Functional analysis and therapeutic targeting of AKT isoforms in BRAF mutant melanoma
McRee, Siobhan.
2018
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Abstract: Despite recent advances, metastatic melanoma remains the deadliest form of skin cancer, and new therapeutic strategies are urgently needed. The PI3K/AKT pathway is known to promote tumor progression and metastatic dissemination in many cancer types, including melanoma. PI3K/AKT signaling is hyperactivated in many melanomas, often through loss of the negative regulator PTEN, which occurs ... read morein 12-20% of cases. PTEN loss cooperates with oncogenic BRAF to induce metastatic melanoma and leads to unrestrained activity of the PI3K effector kinase, AKT. The AKT family of serine/threonine kinases comprises three highly homologous isoforms (AKT1, AKT2, and AKT3) that are major effectors of the PI3K pathway, but despite their unique roles in other cancers, isoform-specific effects in melanoma have yet to be systematically interrogated. We performed shRNA mediated conditional knockdown and CRISPR/Cas9 gene editing of AKTs in a wide variety of human melanoma cell lines, as well as genetic ablation of each isoform in a BRAF-driven mouse model of melanoma. We reveal a role for AKT2 in promoting melanoma cell migration and invasion in vitro and find that AKT2 is required for metastatic seeding in vivo. Additionally, AKT2 specific depletion delays tumor growth and improves survival of mice with metastatic disease after tumor cell seeding, suggesting that AKT2 supports growth or survival in the metastatic niche. We propose several mechanisms whereby AKT2 may mediate these phenotypes, including via regulation of key epithelial-mesenchymal genes, promoting aerobic glycolysis, and responding to hypoxia. In contrast, we observe that the AKT1 isoform specifically promotes melanoma cell proliferation, and genetic ablation of AKT1 in melanoma prone mice prolongs overall survival. We also reveal that the AKT3 isoform may be important in UV-initiated melanomagenesis. Lastly, while non-specific pan-AKT inhibitors are used clinically to moderate benefit, their use is hampered by myriad off-target effects. To increase the efficacy of AKT targeting for clinical benefit, we endeavored to identify AKT isoform-specific substrate effectors that may be mediating differential phenotypes, and therefore potential targets for therapy. We also report efforts to use a tumor specific antigen to target a clinically relevant pan-AKT inhibitor to melanoma tumors in the mouse, and additionally test small molecule inhibitors that synergize with existing targeted therapies. In summary, our work moves toward establishing the contribution of AKT isoforms to melanoma, to improve therapeutic strategies and outcome for this devastating disease.
Thesis (Ph.D.)--Tufts University, 2018.
Submitted to the Dept. of Genetics.
Advisors: Phil Hinds, and Karl Munger.
Committee: Philip Tsichlis, Charlotte Kuperwasser, and Alex Toker.
Keywords: Genetics, Oncology, and Cellular biology.read less - ID:
- cc08hs98z
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