Lyn Tyrosine Kinase Regulates Androgen Receptor Expression and Activity in Castrate Resistant Prostate Cancer Anousheh Zardan, Ka Mun Nip, Daksh Thaper, Paul Toren, Sepideh Vahid, Eliana Beraldi, Ladan Fazli, Francois Lamoureux, Kilian Gust, Michael E. Cox, Jennifer L. Bishop and Amina Zoubeidi Supplementary Figures

Supplementary Figure S1: CRPCNormalPrimary PCa Src Supplementary Figure S1: Src expression in the progression of PCa. Src expression was evaluated in human tissue specimens by immunohistochemistry from normal prostate specimens or patients with primary hormone naïve PCa or CRPC (TURP-Transurethral Resection of Prostate).

Supplementary Figure S2: : Lyn overexpression increases AR transcriptional activity: A) Lyn expression in different stably transfected Lyn clones: Proteins were extracted from LNCaP Lyn-Clone#A, LNCaP Lyn-Clone#B and LNCaP Mock cells and western blot was performed using Lyn antibody, vinculin was used as a loading control. B: LNCaP Lyn-Clone #A, LNCaP Lyn-Clone#B and LNCaP Mock cells were transfected with Probasin-luciferase and were treated +/- R1881 for 24h PSA luciferase activity was determined. The results were reported as mean ± SD; *p<0.05 and ***P< Mock Vinculin Lyn Clone#A Clone#B Lyn overexpression A B Supplementary Figure S2:

Supplementary Figure S3: Lyn knockdown inhibits AR transcriptional activity: A) Lyn expression in different shRNA-Lyn stably transfected clones: Proteins were extracted from LNCaP sh-Lyn#1, LNCaP sh-Lyn#2 and LNCaP sh-Ctr cells and western blot was performed using Lyn antibody, vinculin was used as a loading control. B: from LNCaP sh-Lyn#1, LNCaP sh-Lyn#2 and LNCaP sh-Ctr cells were transfected with Probasin-luciferase and were treated +/- R1881 for 24h PSA luciferase activity was determined. The results were reported as mean ± SD and ***P< sh-Ctr sh-Lyn#1 sh-Lyn#2 Vinculin Lyn A B Supplementary Figure S3:

Supplementary Figure S4: Lyn AR PSA Vinculin LNCaP CtrLyn #1Lyn #2 siRNA Supplementary Figure S4: Lyn knockdown affects AR expression: LNCaP cells were transfected with 10nM of siRNA control, siRNA-Lyn #1 or siRNA-Lyn #2. 48 hours post transfection, total proteins were extracted and western blot was performed Lyn, PSA and AR antibodies and Vinculin was used as a loading control.

Supplementary Figure S5: Supplemental Figure S5: Src kinase siRNA treatment does not affect AR expression. LNCaP cells were transfected with different concentrations of siRNA control, siRNA-Src. 48 hours post transfection, total proteins were extracted and western blot was performed Src, PSA and AR antibodies and Vinculin was used as a loading control. Src kinase AR PSA Vinculin (nM siRNA) CtrSrc kinase

Supplementary Figure S6: Supplemental Figure S6: Lyn knockdown does not affect cell growth: LNCaP sh-Lyn and LNCaP sh-Ctr were grown in FBS and cell growth was assessed using crystal violet over time.

LYNMeanSTDEVCoresCaseSQRTSE BPH Untreated cancer CRPC_TURP Table S1: Statistical data from Lyn TMA analysis Supplementary Table S1:

SrcMeanSTDEVCoresCaseSQRTSE BPH Untreated cancer CRPC_TURP Table S2: Statistical data from Src TMA analysis Supplementary Table S2: