ERA-NET “E-Rare” (http://www.erare.eu/) (grant 01GM1407B) to M.F .; French National Research Agency (ANR) through the LOCALDO (ANR-15-CE14-0017- 01) and “Investments for the Future” LABEX SIGNALIFE (ANR-11-LABX-0028-01) grants to E.L. C.R. was a recipient from Ville de Nice and Fondation de France (grant 00057927) post- doctoral fellowships.
*Correspondence to: Silviu Sbiera, Universitätsklinikum Würzburg - Medizinische Klinik und Poliklinik I, Endokrinologie Forschung ZIM, A4 .- 3.949,
Oberdürrbacherstrasse 6, 97080 Würzburg, Germany, Tel. +49 (0)931-201-39702; Fax +49 (0)931-201-639702; Email: Sbiera_S@ukw.de or Enzo Lalli, Institut de Pharmacologie Moléculaire et Cellulaire CNRS UMR7275, 660 route des Lucioles - Sophia Antipolis, 06560 Valbonne, France, Tel: +33 4 93 95 77 55; Fax: +33 4 93 95 77 08; Email: ninino@ipmc.cnrs.fr
Disclosure summary: The authors have nothing to disclose.
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33. Liu-Chittenden Y, Patel D, Gaskins K, Giordano TJ, Assie G, Bertherat J, Kebebew E. Serum RARRES2 Is a prognostic marker in patients with adrenocortical carcinoma. J Clin Endocrinol Metab. 2016;101:3345-3352.
Figure 1. Examples of various intensities of VAV2 staining in ACC specimens. H-score value is indicated for each image, respectively. Scale bar, 400 um (images in left column); 50 um (images in right column).
Figure 2. Correlation of VAV2 expression (H-score) and Ki67 LI with PFS and OS in our ACC series. (A) PFS in low VAV2 expression (H-score <2) group (green line) 127±15.9 months; high VAV2 expression (H-score ≥2) group (red line) 25.7+4.1 months. p<0.001, Kaplan-Meier method. (B) OS in low VAV2 expression (H-score <2) group (green line) 180±22 months; high VAV2 expression (H-score ≥2) group (red line) 87.4±13 months. p=0.001, Kaplan- Meier method. (C) PFS in low Ki67 LI (<20%) group (green line) 137±17.9 months; high Ki67 LI (≥20%) group (red line) 68.5±14.3 months. p<0.001, Kaplan-Meier method. (D) OS in low Ki67 LI (<20%) group (green line) 187.5±22.9 months; high Ki67 LI (≥20%) group (red line) 96.2±17 months. p=0.001, Kaplan-Meier method. The numbers of cases analyzed for each group are reported in parentheses.
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Figure 3. Correlation of combined VAV2 expression (H-score) and Ki67 LI with PFS and OS in our ACC series. (A) PFS in low VAV2 expression (H-score <2)-low Ki67 LI (<20%) group (green line) 159.7±23.2 months; high VAV2 expression (H-score ≥2)-low Ki67 LI (<20%) group (yellow line) 50.7±8.4 months; low VAV2 expression (H-score <2)-high Ki67 LI (≥20%) group (pale green line) 96.6±26.3 months; high VAV2 expression (H-score ≥2)-high Ki67 LI (≥20%) group (red line) 20.8±5.8 months. Compared to low VAV2-low Ki67 LI: high VAV2- low Ki67 LI HR=2.55 (1.09-5.97), p=0.030; low VAV2-high Ki67 LI HR=2.46 (0.97-6.23), p=0.058; high VAV2-high Ki67 LI HR=6.75 (2.97-15.31), p<0.001; Kaplan-Meier method. (B) OS in low VAV2 expression (H-score <2)-low Ki67 LI (<20%) group (green line) 203.7±29.6 months; high VAV2 expression (H-score ≥2)-low Ki67 LI (<20%) group (yellow line) 120.4±20.5 months; low VAV2 expression (H-score <2)-high Ki67 LI (≥20%) group (pale green line) 126±26.7 months; high VAV2 expression (H-score ≥2)-high Ki67 LI (≥20%) group (red line) 41.6±5.1 months. Compared to low VAV2-low Ki67 LI: high VAV2-low Ki67 LI HR=2.66 (1.08-6.52), p=0.032; low VAV2-high Ki67 LI HR=3.51 (1.38-8.91), p=0.008; high VAV2-high Ki67 LI HR=5.38 (2.33-12.40), p<0.001; Kaplan-Meier method. (C) PFS in low VAV2 expression (H-score <2)-low Ki67 LI (<20%) group (green line) 159.7+23.2 months; high VAV2 expression (H-score ≥2)-high Ki67 LI (≥20%) group (red line) 20.8±5.8 months; all other patients with dissociated VAV2 expression-Ki67 LI group (grey line) 90.3±15.7 months. Compared to low VAV2-low Ki67 LI: other HR=2.51 (1.17-5.39), p=0.018; high VAV2-high Ki67 LI HR=6.75 (2.97-15.31), p<0.001; Kaplan-Meier method. (D) OS in low VAV2 expression (H-score <2)-low Ki67 LI (<20%) group (green line) 203.7±29.6 months; high VAV2 expression (H-score ≥2)-high Ki67 LI (≥20%) group (red line) 41.6±5.1 months; all other patients with dissociated VAV2 expression-Ki67 LI group (grey line) 130.3±18.1 months. Compared to low VAV2-low Ki67 LI: other HR=2.99 (1.32-6.73), p=0.008; high VAV2-high Ki67 LI, HR=5.38 (2.33-12.40), p<0.001; Kaplan-Meier method. The numbers of cases analyzed for each group are reported in parentheses.
