G9a: A new marker to predict and treat aggressive ovarian cancer 

Associate Professor Jason Lee is using his expertise in epigenetics to find a new treatment solution for ovarian cancer patients who have become resistant to available treatments.  

Epigenetics focuses on factors that can control genetic activity without changing the DNA sequence of a gene. In the cancer space, this includes studying factors outside the gene that could be allowing cancer cell development or resistance to therapy. 

Previously, Associate Professor Lee’s team studied high-grade serous ovarian cancer samples, the most common subtype, and found that 50% of tumours had high levels of a protein called G9a.  

G9a protein is an enzyme that protects tumour cells from stress factors (like cancer treatments), by overcoming the genes that work to suppress cancer cells in two ways. Firstly, G9a helps cancer cells grow faster by reducing factors that slow down the cell cycle. Secondly, G9a helps stabilise conditions for tumour cells by recycling energy and nutrients so they can survive better and become more aggressive.  

On a molecular level, G9a protein makes small marks on chromatin (a combination of DNA and proteins that form chromosomes within the cell). These marks prompt a process called ‘methylation’, which is the addition of small ‘methyl’ groups to genes. The addition of these ‘methyl’ groups to certain genes can lead to the silencing of genes that are required to stop cancer from developing in healthy cells. 

To combat the effects of the G9a protein in ovarian cancer, the team have developed a new G9a inhibitor (G9ai). The inhibitor is a drug that blocks the action of the G9a protein, which allows several anti-cancer genes to reactivate and stop cancer cells from resisting treatment and surviving.  

With OCRF funding the team will assess whether their G9ai could effectively treat high-grade serous ovarian cancer by: 

• Validating that the G9a protein makes a suitable ovarian cancer treatment target.  
• Confirming whether levels of the G9a protein could help predict recurrence of ovarian cancer.  
• Determining how exactly their G9ai works against cancer. The team have shown in preliminary studies that the G9ai inhibits the growth of ovarian cancer cell lines, but here they seek to uncover exactly how it is working. 
• Determining how effective their G9ai is at causing cancer cell death in chemo-resistant ovarian cancer samples.  
• Testing, in preclinical models including cell lines and mouse models, how well G9ai works in combination with chemotherapy.  
• Testing, in preclinical models, how well G9ai can improve responses to immunotherapy.  

Data analysis to investigate the correlation between G9a and shorter cancer-free periods: 

The team will examine extensive patient datasets including those from The Cancer Genome Atlas and the International Cancer Genome Consortium. They will group patient samples depending on their level of G9a to see whether higher levels of G9a correlate with a lower relapse-free period. If so, it would indicate that high levels of G9a are associated with recurrence. This will help the team determine whether G9a can be used to predict recurrence earlier so that recurrent disease can be treated quickly. 

Confirming G9ai effectiveness in chemo-resistant samples: 

The team’s preliminary studies suggested the G9ai was effective, but this funding will allow Associate Professor Lee to validate it in 15 clinically relevant in vivo models, supplied by Professor Clare Scott from the Walter and Eliza Hall Institute of Medical Research.  

Combining G9ai with chemotherapy: 

Associate Professor Lee will trial whether using G9ai in combination with the chemotherapy drug carboplatin is effective at killing cancer cells without having to use a full dose of carboplatin. If this is the case, it could mean that G9ai can be used to both enhance the anti-cancer effects of chemotherapy and reduce its side effects. 

Combining G9ai with immunotherapy: 

The team’s preliminary studies indicated that epigenetic factors could exhaust T-cells, which the immune system relies on to effectively eliminate cancer. Here, the team want to see if their G9ai combination can stop the factors exhausting the T-cells and reactivate them, so that they can efficiently eliminate the tumour. They will do this by examining how sensitive ovarian cancer samples are to immunotherapy alone, compared to when immunotherapy is used with their G9ai.  

Because there is no early detection test for ovarian cancer, many cases are diagnosed in the later stages when it is more likely that the cancer will recur after initial treatment. When the cancer comes back it can be resistant to available treatments, such as chemotherapy. This means that projects like this that focus on new solutions for chemo-resistant patients are urgently required.  

OCRF funding allowed Associate Professor Lee’s team to gain further evidence that their G9ai drug combination could be highly effective in treating some chemo-resistant ovarian cancers. They demonstrated: 

• When they used the G9ai, the factors that had been shut down by cancer were reactivated, removing the advantage the cancer cells had and causing cancer cell death. 
• That very few healthy cells were affected when the G9ai was used, which means that as a treatment it has a very low toxicity and will likely have reduced side effects compared to chemotherapy. 
• That G9ai works by inducing the expression of a tumour suppressor and immune marker called Interleukin-24 (IL-24). This helped them understand why their G9ai is not very toxic for patients. Healthy cells already have IL-24 which means when the G9ai reactivates it in cancerous cells the healthy cells are not impacted. 
• Importantly, the team trialled another inhibitor that stops the EZH2 protein function. The inhibitor called Tazemetostat was found to be very toxic to healthy cells in ovarian cancer samples so the team could rule out further study with it. 

OCRF funding provided the preliminary data needed for Associate Professor Lee to secure government funding of $930,000 from the National Health and Medical Research Council. The project continues as the team tests their G9ai in combination with existing treatments.  

The team have shown in ovarian cancer cell lines that using G9ai means that the dosage of the chemotherapy drug carboplatin could be reduced by a third, significantly reducing side effects without reducing the rate or number of cancer cell death. 

Although in the past immunotherapies have not effectively helped ovarian cancer patients in most cases, the team demonstrated that their G9ai impacts immune cells and activates T-cells to elevate immune function. This indicates that it has the potential to enhance the effect of immunotherapies on ovarian cancer.  

The team are in the early stages of testing G9ai in combination with PARP inhibitors. These drugs have been shown to be effective in treating patients, particularly with the BRCA1 and BRCA2 gene mutations as cancer cells in these cases have increased susceptibility to G9a inhibition.  

Next phases: 

Not only were the team able to demonstrate that their G9ai works but the small molecule formulation is also relatively inexpensive which will likely make it more affordable for those who need it when it has undergone further testing. The team are now working to commercialise their G9ai treatment approach.