1. AntibodyCatalog #Company Desminab15200Abcam Smooth muscle actinab5694Abcam S100ab868Abcam Cytokeration(AE1/AE3)ab961Abcam P53sc-6243Santa Cruz Calponinab46794Abcam USP184813Cell Signaling MYC5605Cell Signaling pStat3(Y705)9145Cell Signaling Actinsc-1615Santa Cruz Supplementary Table 1: Antibodies used for immunohistochemistry studies.

2. Supplementary Table 2: Immunohistochemical staining of USP18 in clinical leiomyosarcoma samples. Case USP18 intensity % cells positive Histologic gradeSpecimen sitePrimary sitePrimary vs. recurrence 1140highIntraperitonealUterinemetastasis 200highSmall bowelAbdomenrecurrence 300highUterine primary 400highPelvisUterinerecurrence 500highHipReptroperitoneummetastasis 6280highColonUterinemetastasis 735highRight flankFlankprimary 8290intermediateThighSubcutisprimary 9180highSkin, abdomenSubcutisprimary 101 highThighSoft tissueprimary 11390intermediateLower legCutaneousrecurrence 1200intermediateSkin faceVenousmetastasis 132100highJejunumSmall bowelprimary 142100highRetroperitoneum primary 153100intermediatePelvisUterinemetastasis 16150highFascia, abdomenGImetastasis 17180highColon primary 18290highJejunum metastasis 193100highSmall bowel met/recurrence 20280highDermis primary 212100highRetroperitoneum primary

3. ----++++ USP18 Actin AB Supplementary Figure 1: A: Dystrophic calcifications in USP18 null mice. A representative image of a USP18-/- mouse is shown. B: KHC-2 cells with reconstituted USP18 expression maintained expression for the duration of growth in mice. Immunoblot analysis of protein isolated from 4 control and 4 USP18 overexpressing independent orthotopic sarcomas harvested from mice. The immunoblot showed representative analysis of KHC- 2 cells and this finding was also seen in KHC-1 cells (data not shown).

4. Supplementary Figure 2: USP18 null leiomyosarcoma cell lines are sensitive to treatment with the JAK2-STAT3 inhibitor, JSI-124. A: Immunoblot analysis of pSTAT3, STAT3, CDK4 and USP18 levels in KHC-1 cells with and without stably restored USP18 activity. B: Growth analysis of KHC-1 cells with JAK2-STAT3 inhibitor, JSI-124. Similar effects were seen in KHC-2 cells (data not shown). Validation of JSI-124 repression of JAK2-STAT3 pathway C: Immunoblot analyses of phosphorylated JAK2 (pJAK2), JAK2, and actin with relative level of pJAK2/JAK2 calculated relative to control. D: Immunoblot analysis of phosphorylated STAT3 (pSTAT3), STAT3, cyclin D1 and actin levels. pSTAT3 USP18 STAT3 CDK4 Actin pJAK2 (Y1007/1008) JAK2 Actin Time (min) 100nM 200nM pSTAT3 (Y701) Cyclin D1 STAT3 Actin Time(min ) 0 15 30 60 120 15 30 60 120 100nM 200nM 0 15 30 60 120 15 30 60 120 -+- --+ Empty vector USP18 A B C D 10.77 0.67 0.72 0.85 0.34 0.23 0.49 0.1 pJAK2/JAK2

5. Supplementary Figure 3: USP18 null leiomyosarcoma cell line KHC-1 and human leiomyosarcoma cell line SK-LMS-1 growth in response to interferon-β (500Units/ml IFNB) or doxycycline (0.2μM Dox) treatment over 3 days. Results expressed as fold relative to vehicle treated cells. Each experiment was performed in triplicate 3 separate times. KHC-1SK-LMS-1 0 0.2 0.4 0.6 0.8 1 24 48 72 vehicle IFNB Growth relative to vehicle vehicle Dox 0 0.2 0.4 0.6 0.8 1 24 48 72 244872 244872 Growth relative to vehicle 0 0.2 0.4 0.6 0.8 1 0 0.2 0.4 0.6 0.8 1 Hours A B **

6. 0 0.2 0.4 0.6 0.8 1 244872 Control (vehicle) KHC-1 KHC-1+Empty vector KHC-1+USP18 Growth with IFNB relative to vehicle N.S. Hours Supplementary Figure 4: USP18 null leiomyosarcoma cell line KHC-1 with restored USP18 expression did not affect response to IFN  (500Units/ml). Results expressed as fold relative to vehicle treated cells. (N.S. = not significant).