Figure 4. Prognosis of high-risk (high VAV2 expression-high Ki67 LI) vs. other ACC patients. (A) OS in the whole cohort of ACC patients for the high VAV2 expression (H-score ≥2)-high Ki67 LI (≥20%) group (red line) 41.5±5 months; all other patients (green line) 175.5±19.8 months. p<0.001, Kaplan-Meier method. (B) PFS in RO patients for the high VAV2 expression (H-score ≥2)-high Ki67 LI (≥20%) group (red line) 20.8+5.8 months; all other patients (green line) 127.3±15.7 months. p<0.001, Kaplan-Meier method. (C) OS in RO patients for the high VAV2 expression (H-score ≥2)-high Ki67 LI (≥20%) group (red line) 47.5±6 months; all other patients (green line) 194.8±21.7 months. p=0.005, Kaplan-Meier method. The numbers of cases analyzed for each group are reported in parentheses.
| PFS | Univariate analysis | Multivariate analysis | ||||||
|---|---|---|---|---|---|---|---|---|
| Variables | HR | 95% CI | p | HR | 95% CI | p | ||
| age (n=113; n=99) | 1.00 | 0.98-1.02 | 0.77 | 0.99 | 0.98-1.01 | 0.91 | ||
| sex | ||||||||
| female (n=78; n=69) | ||||||||
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| male (n=35; n=30) | 1.19 | 0.69-2.04 | 0.52 | 1.17 | 0.64-2.13 | 0.59 | ||
|---|---|---|---|---|---|---|---|---|
| Tumor stage | ||||||||
| I (n=12; n=8) | ||||||||
| II (n=66; n=61) | 7.19 | 0.98-52.52 | 0.05 | 5.32 | 0.71-39.39 | 0.10 | ||
| III (n=23; n=19) | 6.29 | 0.81-48.65 | 0.07 | 5.03 | 0.64-39.65 | 0.12 | ||
| IV (n=11; n=11) | 27.51 | 3.40-222.11 | 0.002 | 14.65 | 1.74-122.96 | 0.01 | ||
| VAV2 expression | ||||||||
| VAV2 low (H-score 0- 1) (n=52; n=52) | ||||||||
| VAV2 high(H- score 2-3) (n=48; n=47) | 2.80 | 1.57-4.98 | <0.001 | 2.83 | 1.54-5.21 | 0.001 | ||
| Ki67 LI | ||||||||
| Ki67 low (<20%) (n=63; n=62) | ||||||||
| Ki67 high (≥20%) (n=42; n=42) | 2.77 | 1.58-4.86 | <0.001 | 2.43 | 1.37-4.31 | 0.002 | ||
| OS | Univariate analysis | Multivariate analysis | ||||||
| Variables | HR | 95% CI | p | HR | 95% CI | p | ||
| age (n=156; n=74) | 1.01 | 0.99-1.03 | 0.14 | 1.01 | 0.98-1.03 | 0.35 | ||
| sex | ||||||||
| female (n=98; n=79) | ||||||||
| male (n=58; n=45) | 1.27 | 0.76-2.12 | 0.35 | 1.39 | 0.80-2.41 | 0.24 | ||
| Tumor stage | ||||||||
| I (n=12; n=8) | 3.71 | 0.50-27.60 | 0.2 | |||||
| II (n=72; n=65) | 3.71 | 0.50-27.60 | 0.2 | 3.77 | 0.49-28.84 | 0.20 | ||
| III (n=35; n=29) | 4.52 | 0.59-34.49 | 0.14 | 3.85 | 0.49-29.87 | 0.19 | ||
| IV (n=24; n=22) | 19.07 | 2.52-144.30 | 0.004 | 13.74 | 1.77-106 | 0.01 | ||
