SLIDE 1
Somatic Mutation of clear cell Renal Cell Carcinoma (ccRCC)
(Sato Y. et al., Nat Genet, 2013)
New VHL Regulatory Pathways and Therapeutic Implications in ccRCC - - PowerPoint PPT Presentation
New VHL Regulatory Pathways and Therapeutic Implications in ccRCC - - PowerPoint PPT Presentation
14 th International VHL Medical/Research Symposium New VHL Regulatory Pathways and Therapeutic Implications in ccRCC Qing Zhang, Ph.D Associate Professor, Dept. Pathology UT Southwestern Medical Center 10/29/2020 Somatic Mutation of clear
SLIDE 2
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Choueiri TK and Kaelin WG., Nature Medicine, 2020
Target Therapies in Kidney Cancer
William Kaelin Toni Choueiri
SLIDE 4
Targeting VEGFR/PDGFR in Kidney Cancer
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“...we are ignoring other aspects of ccRCC that may need to be targeted to achieve maximal therapeutic efficacy....”
Linehan WM et al., Nature Reviews Clinical Oncology 10, 614–615, (2013)
Can we identify additional therapeutic avenues in ccRCC regulated by VHL? How?
- W. Marston Linehan (NCI)
Berton Zbar (NCI)
SLIDE 8
VHL and P-OH HIF1α-564 Binding
Based on the work from Kaelin, Ratcliffe and Pavletich
Hydroxylation of HIF1α on proline residue 564 (p-OH HIF) primes its binding with VHL ubiquitin E3 ligase complex (including pVHL, Elogin B and C, VBC)
SLIDE 9
~17,000 cDNA ORF bacterial stock 24 unique cDNA/pool, 700 pools 96 well plate, 8 plates
35S Methionine
In Vitro Translated Proteins Mini-Preps
A Genome-Wide In Vitro Expression Cloning Strategy
Marc Kirschner, HMS William Kaelin, DFCI
SLIDE 10
35S Labeled Proteins
700 pools (24 genes/pool) GST-VHL complex
x
P-OH HIF Peptide WT HIF Peptide
x
IVT GST-VHL complex WT P-OH
Run SDS-PAGE & autoradiography De-convolution Individual binding assay Identification of novel VHL substrates
Screening Strategy for Potential VHL Substrates
ZHX2 SFMBT1
SLIDE 11
Zhang J et al., Science, 2018, PMID 30026228
Zinc fingers and homoboxes protein 2 (ZHX2)
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Scm-Like With Four Malignant Brain Tumor Domains Protein 1 (SFMBT1)
Nady N et al., Journal of Molecular Biology, 2012
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VHL Regulates SFMBT1 Protein Levels
11/2/2020 13
Xijuan Liu, Ph.D
Mol Cell , Feb 4th, 2020
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SFMBT1 Depletion Leads to Decreased ccRCC Cell Growth
11/2/2020 14
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SFMBT1 Depletion Leads to Decreased ccRCC Tumor Growth
11/2/2020 15
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VHL mutant ccRCC Tumors Displays Upregulated SFMBT1 Protein Levels
Missense VHL mutant
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Integrated Analyses for SFMBT1 Regulated Events in ccRCC
Jeremy Simon Chris Fan
SPHK1
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SPHK1 is an SFMBT1 Direct Target Gene in ccRCC
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11/2/2020 19
SPHK1 is a Key SFMBT1 Downstream Target Gene in ccRCC
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11/2/2020 20
SPHK1 Inhibitor Represses ccRCC Cell Proliferation
PF-543: SPHK1 inhibitor
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SPHK1 Inhibitor Represses ccRCC Tumorigenesis
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Summary 1: VHL-SFMBT1-SPHK1 Signaling Drives Oncogenesis in ccRCC
Mol Cell, 2020, PMID 32023483
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How to Target ccRCC that are Mostly VHL Null
- 1. Target VHL regulated pathways, such as HIF2α, ZHX2 and SFMBT1
Challenge: It is difficult to target transcriptional factors Not all of ccRCC cell lines respond to HIF2α inhibitors (~50%)
- 2. Target synthetic lethality pathways for VHL loss
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Synthetic Lethality Targeting in Cancer
Brunen D and Bernards R., Nature Reviews Clinical Oncology 2017
VHL TBK1
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TBK1 in Innate Immunity
Liu s et al., Science, 2015
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TBK1 May be a Synthetic Lethality Partner for VHL Loss in ccRCC
Lianxin Hu, Ph.D
Cancer Discovery, 2020, PMID: 31810986
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TBK1 VHL WT
- r Normoxia
TBK1
OH
P
VHL
P
EglN1
A
TBK1
TBK1 P
OH
PPM1B
P
VHL Loss/Hypoxia Activates TBK1 Oncogenic Signaling in ccRCC
VHL Loss TBK1
P
TBK1
P
TBK1
OH
P
Hypoxia TBK1
P
TBK1
OH
P
EglN1 EglN1 VHL
PPM1B
B
Tumorigenesis
℗-p62
Cancer Discovery, 2020, PMID: 31810986
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11/2/2020 28
ccRCC Tumors Display Higher TBK1 Activity Compared to Paired Normals
Normal Tumor TBK1 pTBK1 Patient #1 Normal Tumor Patient #2
P=0.0016 P=0.0062 P=0.0062
TMA1 TMA2
Collaboration with Kan Gong, Peking University
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TBK1 Depletion Leads to Decreased ccRCC Tumor Growth and Metastasis
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DMSO CMPD1
0.0 0.5 1.0 1.5
Ev VHL
**
Fold change
TBK1 Inhibitor Selectively Suppresses VHL Null Cancer Cells
EV VHL UMRC6
Jenkins, R. W. et al., Cancer Discov 8 (2018).
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TBK1 PROTACs Selectively Suppresses VHL Null ccRCC Cell Growth
Frances Potjewyd, Ph.D Lindsey James, Ph.D
DMSO UNC6587
0.0 0.5 1.0 1.5 2.0
Ev VHL
**
Fold change
Cereblon
DMSO UNC6587 EV VHL UMRC6 DMSO UNC6587
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Summary II and Overall Future Directions
- 1. VHL Null cells are sensitive to TBK1 depletion/inhibition
- 2. VHL loss leads to hyper-activation of TBK1
- 3. ccRCC patients display increased TBK1 phosphorylation
- 4. TBK1 is essential for ccRCC tumorigenesis by phosphorylating p62 on Ser366
Study SFMBT1 signaling axis and its therapeutic implication in ccRCC Study the role of TBK1 on tumor and tumor microenvironment in ccRCC Potential combination therapies in kidney cancer
Future Directions
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