DOI: 10.5061/dryad.15dv41p3b
title: Identification of immune-related candidate biomarkers in plasma of patients with sporadic vestibular schwannoma

description: Vestibular schwannoma (VS) is an intracranial tumor arising from
 neoplastic Schwann cells, and typically presenting with hearing loss. The
 traditional belief that hearing deficit is caused by physical expansion of
 the VS, compressing the auditory nerve, does not explain the common
 clinical finding that patients with small tumors can have profound hearing
 loss, suggesting that tumor-secreted factors could influence hearing
 ability in VS patients. We conducted profiling of patients' plasma
 for 66 immune-related factors in patients with sporadic VS (N&gt;170)
 and identified and validated candidate biomarkers associated with tumor
 size (S100B) and hearing (MCP-3). We further identified a 9-biomarker
 panel (TNR-R2, MIF, CD30, MCP-3, IL-2R, BLC, TWEAK, eotaxin, S100B) with
 outstanding discriminatory ability for VS. These findings revealed
 possible therapeutic targets for VS, providing a unique diagnostic tool
 that may predict hearing change and tumor growth in VS patients, and may
 inform the timing of tumor resection to preserve hearing.
description: Study population and specimen collection
 From 07/2015–04/2021, blood was prospectively collected from
 patients undergoing VS resection at Massachusetts Eye and Ear (MEE) in
 Boston, MA on the day of surgery, typically within 30 minutes of inducing
 general anesthesia and ≥1 hour before tumor microdissection (discovery
 cohort). Blood was similarly prospectively collected from 08/2021–06/2023
 at Stanford Hospital in Palo Alto, CA (validation cohort). Blood from
 controls was collected at the Massachusetts General Hospital (MGH) Blood
 Donor Center in Boston, MA, and the Research Blood Components in
 Watertown, MA for the discovery cohort, and from the Stanford Hospital
 Blood Donor Center in Palo Alto, CA for the validation cohort. At
 collection, fresh blood was stored in EDTA vacutainer tubes (Becton
 Dickinson, NY, US) and kept at 4˚C without freezing. The whole blood
 samples were centrifuged at 2000 g for 10 min at 4°C. Plasma was separated
 and spun at 2000 g for 5 min at 4°C. Centrifuged plasma was filtered
 through 0.8 μm filter units (MF-Millipore MCE membrane, SLAA033SB;
 Millipore, Burlington, MA, US) and stored at -80°C until further
 use. Eligible patients had unilateral, sporadic VS that
 had not been previously resected or irradiated. Of 186 and 50 enrolled VS
 patients in the discovery and validation cohorts, 163 and 50 met inclusion
 criteria, respectively, and were included in the analyses for comparison
 with 70 and 43 controls, respectively. Clinical
 data Clinical and demographic data were collected from
 patient charts, operative reports, pathology reports, and pre-operative
 radiographic imaging. Patient variables included age at tissue collection;
 pre-surgical tumor volume measured via high-resolution axial
 contrast-enhanced T1-weighted brain MRI; internal auditory canal protocol;
 and pre-surgical pure-tone audiometric threshold and WR measurements. MRI
 and hearing tests were those nearest to resection, typically ≤3 months. WR
 was defined as the percentage of spoken monosyllabic words discernable
 from a list typically read at 70 dB or the level at which a patient's
 speech intelligibility curve plateaus. Pure-tone audiometric thresholds at
 0.5, 1, 2, and 3 kHz were used to calculate the PTA. Hearing groups were
 defined according to the American Academy of Otolaryngology–Head and Neck
 Surgery (AAO-HNS) Hearing Classification Guidelines. GH was defined as
 WR&gt;70% and PTA&lt;30 dB (AAO-HNS class A hearing). Otherwise,
 patients were classified as having PH (AAO-HNS class B, C and D hearing).
 A deaf ear was assigned a PTA of 125 dB and WR score of 0%. Serviceable
 hearing (SH) was defined as WR&gt;50% and PTA&lt;50 dB (AAO-HNS
 class A and B hearing). Otherwise, patients were classified as having
 non-serviceable hearing (NSH) (AAO-HNS class C and D hearing). Tumor
 resection was defined as GTR if there was complete tumor removal or a
 small tumor remnant no greater than 5x5x2 mm was left behind to preserve
 nerve integrity. Otherwise, it was defined as STR.
 Biomarker measurements Luminex (65
 cytokines, chemokines, growth factors, and soluble receptors);
 electrochemoluminescence (IL-18, TNF-α, and FGF-2); and ELISA (S100B)
 assays were conducted as described in detail below. Potential biomarkers
 were defined as those detectable in the plasma of &gt;75% of
 patients Luminex assay
 Simultaneous multiplex profiling of 65 immune-related factors
 composed of cytokines, chemokines, and growth factors was performed using
 a customized multiplex bead-based immunoassay - Immune Monitoring 65-Plex
 Human ProcartaPlex™ Panel (#EPX650-10065-901; ThermoFisher Scientific,
 Waltham, MA, US) according to manufacturer's instructions. The
 fluorescence-based signal was acquired on the Magpix instrument (Luminex,
 Austin, TX, US), and the values of analytes were calculated using
 ProcartaPlex Analyst 1.0 Software (ThermoFisher Scientific). The analytes
 with values below the lower limit of quantification in more than 75% of
 all samples were excluded from the analysis. For the accepted analytes,
 the values between 0 pg/mL and the lower limit of quantification, and
 those exceeding the upper limit of quantification, were approximated with
 the lowest and highest concentration representing these limits,
 respectively. Electrochemiluminescence
 assay The following candidate biomarkers were measured
 using electrochemiluminescence-based human assays from Meso Scale
 Diagnostics (MSD, Rockville, MD, US): IL-18 (U-PLEX Human IL-18), and
 TNF-α (U-PLEX Human TNF-α). All assays were conducted according to the
 manufacturer's protocols and signal detection was performed on the
 QuickPlex SQ 120 device (MSD). Pre-analytical data processing was done
 using MSD Discovery Workbench software (v4.0.12).
 ELISA assay The S100B ELISA Kit (#EZHS100B-
 33K; EMD Millipore, Billerica, MA, US) was used to measure S100B protein
 levels in the plasma of VS patients and controls. ELISA assays were
 performed by adhering to the manufacturer's protocol. Absorbance was
 measured using Spectra MAX 190 plate Reader (Molecular Devices, Sunnyvale,
 CA, US), and the standard curve was plotted using SoftMax Pro software
 (v5.2; Molecular Devices, San Jose, CA, US).
description: This README file was generated on 2023-10-24 by Sasa Vasilijic. GENERAL
 INFORMATION 1. Title of Dataset: Identification of Immune-Related
 Candidate Biomarkers in Plasma of Patients with Sporadic Vestibular
 Schwannoma 2. Author Information First author Contact Information Name:
 Sasa Vasilijic Institution: Stanford University School of Medicine
 Address: 801 Welch Rd, Palo Alto, CA 94304, USA Email:
 vasilije@stanford.edu 3. Date of sample/data collection (single date,
 range, approximate date): * Discovery cohort: 07/2015–04/2021 * Validation
 cohort: 08/2021-06/2023 Study subjects * patients with sporadic vestibular
 schwannoma organized in discovery and validation cohorts * healthy blood
 donors organized in discovery and validation cohorts Samples * human
 peripheral blood Data * demographics data: age and sex; * clinical
 preoperative data: concentration of immuno-related factors detected in the
 plasma, pure tone average, word recognition score, tumor volume;
 Abbreviations: VS: vestibular schwannoma PTA: pure tone average WR: word
 recognition 4. Geographic location of sample/ data collection: * Discovery
 cohort patients: Massachusetts Eye and Ear (MEE), Boston, MA, USA *
 Discovery cohort healthy controls: Research Blood Components, Watertown,
 MA, USA * Validation cohort patients: Stanford Hospital, Palo Alto, CA,
 USA * Validation cohort healthy controls: Stanford Hospital Blood Donor
 Center, Palo Alto, CA, USA 5. Information about funding sources that
 supported the collection of the data: National Institute on Deafness and
 Other Communication Disorders grant R01 DC015824; Bertarelli Foundation
 Endowed Professorship at Stanford University; Remondi Foundation, and
 Larry Bowman. SHARING/ACCESS INFORMATION 1. Licenses/restrictions placed
 on the data: CC0 1.0 Universal (CC0 1.0) Public Domain 2. Links to
 publications that cite or use the data: Vasilijic S, Atai NA, Hyakusoku H,
 Worthington S, Ren Y, Sagers JE, Sahin MI, Fujita T, Landegger LD, Lewis
 R, Welling DB, Stankovic KM. Identification of Immune-Related Candidate
 Biomarkers in Plasma of Patients with Sporadic Vestibular Schwannoma.
 Science Advances 2023, http://www.science.org/doi/10.1126/sciadv.adf7295.
 3. Links to other publicly accessible locations of the data: None 4.
 Links/relationships to ancillary data sets: None 5. Was data derived from
 another source? No A. If yes, list source(s): NA 6. Recommended citation
 for this dataset: Vasilijic S, Atai NA, Hyakusoku H, Worthington S, Ren Y,
 Sagers JE, Sahin MI, Fujita T, Landegger LD, Lewis R, Welling DB,
 Stankovic KM. Data from: Identification of Immune-Related Candidate
 Biomarkers in Plasma of Patients with Sporadic Vestibular Schwannoma.
 Dryad Digital Repository. https://doi.org/10.5061/dryad.15dv41p3b. DATA
 & FILE OVERVIEW 1. File List:
 adf7295\_manuscript\_data\_101723.xlsx 2. Relationship between files, if
 important: None 3. Additional related data collected that was not included
 in the current data package: None 4. Are there multiple versions of the
 dataset? No A. If yes, name of file(s) that was updated: NA i. Why was the
 file updated? NA ii. When was the file updated? NA
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_1A (Demographic and clinical
 characteristics of the VS patients and controls - Discovery cohort) 1.
 Number of variables * Vestibular schwannoma: 8 * Healthy donors: 8 2.
 Number of cases/rows * Vestibular schwannoma: 159 * Healthy donors: 70 3.
 Variable List: * Age (years) * Sex (F-female, M-male) * Tumor Size (cm3) *
 PTA (dB) Ipsilateral ear * WR (%) Ipsilateral ear * PTA (dB) Contralateral
 Ear * WR (%) Contralateral Ear * Hearing (good, poor) 4. Missing data
 codes: N/A (data not available) 5. Specialized formats or other
 abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_1B (Demographic and clinical
 characteristics of the VS patients and controls - Validation cohort) 1.
 Number of variables * Vestibular schwannoma: 8 * Healthy donors: 8 2.
 Number of cases/rows * Vestibular schwannoma: 50 * Healthy donors: 43 3.
 Variable List: * Age (years) * Sex (F-female, M-male) * Tumor Size (cm3) *
 PTA (dB) Ipsilateral ear * WR (%) Ipsilateral ear * PTA (dB) Contralateral
 Ear * WR (%) Contralateral Ear * Hearing (good, poor) 4. Missing data
 codes: N/A (data not available), incomplete data 5. Specialized formats or
 other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_2A (candidate biomarker plasma levels
 in VS patients and controls- Discovery cohort) 1. Number of variables *
 Vestibular schwannoma: 22 * Healthy donors: 22 2. Number of cases/rows *
 Vestibular schwannoma: 159 * Healthy donors: 70 3. Variable List: * Age
 (years) * Sex (F-female, M-male) * IL-2R (pg/mL) * SDF-1α (pg/mL) * TWEAK
 (pg/mL) * IL-18 (pg/mL) * CD30 (pg/mL) * APRIL (pg/mL) * IL-16 (pg/mL) *
 TNF-R2 (pg/mL) * MDC (pg/mL) * MCP-1 (pg/mL) * BLC (pg/mL) * MIF (pg/mL) *
 S-100B (pg/mL) * HGF (pg/mL) * Eotaxin-2 (pg/mL) * VEGF-A (pg/mL) * MCP-3
 (pg/mL) * IP-10 (pg/mL) * Eotaxin (pg/mL) * MCP-2 (pg/mL) 4. Missing data
 codes: NA (data not available) 5. Specialized formats or other
 abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_2B (candidate biomarker plasma levels
 in VS patients and controls- Validation cohort) 1. Number of variables *
 Vestibular schwannoma: 15 * Healthy donors: 15 2. Number of cases/rows *
 Vestibular schwannoma: 50 * Healthy donors: 43 3. Variable List: * Age
 (years) * Sex (F-female, M-male) * IL-2R (pg/mL) * SDF-1α (pg/mL) * TWEAK
 (pg/mL) * CD30 (pg/mL) * APRIL (pg/mL) * IL-16 (pg/mL) * TNF-R2 (pg/mL) *
 BLC (pg/mL) * MIF (pg/mL) * S100B (pg/mL) * MCP-3 (pg/mL) * Eotaxin
 (pg/mL) * MCP-2 (pg/mL) 4. Missing data codes: NA (data not available) 5.
 Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_2C (candidate biomarker plasma levels
 ratio between VS patients and controls - Discovery cohort) *** Data set-1
 1. Number of variables: 14 2. Number of cases/rows * Vestibular
 schwannoma: 53 3. Variable List: * Age (years) * Sex (F-female, M-male) *
 TNF-R2 (pg/mL) * IL-2R (pg/mL) * MCP-3 (pg/mL) * CD30 (pg/mL) * TWEAK
 (pg/mL) * MIF (pg/mL) * APRIL (pg/mL) * IL-16 (pg/mL) * SDF-1α (pg/mL) *
 MCP-2 (pg/mL) * Eotaxin (pg/mL) * BLC (pg/mL) 4. Missing data codes: NA
 (data not available) 5. Specialized formats or other abbreviations used:
 None *** Data set-2 1. Number of variables: 14 2. Number of cases/rows *
 Healthy donors: 53 3. Variable List: * Age (years) * Sex (F-female,
 M-male) * TNF-R2 (pg/mL) * IL-2R (pg/mL) * MCP-3 (pg/mL) * CD30 (pg/mL) *
 TWEAK (pg/mL) * MIF (pg/mL) * APRIL (pg/mL) * IL-16 (pg/mL) * SDF-1α
 (pg/mL) * MCP-2 (pg/mL) * Eotaxin (pg/mL) * BLC (pg/mL) 4. Missing data
 codes: NA (data not available) 5. Specialized formats or other
 abbreviations used: None *** Data set-3 Comment: The data set represents
 the plasma level ratio of biomarker concentrations by dividing values from
 data set-1 with values in data set-2. Cases/rows in data set-1 and data
 set-2 are arranged that represent sex and age-matched pairs. The outlier
 values of the plasma ratio (gray labeled values) were identified using the
 Rout method in GraphPad Prism (v9.3.1; GraphPad Software, La Jolla, CA,
 US) by setting Q to 1% (alpha to 0.01). The outlier values are omitted
 from the analysis. 1. Number of variables: 12 2. Number of cases/rows:53
 3. Variable List * TNF-R2 (ratio) * IL-2R (ratio) * MCP-3 (ratio) * CD30
 (ratio) * TWEAK (ratio) * MIF (ratio) * APRIL (ratio) * IL-16 (ratio) *
 SDF-1α (ratio) * MCP-2 (ratio) * Eotaxin (ratio) * BLC (ratio) 4. Missing
 data codes: NA (data not available) 5. Specialized formats or other
 abbreviations used: None *** Data set-4 1. Number of variables: 3 2.