| VAV2 expression | ||||||||
| VAV2 low (H-score 0- 1) (n=66; n=60) | ||||||||
| VAV2 high (H-score 2- 3) (n=76; n=64) | 1.64 | 1.01-2.66 | 0.042 | 2.03 | 1.07-3.83 | 0.02 | ||
| Ki67 LI | ||||||||
| Ki67 low | ||||||||
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| (<20%) (n=77; n=72) | ||||||||
|---|---|---|---|---|---|---|---|---|
| Ki67 high (≥20%) (n=68; n=60) | 2.94 | 1.67-5.19 | <0.001 | 2.31 | 1.24-4.30 | 0.008 |
Abbreviations: CI, confidence interval; HR, hazard ratio; PFS, progression free-survival; OS, overall survival.
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ENDOCRINE SOCIETY
ADVANCE ARTICLE: JCEM
AD
H-score=3
H-score=2
H-score=1
H-score=0
MANGE ARTICLE
A
B
Progression-free survival (%)
100
100
80
Overall survival (%)
80
60
low VAV2 (n=52)
60
low VAV2 (n=66)
40
p<0.001
40
p=0.001
20
20
high VAV2 (n=76)
high VAV2 (n=48)
0
0
0
50
100
150
200
250
300
0
50
100
150
200
250
300
C
Time (months)
D
Time (months)
Progression-free survival (%)
100
100
80
Overall survival (%)
80
60
low Ki67 LI (n=63)
60
low Ki67 LI (n=77)
40
p<0.001
40
p=0.001
20
high Ki67 LI (n=42)
20
high Ki67 LI (n=68)
0
0
0
50
100
150
200
250
0
50
100
150
200
250
300
Time (months)
Time (months)
ADVANCE
ADVANCE ARTICLE: JCEM
ENDOCRINE SOCIETY
A
B
Progression-free survival (%)
100
100
VAV2 low-Ki67 LI low(=34)
VAV2 low-Ki67 LI low (=41)
VAV2 high-Ki67 LI low (=23)
VAV2 high-Ki67 LI low (=30)
VAV2 low-Ki67 LI high (=14)
VAV2 low-Ki67 LI high (=21)
80
VAV2 high-Ki67 LI high (=21)
Overall survival (%)
80
VAV2 high-Ki67 LI high (=39)
60
60
p=0.032
40
p=0.058
40
p=0.008
p=0.030
20
20
p<0.001
p<0.001
0
0
0
50
100
150
200
250
300
0
50
100
150
200
250
300
C
Time (months)
D
Time (months)
Progression-free survival (%)
100
100
80
Overall survival (%)
80
VAV2 low-Ki67 LI low (n=41)
VAV2 low-Ki67 LI low (n=34)
60
60
40
other (n=37)
40
other (n=51)
p=0.008
p=0.018
20
p<0.001
20
p<0.001
VAV2 high-Ki67 LI high (n=21)
VAV2 high-Ki67 LI high (n=39)
0
0
0
50
100
150
200
250
300
0
50
100
150
200
250
300
Time (months)
Time (months)
ADVANCE
A
100
Overall survival (%)
80
60
other (n=92)
40
Progression-free survival (%)
CLE
p<0.001
20
VAV2 high-Ki67 LI high (n=39)
0
0
50
100
150
200
250
300
B
Time (months)
C
100
100
80
Overall survival (%)
80
60
other (n=71)
60
other (n=71)
40
p<0.001
40
p=0.005
20
20
V2 high-Ki67 LI high (n=21)
VAV2 high-Ki67 LI high (n=21)
0
0
0
50
100
150
200
250
0
50
100
150
200
250
300
Time (months)
Time (months)
ADVANCE
ENDOCRINE SOCIETY