A Closer Look
Partners in Crime: Using Synthetic Lethality to Identify New Cancer Targets
One of the big stories of the past year in cancer research has been the emergence of several agents that target an enzyme known as PARP1. Early-stage clinical trials involving women who have particularly difficult-to-treat types of breast cancer, including tumors with a mutated form of either the BRCA1 or BRCA2 gene, have shown that treatment with drugs that block PARP1 produced promising results. (See the “Also in the News” sidebar below.)
A cancer cell with mutations in the BRCA gene has a limited capacity for repairing damaged DNA, but can still remain viable. If another route of DNA repair is also blocked, such as the type performed by the PARP1 enzyme, the cancer cell loses an important fail-safe and is more likely to die.
Similar to PARP1, the two BRCA genes produce proteins that help repair damaged DNA. When these genes are mutated, they can transform a healthy cell into a cancerous one. However, combining a BRCA mutation with a PARP1 enzyme that is under therapeutic assault eliminates one potential fail-safe mechanism that the genetically unstable but still-thriving cancer cell needs to survive. The mutated genes and PARP1 enzyme have what is called a “synthetic lethal” relationship: when either one alone is blocked or damaged, the cancer cell is all right; when both are blocked or damaged, the cancer cell dies.
A number of oncology research groups are using relatively new tools such as heavy-duty bioinformatics and small-molecule or RNA interference (RNAi) screens to identify synthetic lethal relationships between well-established therapeutic targets and, in some cases, lesser-known components of cancer cells’ signaling networks. They believe these discoveries could lead to new treatment options that overcome age-old problems in oncology, such as excessive toxicity, treatment resistance, and “undruggable” targets.
Prior to the clinical trial results with the PARP1 inhibitors, synthetic lethality was “a promising theoretical concept that people thought should work [in humans] because it works in model organisms,” said Dr. William Hahn of the Dana-Farber Cancer Institute. Now, he continued, it’s been “taken all the way to the patient” and shown “to have a clinical benefit, and that’s why people are excited about it.”
Finding a Hit in a Signaling Network
The concept of synthetic lethality is very attractive, said Dr. Igor Astsaturov, a researcher at Fox Chase Cancer Center who is involved in several synthetic lethality-related projects. Most normal cells are dormant for long periods, he explained, and they use growth-related signaling pathways only intermittently. Genetically unstable cancer cells, on the other hand, “are continuously exploiting these signaling systems to their advantage, so they are vulnerable to inhibition.”
Also in the News: PARP Inhibitor Shows Promise in Difficult-to-treat Breast Cancer
An investigational drug that targets PARP-mediated DNA repair improved survival in women with metastatic, triple-negative breast cancer, U.S. researchers reported last week at the European Society for Medical Oncology Congress in Milan. Updated results from a 121-patient phase II clinical trial showed that a combination of chemotherapy and the PARP inhibitor iniparib improved overall survival by 5 months (12.3 months versus 7.7 months) in women with this difficult-to-treat type of breast cancer compared with women who only received chemotherapy.
Women in the trial who received iniparib also had superior response rates and longer progression-free survival compared with women who only received chemotherapy. The addition of iniparib to chemotherapy did not appear to produce any additional side effects, the researchers reported.
“These data are promising and suggest that iniparib may provide a potential new treatment option for patients with metastatic triple-negative breast cancer, which currently has limited therapeutic options,” said the trial’s lead investigator, Dr. Joyce O’Shaughnessy, who directs the Breast Cancer Research Program at the Baylor Charles A. Sammons Cancer Center in Texas, in a news release.
Based in large part on findings reported by Dr. Astsaturov and his colleagues in Science Signaling last month, several human trials testing treatments based on synthetic lethality are being planned at Fox Chase. Their study was centered on the epidermal growth factor receptor (EGFR), a protein that is overexpressed in a number of cancers and is the target of several FDA-approved cancer drugs.
To conduct the study, explained co-investigator Dr. Erica Golemis, the team exhaustively mined gene and protein databases to construct a large but still limited “library” of 638 proteins that make up a broad intracellular signaling network in which EGFR is a critical component. Using an RNAi-based screen, they identified synthetic lethal relationships between the products of several genes and EGFR: when EGFR was inhibited by a drug and expression of each of the other genes was simultaneously “knocked down” by small interfering RNAs, there was an increase in cancer cell death. (See the box at the end of this article.)
