Functional characterisation of OPCML clinical mutations in ovarian cancer and correlate to OPCML crystal structure

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Title: Functional characterisation of OPCML clinical mutations in ovarian cancer and correlate to OPCML crystal structure
Authors: Alomary, Mohammad Nasser
Item Type: Thesis or dissertation
Abstract: Ovarian cancer is a heterogeneous disease, considered the most lethal of all gynaecological malignancies. With an initially high response rate to treatment, ovarian cancer can suddenly relapse, with advanced stages showing loco-regional dissemination. Despite enormous efforts to tackle ovarian cancer, current treatments fail to bring complete remission or significantly extend patient life. OPCML is a promising candidate in ovarian cancer treatment, identified as a tumour suppressor frequently inactivated in ovarian and other cancers. OPCML is a negative regulator of RTKs and regulates different cellular activities. The aims of this PhD were to identify and functionally characterise clinical OPCML mutations in ovarian cancer guided by the newly resolved crystal structure of OPCML. The project describes how TCGA and COSMIC databases have been used here to identify somatic missense mutations in several tumours. A panel of OPCML mutant constructs was then created and these selected constructs were utilised to transduce three ovarian cancer cell lines (SKOV3, PEA1, and PEO1) for domain I, and SKOV3 cells for domain II and III. Selected transduced mutants were used to characterise the mutants’ localisation, interaction with receptor tyrosine kinases, proliferation, cellular transformation, invasion, migration, and their potential role in the adhesion to the extracellular matrix in vitro. Furthermore, two in vivo models were used to characterise OPCML mutants: the athymic mice model and the chick embryo chorioallantoic membrane assay. The results showed that the OPCML mutants in domain I behave differently when they interact with receptor tyrosine kinase AXL and FGFR1 compared to wild-type OPCML. These OPCML mutants preserve the ability to inhibit growth in vitro, but have a complete loss of function in vivo. They also block the OPCML inhibitory activity on anchorage-independent growth, invasion, migration, and adhesion to extracellular matrix components laminin I, fibronectin, and fibrinogen. The OPCML mutants in domain II showed partial loss of function in adhesion to collagen I and IV, and complete loss of tumour suppressor function in cell proliferation in vitro and in vivo, anchorage-independent growth, invasion, migration, and adhesion to laminin I, fibronectin, and fibrinogen. The OPCML mutants in domain III preserve adhesion to collagen I and IV, but show complete loss of function in the inhibition of tumour growth in vitro and in vivo, anchorage-independent growth, invasion, migration, and adhesion to laminin I, fibronectin, and fibrinogen. Therefore, this study identifies a new mechanism of OPCML inactivation by mutations and shows that single amino acid change can abolish OPCML function. These findings suggest that OPCML is an excellent candidate for a new and novel therapeutic strategy in ovarian cancer and other malignancies, as well as other non-cancer diseases.
Content Version: Open Access
Issue Date: Dec-2017
Date Awarded: Apr-2018
URI: http://hdl.handle.net/10044/1/68395
Supervisor: Gabra, Hani
Recchi, Chiara
Sponsor/Funder: King Abdulaziz City for Science and Technology (KACST)
Ovarian Cancer Action
Department: Department of Surgery & Cancer
Publisher: Imperial College London
Qualification Level: Doctoral
Qualification Name: Doctor of Philosophy (PhD)
Appears in Collections:Department of Surgery and Cancer PhD Theses



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