Dr. Qun Lu Leo Jenkins Cancer Center at BSOM UNC Lineberger Comprehensive Cancer Center Contextual Contributions of Oncogenic and Tumor Suppressive Pathways: - Our Opportunities and Challenges in Therapeutic Development -

Our Progresses in The Past Four Decades * * * * # # #  Target specific drugs such as Gleevec and Iressa By specifically blocking the abnormal protein called BCR-ABL, Gleevec kills the leukemia cells. By inhibiting tyrosine kinase activity of EGFR, Iressa suppresses NSCLC. Remarkable Treatment Successes  Cytotoxic chemotherapy drugs such as Cisplatin Before cisplatin was available, metastatic testicular cancer had a mortality rate of 95%. Cisplatin based regimens resulted in a 60% cure rate for this disease.

What are the modern strategies for drug discovery against cancer?  What are the modern strategies for drug discovery against cancer? Most strategies for cancer drug discovery focus on attacking specific elements of a given biological pathway of cancer cells to achieve target specificity with minimized toxicity. This approach, although with long term merits, has not produced the desired numbers of viable drugs because most such leads succeeded in vitro or in animal studies do not show the same efficacy in human trials. Consequently, few viable cancer drugs enter markets in recent years leading to skyrocketed R&D cost. Billions of dollars have been invested on R&D, why are we still so far away from conquering cancer?

(2009) (2010) (2010) Dichotomy of Oncogenes and Tumor Suppressors Challenges Anti-cancer Therapies

Surprising Discoveries in Cancer Research  Oncogenic Versus Tumor Suppressive Signaling Pathways When oncogenes were identified, we thought that we have found the bad guys. So the theory was that if we inhibit the functions of oncogenes, we can stop the disease. It was also thought that since there are tumor suppressors, we can increase the function of the tumor suppressors to inhibit the disease progression. However: Many genes and many signaling pathways can be oncogenic in one context, but be tumor suppressive at a different context or at different stages.

Myc-pRb-E2F axis Myc oncogene expression is deregulated in 15 to 30 % of human cancers, resulting in elevated levels of Myc protein.  As mitogenic promoter: Myc perturbs the decision to phosphorylate tumor suppressor pRb and thereby alter cell cycle progression.  As pro-apoptotic factor: Myc induces cell to undergo apoptosis unless they have been protected from apoptosis by some other, previously acquired anti-apoptotic allele (e.g., a ras oncogene). In the presence of the anti-apoptotic mutation, the strong mitogenic effects of the myc oncogene then become apparent.  As proliferation promoter: both Myc and Ras can activate E2F transcription factor, which can initiate cell proliferation.  As pro-apoptotic factor: both Myc and Ras can also activate E2F transcription factor, which can initiate cell death. Indeed, the oncogenic roles of E2F could be context-dependent. Zhang et al., Future Oncology

TGF  1 signaling axis TGF  1 signaling axis  As a tumor suppressor: for most normal epithelial cells and at the early stages of tumor development. This process occurs largely through the activation of TGF  1 receptor complex and phosphorylation of Smads and other signaling pathways.  As metastasis promoter: elevated expression of TGF  1 may enhance malignant properties of tumor cells through effects on cell invasion, metastasis, epithelial to mesenchymal transition (EMT), or anti-tumor immunity. This alteration in functions of TGF  1 in cancer development is likely due to its interaction with other signaling pathways such as ras oncogene that becomes activated during tumor progression. Phosphatidylinositol 3-kinases (PI3Ks) signaling axis  Oncogenic functions of PI3K: In cell-based assays, PIK3CA mutations confer a gain of function as measured by lipid kinase activity, constitutive activation of Akt, and cellular transformation. Therefore, the frequency of PIK3CA mutation is consistent with the significance of PI3Ks in breast cancer pathogenesis.  Positive prognostic significance of PIK3CA mutations: Some studies indicate that PIK3CA can even be a “good” activating mutation in cancer. This is clinically relevant, because it could significantly affect the design of clinical trials planned for PI3K-targeted therapy. Zhang et al., Future Oncology

 -Catenin p120-catenin p0071 ARVCF  -Catenin β-Catenin Armadillo 843 aa 781 aa 745 aa 968 aa 962 aa 1211 aa 1225 aa  -Catenin in comparison to other armadillo domain containing proteins The Armadillo/β-catenin superfamily DSWV

Incompatible Observations on Human tissues and Cultured Cell Lines - Make drug discovery more challenging Esophagus 食管 Benign Cancer Breast 乳腺 Prostate 前列腺 Lung Lu et al., 2005 (180 cases TMA) Zhang et al., cases NSCLC

Promoter Mutations Can Increase  -CateninExpression Promoter Mutations Can Increase  -Catenin Expression in Prostate Cancer (PCa) Cells Wang et al., Oncogene

In Culture,  -Catenin Expression Is Moderate But Its Expression Increases PCa Cell Viability and Tumor Xenograft Growth Zeng et al., Mole Cancer

MDCK Cells However,  -Catenin Also Displays Tumor Suppressive Activities Lu et al., 1999; J. Cell BiolWestbrook et al., 2005; Cell HMEC Cells PCa Cells Zeng and Lu, unpublished

So, Why Does  -Catenin Display Opposite Functions ? Does Its Mutations Play a Role in PCa Cell Fate and Behavior? -Identification of  -Catenin Mutations in Cancer- Nopparat and Zhang et al., Oncogene

 -Catenin 1225 aa Identification of Functional  -Catenin Mutations in Human Prostate Cancer , , 816  Mutations occur at very high frequency;  Mutations occur in other cancer types as well;  May affect  -catenin functions. Patent pending

Ovary cancer: P1218L (Primary tumor) Large intestine: Y1128fs*6 (Culture sample) Large intestine: P1159S (Culture sample) NSCLC: K703R (Culture sample) Pancreas cancer: A685V (Culture sample) Skin cancer: F1172F (Culture sample) Majority of Cancer Associated Mutations Are Limited to the Carboxyl Half of  -Catenin Prostate cancer: K811N, K811Q, K1027Q, S1095S (Primary tumor) Prostate cancer: K816R, N1096R, S1140Q Between exon 21 and 22—EGFR (Primary tumor) No-sense or Missense Mutations: Deletion or insertion:

Lu et al., 1999 Zhang and Lu, unpublished Mutations Can Alter  -CateninLocalization in Cells Mutations Can Alter  -Catenin Localization in Cells 811, 816 Differential  -CateninLocalization May be Driven by Mutations in Prostatic Tumors Differential  -Catenin Localization May be Driven by Mutations in Prostatic Tumors

Some  -Catenin Mutations May Lead to Gain-of- Functions in Cell Proliferation/Survival, Adhesion, and Energy Metabolism

Compound Transgenic Mice Expressing Both Oncogenic Myc and δ-Catenin Mutation

Acknowledgements The Lu Lab Amy Friesland Amy Friesland Joyce Zhang Joyce Zhang Christi Boykin Christi Boykin Jian-Ping Lu Jian-Ping Lu Tao Wang Tao Wang Yan Zeng Yan Zeng Kwonseop Kim Kwonseop Kim Sarah James Sarah James Sonja Bareiss Sonja Bareiss Collaborators Yan-hua Chen Yan-hua Chen Lei Ding Lei Ding Huchen Zhou Huchen Zhou Robert Matusik Robert Matusik Scott Shappell Scott Shappell Frank Longo Frank Longo Baoan Chen Baoan Chen Xuemei Wang Xuemei Wang Contextual Contributions of Oncogenic and Tumor Suppressive Pathways