“The study generated a lot of solid leads,” Dr. Astsaturov said. Most of the gene products that had a robust synthetic lethal relationship with EGFR tended to cluster near to EGFR or had direct or indirect interactions with it. Several of the strongest hits “pointed directly to logical targets for clinical exploration of combination therapies,” he continued, including Aurora kinase A and STAT3, both of which are targets of available investigational drugs. The Fox Chase trials will combine an EGFR inhibitor with an Aurora kinase A inhibitor in patients with metastatic lung cancer and head and neck cancer.
Getting Around the Shortcomings of Current Treatments
One of the theoretical advantages of cancer treatments based on synthetic lethality—now with some clinical evidence to support it—is that they should have minimal toxicity, because only cells with the “impairments” that comprise the synthetic lethal relationship (e.g., a mutated gene and a therapeutically inhibited enzyme) should be affected. Those cells should almost exclusively be cancer cells. In trials that tested PARP1 inhibitors, the drugs appeared to add little in the way of side effects.
Treatments based on synthetic lethality, stressed Dr. Hahn, may also help to overcome the problem of targets that, either due to underlying biology or the targets’ actual physical makeup, are “undruggable” with small molecule and biologic drugs; a number that, according to one recent estimate, could represent as much as 75 percent of the identified molecular targets for cancer.
A prime example of a potentially valuable undruggable target is the oncogene KRAS, which is commonly mutated in a number of cancers, including pancreatic, lung, and colon. But synthetic lethality may offer a route around the problem.
Last year, Dr. Hahn and colleagues published studies in Cell and Nature in which they used RNAi screens in cancer cell lines with mutant KRAS to identify a synthetic lethal relationship between KRAS and two kinase enzymes, STK33 and TBK1. Kinases have proven to be highly druggable targets. In the same issue of Cell, Dr. Steven Elledge from Harvard Medical School and colleagues identified another kinase called PLK1 that had a strong synthetic lethal relationship with mutant KRAS.
Although many synthetic lethality studies rely on small-molecule and RNAi screens, the synthetic lethal relationship between PARP1 and BRCA mutations was actually deduced from previous research on DNA repair and then validated in cell lines and animal models before moving into human trials. (Researchers are already searching for similar relationships between PARP1 and other DNA-repair targets.) While it is possible that other synthetic lethal relationships can and will be identified in this way, it will be difficult to do, said Dr. William Kaelin from the Dana-Farber Cancer Institute.
Although the understanding of the biology that underlies cells’ molecular networks has improved substantially, it is “far from complete,” Dr. Kaelin said. “The lesson from the screens in model organisms, such as yeast, is that it’s difficult to predict synthetic lethal interactions.”
And although Dr. Kaelin believes that identifying and targeting individual “drivers” of disease is and will continue to be a fruitful therapeutic avenue, synthetic lethality research underscores the importance of the context in which those targets reside. “At the end of the day, we are dealing with molecular networks rather than simple linear pathways,” he said.
Improved understanding of how those networks operate may yield yet another benefit: preventing the emergence of resistance to therapy, something that can be achieved “by using combinations of agents that are not cross resistant and have unique mechanisms of action,” Dr. Kaelin said.
The parallel that is often raised is the “cocktail” therapy approach used to treat HIV. The idea is to “hold things down so that the tumor can’t select for resistance,” Dr. Golemis explained. “Whether you do that by sequential treatments or by developing good combination treatments that hit multiple hubs in the signaling network, we’re gradually accumulating the tools to be able to do this.”
Anatomy of an RNAi Screen for Synthetic Lethality
From the initial library of 638 proteins chosen for the screen in the Fox Chase study, the team identified more than 60 proteins that cancer cells under assault with an EGFR inhibitor rely on to survive. The screens use RNA interference in several cancer cell lines to, one at a time, knock down the expression of the genes that produce these proteins. The cell lines are simultaneously treated with an EGFR inhibitor or a control drug that does not affect cancer cell viability. The team then looked at the effect of the knockdown of each individual gene on the extent of cell death in both EGFR-inhibitor treated and non-treated cells. From the initial group of “hits”—genes that, when silenced, yielded increases in cancer cell death in the inhibitor-treated cells—the team did additional validation testing with other small interfering RNAs to confirm the findings and isolate the most robust synthetic lethal relationships.
The entire study took more than 3 years to conduct, Dr. Golemis explained. Although it is a cumbersome process, she added, “RNAi screening is becoming a dominant way of approaching biological networks.”
One of the first groups to use this approach was the laboratory of Dr. Lou Staudt in NCI’s Center for Cancer Research. As described in a paper published in 2006 in Nature, they used the approach—what they called an “Achilles’ heel genetic screen”—in cell lines of diffuse large B-cell lymphoma to identify genes aside from known oncogenes and tumor suppressors that cancer cells need to survive.