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Title: The role of FKBPL and its peptide derivatives in targeting stemness
Author: Annett, Stephanie Louise
ISNI:       0000 0004 6423 2302
Awarding Body: Queen's University Belfast
Current Institution: Queen's University Belfast
Date of Award: 2017
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FKBPL is a secreted protein with well-established anti-angiogenic activity and a novel therapeutic peptide, ALM201, derived from the protein has entered Phase l/ll clinical trial in ovarian cancer patients. Ovarian cancer is the most lethal gynaecologic cancer with a high incidence of recurrent chemo-resistant disease and this has been attributed to a subpopulation of cancer stem cells (CSCs), which escape standard therapies and drive metastatic spread. ALM201 binds to the cell surface receptor antigen, CD44, a classic marker of CSCs, and for the first time, we demonstrate ALM201's ability to target ovarian CSCs. Tumoursphere assays have demonstrated that ALM201 is effective at reducing ovarian CSCs in a range of cell lines and primary patient samples in vitro and reduced the CD44+CD117+ ovarian CSC subpopulation. Clonogenic assays suggest that ALM201 mediates ovarian CSC differentiation; a similar observation was previously noted in breast cancer. In vitro ALM201 displayed potent anti-CSC activity in the high grade serous (HGS) ovarian cancer cell line, OVCAR3, however, it displayed no anti-angiogenic or anti-CSC efficacy in in vivo models. In contrast, ALM201 treatment of Kuramochi xenografts resulted in significant growth delay and a 10 fold decrease in CSCs in in vivo experiments. Upon CD31/PAS staining, the Kuramochi xenografts displayed an extensive CD31+ vasculature network, which was disrupted by treatment with ALM201. On the other hand, the OVCAR3 xenografts had relatively few CD31+ blood vessels but had extensive PAS+ vasculogenic mimicry (VM) networks; which ALM201 did not target. Furthermore, OVCAR3 xenografts dramatically up regulated the inflammatory cytokines IL-6 and IL-8, in comparison to the Kuramochi xenografts, and as a result ALM201 had no effect on the CSC sub-population. In a tissue microarray of HGS ovarian cancer patients, high FKBPL expression correlated with an increase in progression free interval thus indicating a role for FKBPL as a prognostic biomarker in the clinic. Endocrine therapies are commonly used to prevent ER+ breast cancer release however, findings suggest that they may be increasing treatment resistant breast CSCs. FKBPLs preclinical peptide AD-01 has previously shown potent anti-CSC activity in the breast cancer setting and here we demonstrate its ability to abrogate endocrine enrichment of CSCs in both cell lines and patient samples. Furthermore, we demonstrate using in vivo limiting dilution models that ALM201 alone or in combination with tamoxifen was effective at delaying tumour recurrence by 12 days and 21 days, respectively. FKBPL and its peptide derivatives down regulate the DLL4/Notch pathway, a regulator of CSC self-renewal, thus indicating a novel mechanism of action. In the haemopoietic setting, ALM201 inhibited migration of chronic lymphocytic leukaemia (CLL) - like cell lines without inducing apoptosis. In an in vivo model of CLL, treatment of ALM201 resulted in a compartmental shift of the cells from the bone marrow to the peripheral blood and spleen. Furthermore, in vitro studies of acute lymphocytic leukaemia (AML) cell lines show that the combination of ALM201 and all trans retinoic acid (ATRA) significantly reduces cell viability, compared to ATRA alone. Analysis of AML cell morphology indicates that ALM201 may be inducing cellular differentiation and thus overcoming the characteristic maturation arrest of this disease. In summary, high FKBPL levels is associated with progression free survival in HGS ovarian cancer patients and the clinical FKBPL peptide, ALM201, effectively targets angiogenesis and CSCs in well-vascularised HGS ovarian cancer thus enhancing the efficacy of this agent in the clinic. In addition, we have demonstrated additional indications for ALM201 in treating endocrine resistance in breast cancer and a novel role in haematopoietic malignancies.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available