 Number of cases/rows * Vestibular schwannoma: 36 3. Variable List: * Age
 (years) * Sex (F-female, M-male) * S100B (pg/mL) 4. Missing data codes: NA
 (data not available) 5. Specialized formats or other abbreviations used:
 None *** Data set-5 1. Number of variables: 3 2. Number of cases/rows *
 Healthy donors: 36 3. Variable List: * Age (years) * Sex (F-female,
 M-male) * S100B (pg/mL) 4. Missing data codes: NA (data not available) 5.
 Specialized formats or other abbreviations used: None *** Data set-6
 Comment: The data set represents the plasma level ratio of biomarker
 concentrations by dividing values from data set-4 with values in data
 set-5. Cases/rows in data set-4 and data set-5 are arranged that represent
 sex and age-matched pairs. The outlier values of the plasma ratio (gray
 labeled values) were identified using the Rout method in GraphPad Prism
 (v9.3.1; GraphPad Software, La Jolla, CA, US) by setting Q to 1% (alpha to
 0.01). The outlier values are omitted from the analysis 1. Number of
 variables: 1 2. Number of cases/rows: 36 3. Variable List: * S100B (ratio)
 4. Missing data codes: NA (data not available) 5. Specialized formats or
 other abbreviations used: None *** Data set-7 (Fig\_2C data) 1. Number of
 variables: 13 2. Number of cases/rows:53 Comment: Data set-3 and data
 set-6 were pooled, the outlier values were omitted and their place in a
 new data set is indicated with gray-colored cells named
 "Outlier". 3. Variable List * TNF-R2 (ratio) * IL-2R (ratio) *
 MCP-3 (ratio) * CD30 (ratio) * TWEAK (ratio) * MIF (ratio) * APRIL (ratio)
 * S100B (ratio) * IL-16 (ratio) * SDF-1α (ratio) * MCP-2 (ratio) * Eotaxin
 (ratio) * BLC (ratio) 4. Missing data codes: NA (data not available) 5.
 Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_2D (candidate biomarker plasma levels
 ratio between VS patients and controls - Validation cohort) *** Data set-1
 1. Number of variables: 15 2. Number of cases/rows * Vestibular
 schwannoma: 32 3. Variable List: * Age (years) * Sex (F-female, M-male) *
 TNF-R2 (pg/mL) * MIF (pg/mL) * APRIL (pg/mL) * SDF-1α (pg/mL) * IL-2R
 (pg/mL) * MCP-2 (pg/mL) * CD30 (pg/mL) * IL-16 (pg/mL) * TWEAK (pg/mL) *
 BLC (pg/mL) * Eotaxin (pg/mL) * MCP-3 (pg/mL) * S100B (pg/mL) 4. Missing
 data codes: NA (data not available) 5. Specialized formats or other
 abbreviations used: None *** Data set-2 1. Number of variables: 15 2.
 Number of cases/rows * Healthy donors: 32 3. Variable List: * Age (years)
 * Sex (F-female, M-male) * TNF-R2 (pg/mL) * MIF (pg/mL) * APRIL (pg/mL) *
 SDF-1α (pg/mL) * IL-2R (pg/mL) * MCP-2 (pg/mL) * CD30 (pg/mL) * IL-16
 (pg/mL) * TWEAK (pg/mL) * BLC (pg/mL) * Eotaxin (pg/mL) * MCP-3 (pg/mL) *
 S100B (pg/mL) 4. Missing data codes: NA (data not available) 5.
