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Proprotein convertase 6: role in embryo implantation and clinical implications

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thesis
posted on 2017-02-28, 05:09 authored by Ho, Huiting
To enable embryo implantation for the establishment of a pregnancy, the uterus must acquire endometrial epithelial receptivity and undergo stromal differentiation known as decidualisation. Proprotein convertase 6 (PC6) is a member of the PC family of proteases that are critical in controlling the post-translational activation of a multitude of precursor proteins through limited proteolysis. Our group has established that uterine PC6 is critical for embryo implantation, as it is essential for endometrial epithelial receptivity (in women) and decidualisation (in both women and mice). In addition, PCs including PC6 are also involved in HIV infectivity, and inhibition of PC activity inhibits HIV infections in cells. We hypothesised that targeting PC6 in the female reproductive tract (vagina, cervix and uterus) could provide a novel approach to the development of non-hormonal and women-controlled contraceptives that could also protect women from HIV infections. The vagina is the entry site of sexually transmitted HIV in women, and transmission can be stopped by vaginal application of anti-HIV drugs. Therefore, vaginal delivery of PC6 inhibitors presents the ideal route of administration to achieve this dual protection. A key requirement to prove this concept is to develop potent and vaginally deliverable PC6 inhibitors, and to test their efficacy in appropriate animal models. This thesis examined the contraceptive potential of two types of PC6 inhibitors, (i) peptide-based and (ii) small molecule compounds. Work described in Chapter 2 examined Poly R (one of the most potent peptidic PC inhibitors) and its PEGylated derivatives. The study revealed that C-terminally PEGylated Poly Rs had comparable potency to the parental Poly R, but with enhanced vaginal absorption and penetration across the vaginal epithelium. However, only one of the C-PEGylated Poly R derivatives, C-30k-PEG Poly R could reach the uterus at a relatively low level following vaginal administration in mice. This was contradictory to expectation, because it has been widely demonstrated in women that vaginally administered drugs preferentially localise to the uterus, a phenomenon called the “first uterine pass effect”. In the absence of a better animal model, the study continued to test C-30k-PEG Poly R in mice for inhibition of embryo implantation following vaginal administration (Chapter 3). Overall, inhibition of implantation was achieved in 71% of mice (24% complete and 47% partial inhibitions). These encouraging results strongly suggest that inhibition could be much higher if efficacy of inhibitor delivery could be improved in a more appropriate animal model. To assist my research and to fill a knowledge gap in the field, a simple, reliable and high-throughput in vitro model for human embryo attachment was established (Chapter 4). This model was proven to be effective and reproducible for the study of the attachment of human trophoblast spheroids to endometrial epithelial cells. This publication has attracted attention from other researchers, highlighting its significant contribution to the field. Chapter 5 described studies on a class of five small molecule compounds that were previously reported as potent inhibitors of furin (another PC member). The work revealed that all five compounds potently inhibited PC6 activity in vitro. However, only one (compound 1o) was able to inhibit PC6-dependent cellular processes (decidualisation and epithelial receptivity) in human cell-based models. Compound 1o is unique among the five compounds as it is the most lipophilic. These findings identified compound 1o as a potent small molecule PC6 inhibitor with pharmaceutical potential to inhibit embryo implantation. Future in vivo studies need to evaluate whether compound 1o is superior to C-PEGylated Poly Rs as a vaginally deliverable PC6 inhibitor for the dual protection. In summary, this thesis established an important foundation for future studies to prove the hypothesis that inhibiting PC6 is a viable strategy for the development of non-hormonal and women-controlled contraceptives.

History

Principal supervisor

Guiying Nie

Year of Award

2013

Department, School or Centre

Biochemistry and Cell Biology

Campus location

Australia

Course

Doctor of Philosophy

Degree Type

DOCTORATE

Faculty

Faculty of Medicine Nursing and Health Sciences

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