The how and why: cancer pathology
Originally from Malaysia, Professor Ui-Soon Khoo obtained her medical degree from University College Galway, Ireland. Thereafter, she joined the Department of Pathology at The University of Hong Kong (HKU) where she has been for 30 years. As a pathologist, she works with clinicians and basic scientists from different disciplines to find the most suitable diagnosis and treatment plan for patients.
“Pathology is interesting because it is the study of diseases; it explains to you the how and why. We play an important role in the management of patients because we are the ones who ultimately give you not only the tissue diagnosis, but also a lot of information that enables clinicians to decide on the best treatment,” says Khoo.
Khoo credits her first Head of Department, Professor Faith Ho for encouraging her to explore pathology research. “Professor Ho was very farsighted; she was the one who encouraged all the pathologists here to get into molecular research. Thanks to her sponsorship, I spent some time in Mount Sinai Hospital in Toronto and learned a great deal about screening for BRCA gene mutations,” said Khoo.
Using this knowledge, Khoo pioneered the study of BRCA mutations of breast and ovarian cancer among Hong Kong Chinese women. Her studies identified a founder mutation and three other recurrent BRCA mutations in Southern Chinese patients, as well as a high incidence of BRCA mutations in sporadic ovarian cancer. These findings on genetic susceptibility screening in Asian populations were novel at that time. For ovarian cancer it would serve as a future potential molecular target predicting treatment response.
The importance of collaboration
Although Khoo specializes in breast cancer research, she works a lot with other departments. “We collaborate a lot; nowadays we can never do anything alone. For a scientist to identify a novel gene, the starting material for analysis has to be correct. The forte of pathologists is our expertise in examining the morphology of tissue samples, ensuring analysis is performed accurately on the diseased parts in question. Clinicians need to provide the clinical follow up data. Findings also need further validation on more tissue samples. A lot of medicine requires the contribution of pathologists. ”
During the SARS epidemic in 2003, Khoo happened to attend a talk where the speaker, using HIV as an example, introduced DC-SIGN, a receptor on the surface of dendritic cells and a subset of macrophages that recognizes microorganisms. She was struck by how it acted as a Trojan horse, leading to the productive infection of interacting immune T cells and contributing to viral spread. DC-SIGN has a long tandem neck (with many repeats) with a carbohydrate recognition domain at the top. L-SIGN is a homologue of DC-SIGN and shares 77% identity in their coding sequence, including the long tandem neck repeats. Both were subsequently shown to bind to the SARS virus. In contrast to DC-SIGN, L-SIGN has considerable polymorphism in the tandem neck repeats. Khoo explained. “It occurred to me that the heterozygous variants in which the neck lengths differ could affect the binding capacity of the receptor to the virus and hence different susceptibility to infection. And I thought, why don’t we do a genetic association study for susceptibility to SARS?”
Collaborating with Dr Chen-Lung Steve Lin, a pediatric surgeon, who did the functional studies, Khoo collected blood samples and compared the genotypes of affected and unaffected individuals. Khoo’s research found that homozygosity in the tandem neck repeats (possession of two identical copies, one from each parent) for the L-SIGN receptor plays a protective role in SARS coronavirus infection. “The whole concept was novel. With functional studies to back up our in vivo data on quite a number of subjects genotyped, we were thus able to publish in Nature Genetics ” said Khoo. “It was my windfall! As a researcher, it was quite risky because it was completely out of my normal area of research. But SARS came along, there was a need to understand individual susceptibility to infection, I had this brainwave and went for it.”
Returning to cancer research, Khoo and her team are now seeking to understand Tamoxifen (TMX) resistance among breast cancer patients, focusing on estrogen receptor-positive (ER-positive) breast cancer, the most common type of breast cancer. Cancer cells bearing Estrogen receptors (ER) can be activated by the hormone estrogen, which plays an integral role in stimulating breast cancer growth. Due to low costs, TMX, a Selective Estrogen Receptor Modulator (SERM) is often used as a first line treatment. It successfully reduces breast cancer mortality by 25 to 30%. When TMX is introduced, estrogen receptors are blocked and target genes stimulating the growth of ER-positive breast cancer are inactivated. However, almost 50% of cases will acquire TMX resistance over time.
Khoo has made significant contributions in our ability to understand ER target genes and TMX resistance. Together with TMX, NCOR2, a nuclear receptor co-repressor gene inhibits the activation of ER target genes. Alternative splicing, a post-transcription mechanism that occurs during gene expression can give rise to different protein products from a single gene coding. Splice variants can have different, and possibly antagonistic biological functions. Khoo identified a novel alternative splice variant to the NCOR2 gene: BQ, associated with TMX resistance. Breast cancer cell lines that showed stronger BQ expression were TMX- resistant. This was also confirmed in tissue samples from patients at the time of first diagnosis who later developed TMX resistance.
With a grant awarded by the Hong Kong government’s Innovation and Technology fund, Khoo and her team developed a monoclonal antibody that specifically recognizes the BQ variant. “Prior to this there was no antibody to enable us to directly visualize the protein. We would have to run extracted protein on a Western Blot (a technique used to detect specific proteins in a tissue sample) and compare the different sizes. But you weren’t able to visualize it in a cell. ” The monoclonal antibody can be used for immunohistochemistry staining on Formalin-fixed paraffin embedded section (tissues that are preserved and stored indefinitely), that allows for visualization of the cell morphology together with assessment of localization of the BQ protein as recognized by the antibody. Researchers are thus able to compare and quantitate the distribution of BQ in TMX-resistant and TMX -sensitive cell lines, in cancer xenograft mouse models and in patient tissue samples. “We can use this tool to further our investigation for in vitro and in vivo studies to help us understand the role of BQ in tamoxifen resistance, cancer metastasis and disease relapse.”
In order to test the antibody on more cancer samples, Khoo collaborated with researchers from the United Kingdom. “The beauty in collaboration is that we don’t only have data from Hong Kong Chinese breast cancer patients, but also patients from the UK. Regardless of whether they are Caucasian or Chinese, the results are the same.” This has enabled us to establish BQ as a reliable biomarker to predict Tamoxifen resistance. “With this, we will be able to predict patients who will eventually develop resistance. Instead of waiting for them to develop resistance, we could consider giving them another drug at the outset,” said Khoo. Khoo has successfully filed for a patent.
Regarding future plans to investigate, Khoo explains, “What we don’t know is why BQ is overexpressed in the nucleus of some cancer cells. So far we have identified a nuclear import factor that interacts with a localization signal for BQ. When this receptor is knocked down, BQ is unable to enter the nucleus, enhancing Tamoxifen resistance.”
“We are also interested in finding out the mechanisms that induce splicing in the native tumor. We are looking to identify which mutations may result in BQ splicing,” said Khoo.
Professor Ui-Soon Khoo obtained her medical degree in 1984 from University College Galway, Ireland before joining The University of Hong Kong (HKU) where she completed her postgraduate pathology specialist training and a higher doctorate in research. An accomplished lecturer, Khoo was awarded the Faculty Teaching Medal by the Li Ka Shing Faculty of Medicine at HKU in 2007. She is currently a Clinical Professor at the Department of Pathology, Li Ka Shing Faculty of Medicine at HKU. Khoo was awarded a Croucher Senior Medical Research Fellowship in 2015.
To view Professor Ui-Soon Khoo’s Croucher profile, please click here