 Specialized formats or other abbreviations used: None *** Data set-3
 Comment: The data set represents the plasma level ratio of biomarker
 concentrations by dividing values from data set-1 with values in data
 set-2. Cases/rows in data set-1 and data set-2 are arranged that represent
 sex and age-matched pairs. The outlier values of the plasma ratio (gray
 labeled values) were identified using the Rout method in GraphPad Prism
 (v9.3.1; GraphPad Software, La Jolla, CA, US) by setting Q to 1% (alpha to
 0.01). The outlier values are omitted from the analysis. 1. Number of
 variables: 13 2. Number of cases/rows:32 3. Variable List * TNF-R2 (ratio)
 * MIF (ratio) * APRIL (ratio) * SDF-1α (ratio) * IL-2R (ratio) * MCP-2
 (ratio) * CD30 (ratio) * IL-16 (ratio) * TWEAK (ratio) * BLC (ratio) *
 Eotaxin (ratio) * MCP-3 (ratio) * S100B (ratio) 4. Missing data codes: NA
 (data not available) 5. Specialized formats or other abbreviations used:
 None *** Data set-4 (Fig\_2D data) Comment: The data set represents the
 data for Fig\_2D and contains the data from data set-3 whose outlier
 values were omitted and their place is indicated with gray-colored cells
 named "Outlier". 1. Number of variables: 13 2. Number of
 cases/rows:32 3. Variable List * TNF-R2 (ratio) * MIF (ratio) * APRIL
 (ratio) * SDF-1α (ratio) * IL-2R (ratio) * MCP-2 (ratio) * CD30 (ratio) *
 IL-16 (ratio) * TWEAK (ratio) * BLC (ratio) * Eotaxin (ratio) * MCP-3
 (ratio) * S100B (ratio) 4. Missing data codes: NA (data not available) 5.
 Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_3A, B, C (Association of MCP-3 levels
 and preoperative hearing- Discovery cohort) 1. Number of variables: 6 2.
 Number of cases/rows:78 3. Variable List * MCP-3 (pg/mL) * Sex (F-female,
 M-male) * Age (years) * WR (%) Ipsilateral ear * WR (%) Contralateral Ear
 * Tumor volume (cm3) 4. Missing data codes: N/A (data not available) 5.
 Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_3D, E (MCP-3 levels and hearing
 serviceability- Discovery cohort) 1. Number of variables: 6 2. Number of
 cases/rows * VS patients with serviceable hearing: 30 * VS patients with
 nonserviceable hearing: 45 3. Variable List * MCP-3 (pg/mL) * Sex
 (F-female, M-male) * Age (years) * Tumor volume (cm3) * PTA (dB)
 Ipsilateral Ear * WR (%) Ipsilateral ear 4. Missing data codes: N/A (data
 not available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_3F (WR and hearing serviceability-
 Discovery cohort) 1. Number of variables: 3 2. Number of cases/rows * VS
 patients with serviceable hearing: 59 * VS patients with nonserviceable
 hearing: 59 3. Variable List * Age (years) * Sex (F-female, M-male) * WR
 (%) Ipsilateral ear 4. Missing data codes: N/A (data not available) 5.
 Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_3G, H, I (Association of MCP-3 levels
 and preoperative hearing- Validation cohort-A) 1. Number of variables: 6
 2. Number of cases/rows:50 3. Variable List * MCP-3 (pg/mL) * Sex
 (F-female, M-male) * Age (years) * WR (%) Ipsilateral ear * WR (%)
 Contralateral Ear * Tumor volume (cm3) 4. Missing data codes: N/A (data
 not available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_3J, K (MCP-3 levels and hearing
 serviceability- Validation cohort-A) 1. Number of variables: 6 2. Number
 of cases/rows * VS patients with serviceable hearing: 22 * VS patients
 with nonserviceable hearing: 23 3. Variable List * MCP-3 (pg/mL) * Sex
 (F-female, M-male) * Age (years) * Tumor volume (cm3) * PTA (dB)
 Ipsilateral Ear * WR (%) Ipsilateral ear 4. Missing data codes: N/A (data
 not available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_3L (WR and hearing serviceability-
 Validation cohort-A) 1. Number of variables: 3 2. Number of cases/rows *
 VS patients with serviceable hearing: 15 * VS patients with nonserviceable
 hearing: 15 3. Variable List * Age (years) * Sex (F-female, M-male) * WR
 (%) Ipsilateral ear 4. Missing data codes: N/A (data not available) 5.
 Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_3M, N, O (Association of MCP-3 levels
 and preoperative hearing- Validation cohort-B) 1. Number of variables: 6
 2. Number of cases/rows:63 3. Variable List * MCP-3 (pg/mL) * Sex
 (F-female, M-male) * Age (years) * WR (%) Ipsilateral ear * WR (%)
 Contralateral Ear * Tumor volume (cm3) 4. Missing data codes: N/A (data
 not available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_3P, Q (MCP-3 levels and hearing
 serviceability- Validation cohort-B) 1. Number of variables: 6 2. Number
 of cases/rows * VS patients with serviceable hearing: 27 * VS patients
 with nonserviceable hearing: 30 3. Variable List * MCP-3 (pg/mL) * Sex
 (F-female, M-male) * Age (years) * Tumor volume (cm3) * PTA (dB)
 Ipsilateral Ear * WR (%) Ipsilateral ear 4. Missing data codes: N/A (data
 not available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_3R (WR and hearing serviceability-
 Validation cohort-B) 1. Number of variables: 3 2. Number of cases/rows *
 VS patients with serviceable hearing: 26 * VS patients with nonserviceable
 hearing: 26 3. Variable List * Age (years) * Sex (F-female, M-male) * WR
 (%) Ipsilateral ear 4. Missing data codes: N/A (data not available) 5.
 Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_4A, B, C (Association of S100B levels
 and preoperative tumor volume- Discovery cohort) 1. Number of variables: 4
 2. Number of cases/rows:122 3. Variable List * S100B (pg/mL) * Sex
 (F-female, M-male) * Age (years) * Tumor volume (mm3) 4. Missing data
 codes: N/A (data not available) 5. Specialized formats or other
 abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_4D, E (S100B levels and tumor
 resection- Discovery cohort) 1. Number of variables: 4 2. Number of
 cases/rows * VS patients with gross tumor resection: 88 * VS patients with
 subtotal tumor resection: 34 3. Variable List * S100B (pg/mL) * Sex
 (F-female, M-male) * Age (years) * Tumor volume (mm3) 4. Missing data
 codes: N/A (data not available) 5. Specialized formats or other
 abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_4F (Tumor volume and tumor resection-
 Discovery cohort) 1. Number of variables: 3 2. Number of cases/rows * VS
 patients with gross tumor resection: 36 * VS patients with subtotal tumor
 resection: 36 3. Variable List * Age (years) * Sex (F-female, M-male) *
 Tumor volume (mm3) 4. Missing data codes: N/A (data not available) 5.
 Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_4G, H, I (Association of S100B levels
 and preoperative tumor volume- Validation cohort-A) 1. Number of
 variables: 4 2. Number of cases/rows:50 3. Variable List * S100B (pg/mL) *
 Sex (F-female, M-male) * Age (years) * Tumor volume (mm3) 4. Missing data
 codes: N/A (data not available) 5. Specialized formats or other
 abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_4J, K (S100B levels and tumor
 resection- Validation cohort-A) 1. Number of variables: 4 2. Number of
 cases/rows * VS patients with gross tumor resection: 17 * VS patients with
 subtotal tumor resection: 11 3. Variable List * S100B (pg/mL) * Sex
 (F-female, M-male) * Age (years) * Tumor volume (mm3) 4. Missing data
 codes: N/A (data not available) 5. Specialized formats or other
 abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_4L (Tumor volume and tumor resection-
 Validation cohort-A) 1. Number of variables: 3 2. Number of cases/rows *
 VS patients with gross tumor resection: 11 * VS patients with subtotal
 tumor resection: 11 3. Variable List * Age (years) * Sex (F-female,
 M-male) * Tumor volume (mm3) 4. Missing data codes: N/A (data not
 available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_4M, N, O (Association of S100B levels
 and preoperative tumor volume- Validation cohort-B) 1. Number of
 variables: 4 2. Number of cases/rows:86 3. Variable List * S100B (pg/mL) *
 Sex (F-female, M-male) * Age (years) * Tumor volume (mm3) 4. Missing data
 codes: N/A (data not available) 5. Specialized formats or other
 abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_4P, Q (S100B levels and tumor
 resection- Validation cohort-B) 1. Number of variables: 4 2. Number of
 cases/rows * VS patients with gross tumor resection: 44 * VS patients with
 subtotal tumor resection: 20 3. Variable List * S100B (pg/mL) * Sex
 (F-female, M-male) * Age (years) * Tumor volume (mm3) 4. Missing data
 codes: N/A (data not available) 5. Specialized formats or other
 abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_4R (Tumor volume and tumor resection-
 Validation cohort-B) 1. Number of variables: 3 2. Number of cases/rows *
 VS patients with gross tumor resection: 17 * VS patients with subtotal
 tumor resection: 17 3. Variable List * Age (years) * Sex (F-female,
 M-male) * Tumor volume (mm3) 4. Missing data codes: N/A (data not
 available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_5A (data used in ROC analysis-
 Discovery cohort) 1. Number of variables: 11 2. Number of cases/rows *
 Vestibular schwannoma: 24 * Healthy donors: 24 3. Variable List * Age
 (years) * Sex (F-female, M-male) * TNF-R2 (pg/mL) * MIF (pg/mL) * CD30
 (pg/mL) * MCP-3 (pg/mL) * IL-2R (pg/mL) * BLC (pg/mL) * TWEAK (pg/mL) *
 Eotaxin (pg/mL) * S-100B (pg/mL) 4. Missing data codes: N/A (data not
 available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_5B (data used in ROC analysis-
 Validation cohort) 1. Number of variables: 11 2. Number of cases/rows *
 Vestibular schwannoma: 32 * Healthy donors: 32 3. Variable List * Age
 (years) * Sex (F-female, M-male) * TNF-R2 (pg/mL) * MIF (pg/mL) * CD30
 (pg/mL) * MCP-3 (pg/mL) * IL-2R (pg/mL) * BLC (pg/mL) * TWEAK (pg/mL) *
 Eotaxin (pg/mL) * S-100B (pg/mL) 4. Missing data codes: N/A (data not
