Anti-cancer Research – Dr Virna Leaner
Title of the Project
Identification of nuclear import inhibitors with anti-cancer activity
Aim of the Project
This project is based on the hypothesis that the nuclear import protein, Kpnβ1 is critical for the growth and survival of cancer cells and that a rational drug design approach to identify small molecule inhibitors against Kpnβ1 may yield effective anti-cancer treatments.
Previous work performed in our laboratory used a genomic approach to identify target genes that associate with and are necessary for the growth of cervical cancer cells. We identified the nuclear import protein, Kpnβ1 as a gene that is essential for the growth of cancer cells as inhibition of its expression resulted in cell death via apoptosis. Similar findings have been observed in oesophageal and breast cancer cells where inhibition of Kpnβ1 results in a significant inhibition of the growth of these cancer cells (unpublished data). As no therapeutic agents against Kpnβ1 have been developed to date, in this study we aim to use a rational drug design approach to identify small molecule inhibitors against nuclear import.
The objectives of this project are:
- To use computational chemistry to identify potential small molecule inhibitors against Kpnβ1.
- To develop an in vivo assay to test the efficacy of small molecules that inhibits nuclear import via Kpnβ1.
- To determine if inhibition of nuclear import with small molecules inhibits the biology of cancer cells.
What has been achieved to date?
Click here to read what has been achieved to date.
What is hoped to be achieved in the next year?
To achieve our objectives the project has been divided into two phases. Phase 1 includes objectives 1 and 2, while phase 3 includes objective 3.
PHASE 1: Time frame: 1 year (April 2010 – March 2011).
Objective 1. Use computational chemistry to identify small molecule inhibitors against Kpnβ1
- Preliminary screening to identify potential target regions: complete
- Screening of an in silico library of 10, 000, 000 compounds for small molecules that may bind Kpnβ1: IN PROGRESS.
It is anticipated that the running of individual virtual screens will take approximately 3-4 weeks. Thus screening two separate regions is expected to take 6-8 weeks.
Objective 2. To develop an in vivo assay to test the efficacy of small molecules that inhibits nuclear import via Kpnβ1
- Cloning of the Kpnβ1 target protein, SREBP-2 into the expression vector, pEGFP: complete
- Transient transfection of GFP-SREBP-2 into cancer cell lines to observe cellular localisation of the target protein in the presence of uninhibited Kpnβ1: complete
- Inhibiting Kpnβ1 expression using siRNA and observing the effect on GFP-SREBP-2 localization: IN PROGRESS
- Extending the in vivo screening assay to cancers of different origins (oesophageal, cervix and breast) by preparation of cancer cells lines with stable GFP-SREBP-2 expression: IN PROGRESS.
PHASE 2: Time frame 2 years (April 2011 – March 2013).
Objective 3. To determine if inhibition of nuclear import with small molecules inhibit the biology of cancer cells
- Inhibiting Kpnβ1 expression in cancer cells with stable GFP-SREBP-2 expression using siRNA.
- Testing compounds (~50 per target site ie. 100 compounds for 2 sites) for inhibitory activity against Kpnβ1 function.
- Use compounds that are positive in inhibiting nuclear import to determine their efficacy in blocking the biology of cancer cells. The effect of inhibiting Kpnβ1 on the following biological endpoints will be determined;
- cell growth
- cell morphology
- colony formation
- mechanisms of inducing cell death (apoptosis, autophagy, necrosis).