 available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_6A, C (data used in combinatorial ROC
 analysis, 10-fold cross-validation test, and AUC permutation test-
 Discovery cohort) 1. Number of variables: 11 2. Number of cases/rows *
 Vestibular schwannoma: 24 * Healthy donors: 24 3. Variable List * Age
 (years) * Sex (F-female, M-male) * TNF-R2 (pg/mL) * MIF (pg/mL) * CD30
 (pg/mL) * MCP-3 (pg/mL) * IL-2R (pg/mL) * BLC (pg/mL) * TWEAK (pg/mL) *
 Eotaxin (pg/mL) * S-100B (pg/mL) 4. Missing data codes: N/A (data not
 available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_6B, D (data used in combinatorial ROC
 analysis, 10-fold cross-validation test, and AUC permutation test-
 Validation cohort) 1. Number of variables: 11 2. Number of cases/rows *
 Vestibular schwannoma: 32 * Healthy donors: 32 3. Variable List * Age
 (years) * Sex (F-female, M-male) * TNF-R2 (pg/mL) * MIF (pg/mL) * CD30
 (pg/mL) * MCP-3 (pg/mL) * IL-2R (pg/mL) * BLC (pg/mL) * TWEAK (pg/mL) *
 Eotaxin (pg/mL) * S-100B (pg/mL) 4. Missing data codes: N/A (data not
 available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_S2A (Hearing characteristics of VS
 patients- Discovery cohort) 1. Number of variables: 16 2. Number of
 cases/rows * VS patients: 154 3. Variable List * Audiometric threshold
 (dB) at 250 Hz Ipsilateral ear * Audiometric threshold (dB) at 500 Hz
 Ipsilateral ear * Audiometric threshold (dB) at 1000 Hz Ipsilateral ear *
 Audiometric threshold (dB) at 2000 Hz Ipsilateral ear * Audiometric
 threshold (dB) at 3000 Hz Ipsilateral ear * Audiometric threshold (dB) at
 4000 Hz Ipsilateral ear * Audiometric threshold (dB) at 8000 Hz
 Ipsilateral ear * Audiometric threshold (dB) at 250 Hz Contralateral ear *
 Audiometric threshold (dB) at 500 Hz Contralateral ear * Audiometric
 threshold (dB) at 1000 Hz Contralateral ear * Audiometric threshold (dB)
 at 2000 Hz Contralateral ear * Audiometric threshold (dB) at 3000 Hz
 Contralateral ear * Audiometric threshold (dB) at 4000 Hz Contralateral
 ear * Audiometric threshold (dB) at 8000 Hz Contralateral ear * WR (%)
 Ipsilateral ear * WR (%) Contralateral ear 4. Missing data codes: N/A
 (data not available) 5. Specialized formats or other abbreviations used:
 None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_S2B (Hearing characteristics of VS
 patients- Validation cohort) 1. Number of variables: 16 2. Number of
 cases/rows * VS patients: 33 3. Variable List * Audiometric threshold (dB)
 at 250 Hz Ipsilateral ear * Audiometric threshold (dB) at 500 Hz
 Ipsilateral ear * Audiometric threshold (dB) at 1000 Hz Ipsilateral ear *
 Audiometric threshold (dB) at 2000 Hz Ipsilateral ear * Audiometric
 threshold (dB) at 3000 Hz Ipsilateral ear * Audiometric threshold (dB) at
 4000 Hz Ipsilateral ear * Audiometric threshold (dB) at 8000 Hz
 Ipsilateral ear * Audiometric threshold (dB) at 250 Hz Contralateral ear *
 Audiometric threshold (dB) at 500 Hz Contralateral ear * Audiometric
 threshold (dB) at 1000 Hz Contralateral ear * Audiometric threshold (dB)
 at 2000 Hz Contralateral ear * Audiometric threshold (dB) at 3000 Hz
 Contralateral ear * Audiometric threshold (dB) at 4000 Hz Contralateral
 ear * Audiometric threshold (dB) at 8000 Hz Contralateral ear * WR (%)
 Ipsilateral ear * WR (%) Contralateral ear 4. Missing data codes: N/A
 (data not available) 5. Specialized formats or other abbreviations used:
 None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_S3 1. Number of variables: 15 2.
 Number of cases/rows * Vestibular schwannoma: 159 * Healthy donors: 70 3.
 Variable List: * Age (years) * Sex (F-female, M-male) * MCP-3 (pg/mL) *
 CD30 (pg/mL) * IL-2R (pg/mL) * TWEAK (pg/mL) * TNF-R2 (pg/mL) * S-100B
 (pg/mL) * MIF (pg/mL) * BLC (pg/mL) * SDF-1α (pg/mL) * MCP-2 (pg/mL) *
 Eotaxin (pg/mL) * APRIL (pg/mL) * IL-16 (pg/mL) 4. Missing data codes: NA
 (data not available) 5. Specialized formats or other abbreviations used:
 None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_S4 1. Number of variables: 15 2.
 Number of cases/rows * Vestibular schwannoma: 159 * Good hearing: 34 *
 Poor hearing: 121 * Healthy donors: 70 3. Variable List: * Age (years) *
 Sex (F-female, M-male) * IL-2R (pg/mL) * SDF-1α (pg/mL) * TWEAK (pg/mL) *
 CD30 (pg/mL) * IL-16 (pg/mL) * APRIL (pg/mL) * TNF-R2 (pg/mL) * BLC
 (pg/mL) * MIF (pg/mL) * S-100B (pg/mL) * MCP-3 (pg/mL) * Eotaxin (pg/mL) *
 MCP-2 (pg/mL) 4. Missing data codes: NA (data not available) 5.
 Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_S5A 1. Number of variables: 13 2.
 Number of cases/rows * Vestibular schwannoma (Poor hearing): 121 * Healthy
 donors: 70 3. Variable List: * Age (years) * Sex (F-female, M-male) *
 IL-2R (pg/mL) * SDF-1α (pg/mL) * TWEAK (pg/mL) * CD30 (pg/mL) * TNF-R2
 (pg/mL) * BLC (pg/mL) * MIF (pg/mL) * S-100B (pg/mL) * MCP-3 (pg/mL) *
 Eotaxin (pg/mL) * MCP-2 (pg/mL) 4. Missing data codes: NA (data not
 available) 5. Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_S5B 1. Number of variables: 14 2.
 Number of cases/rows * Vestibular schwannoma (Good hearing): 34 * Healthy
 donors: 70 3. Variable List: * Age (years) * Sex (F-female, M-male) *
 IL-2R (pg/mL) * SDF-1α (pg/mL) * TWEAK (pg/mL) * CD30 (pg/mL) * APRIL
 (pg/mL) * IL-16 (pg/mL) * TNF-R2 (pg/mL) * BLC (pg/mL) * MIF (pg/mL) *
 S-100B (pg/mL) * MCP-3 (pg/mL) * Eotaxin (pg/mL) 4. Missing data codes: NA
 (data not available) 5. Specialized formats or other abbreviations used:
 None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_S6 1. Number of variables: 18 2.
 Number of cases/rows * VS patients with serviceable hearing: 71 * VS
 patients with nonserviceable hearing: 79 3. Variable List: * IL-2R (pg/mL)
 * SDF-1α (pg/mL) * TWEAK (pg/mL) * CD30 (pg/mL) * APRIL (pg/mL) * IL-16
 (pg/mL) * TNF-R2 (pg/mL) * BLC (pg/mL) * MIF (pg/mL) * S-100B (pg/mL) *
 MCP-3 (pg/mL) * Eotaxin (pg/mL) * MCP-2 (pg/mL) * Sex (F-female, M-male) *
 Age (years) * Tumor volume (cm3) * PTA (dB) Ipsilateral ear * WR (%)
 Ipsilateral ear 4. Missing data codes: NA (data not available) 5.
 Specialized formats or other abbreviations used: None
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_S7 (Discovery cohort) *** Data set-1
 1. Number of variables: 11 2. Number of cases/rows * Vestibular
 schwannoma: 24 * Healthy donors: 24 3. Variable List * Age (years) * Sex
 (F-female, M-male) * S-100B (pg/mL) * BLC (pg/mL) * CD30 (pg/mL) * IL-2R
 (pg/mL) * MCP-3 (pg/mL) * MIF (pg/mL) * TNF-R2 (pg/mL) * TWEAK (pg/mL) *
 Eotaxin (pg/mL) 4. Missing data codes: N/A (data not available) 5.
 Specialized formats or other abbreviations used: None *** Data set-2, data
 set-3 Comment: The data sets represent a correlation matrix composed of
 Spearman r coefficients based on the correlation analysis performed for
 the data set-1, using GraphPad Prism (v9.3.1; GraphPad Software, La Jolla,
 CA, US). Correlation coefficients were used for the construction of the
 correlation networks shown in Fig\_S7, using Cytoscape (Shannon P. et al.
 Cytoscape: a software environment for integrated models of biomolecular
 interaction networks Genome Research 2003 Nov; 13(11):2498-504).
 #########################################################################
 DATA-SPECIFIC INFORMATION FOR: Fig\_S7 (Validation cohort) *** Data set-1
 1. Number of variables: 11 2. Number of cases/rows * Vestibular
 schwannoma: 32 * Healthy donors: 32 3. Variable List * Age (years) * Sex
 (F-female, M-male) * S-100B (pg/mL) * BLC (pg/mL) * CD30 (pg/mL) * IL-2R
 (pg/mL) * MCP-3 (pg/mL) * MIF (pg/mL) * TNF-R2 (pg/mL) * TWEAK (pg/mL) *
 Eotaxin (pg/mL) 4. Missing data codes: N/A (data not available) 5.
 Specialized formats or other abbreviations used: None *** Data set-2, data
 set-3 Comment: The data sets represent a correlation matrix composed of
 Spearman r coefficients based on the correlation analysis performed for
 the data set-1, using GraphPad Prism (v9.3.1; GraphPad Software, La Jolla,
 CA, US). Correlation coefficients were used for the construction of the
 correlation networks shown in Fig\_S7, using Cytoscape (Shannon P. et al.
 Cytoscape: a software environment for integrated models of biomolecular
 interaction networks Genome Research 2003 Nov; 13(11):2498-504).
 #########################################################################