We have many great anti-tumor drugs that can do a fantastic job destroying the molecular insides of tumor cells. There is, however, a major catch: tumors have a nasty habit of become drug resistant. Such is the case with the breast cancer chemotherapeutic agent docetaxel. This drug can be effective at stopping breast cancer, but unfortunately many tumors are docetaxel-resistant. 50% of breast cancer patients receiving their initial course of chemotherapy are resistant to docetaxel, and it gets worse for patients who have already had chemotherapy - 70-80% of patients who have already received chemotherapy don't respond to this drug.
Administering docetaxel to resistant patients obviously wastes time that could be spent on other treatments. It also causes needless suffering of side effects. But is there some way to predict in advance who is going to be resistant? Or better yet, is there something we can do to eliminate docetaxel resistance altogether?
A Japanese group from the Japanese National Cancer Research Institute set out to tackle this problem, and their encouraging results have been reported in Nature Medicine. These researchers discovered a gene that makes breast cancer cells resistant to docetaxel, and they used that knowledge to knock out the source of docetaxel resistance. Although this study was largely confined to petri dishes and mice, cancer researchers can now use this result to identify patients who won't respond to docetaxel, and they are ready to test this new therapy target in real human cancers.
Finding the Culprit Gene
What makes cancer cells drug resistant? We have an amazing arsenal of chemicals that attack all sorts of molecular processes inside cancer cells, and yet many tumors manage to evade the onslaught. They do this using a fantastic toxic waste disposal system made of up membrane pumps known as P-glycoproteins. P-glycoproteins sit embedded in the outer membrane of our cells and send toxic molecules packing back out into intercellular space. Oncologists have tried, with limited success, to reduce chemotherapy resistance by knocking out these pumps with more drugs, but the cell is able to stay ahead of the game by making more and more P-glycoprotein.
The Japanese researchers came to this problem hoping to find new drug-resistance genes that could be treatment targets. They started off by comparing breast cancer tissue samples from drug-resistant and non-resistant tumors, and they observed that drug-resistant breast cancer tumors express high levels of a gene called RPN2. Thus, RPN2 may have something to do with making breast cancers docetaxel resistant, but what do we do with this knowledge?
To dig deeper, the scientists moved from human tumor samples to petri dishes. Cells cultured in a petri dish offer a virtually unlimited supply of material for experiments aimed at unraveling the molecular details of cancer. The Japanese researchers used some widely available breast cancer cell lines - petri-dish cells obtained by taking tumor samples and coaxing them to proliferate indefinitely in the lab. Once such cell lines are established, they can be shared around the world, and researchers can treat these cells with drugs, break them up and analyze their genes, or even inject them into mice and study their tumor-forming properties.
Now that the scientists had identified RPN2 as a possible source of drug resistance, they knocked out RPN2 in their petri dish cells and looked at what happened. They took a line of docetaxel-resistant breast cancer cells and treated it with a gene-knockdown agent called siRNA. An siRNA is a little snippet of nucleic acid that is able eliminate the RNA template necessary for making a particular protein. In this case, RPN2 siRNA eliminated any RPN2 template floating around, thus preventing any Rpn2 protein from being made. And as the researchers had hoped, once RPN2 was knocked down, the petri dish cells were now sensitive to docetaxel. Knocking out RPN2 somehow let docetaxel get around the toxic waste disposal system.
How Does It Work?
This result is exciting, but how does it work? The function of RPN2 offered the researchers a clue: RPN2 puts sugars on proteins. P-glycoprotein is one protein that is heavily decorated by sugars. These sugars play two major roles: they are involved in quality control, and they help make sure P-glycoprotein ends up in the right place in the cell, on the outer membrane. If the sugars on P-glycoprotein aren't right, then P-glycoprotein doesn't make it out to the membrane, and it also gets scrapped by the quality control process of the protein manufacturing system. When the scientists knocked out RPN2, they found that P-glycoprotein wasn't making it to the membrane.
And this isn't just true in a petri dish. The researchers also checked P-glycoprotein in their tumor tissue samples. Tumor cells that were docetaxel resistant, which had high levels of RPN2, also had a lot of P-glycoprotein in the membrane. Tumor cells that were sensitive to docetaxel had low levels of RPN2, and P-glycoprotein was not found in the membrane. This correlation is explained by the results found in the petri dish cell lines - knock out RPN2, and you prevent P-glycoprotein from doing its job as a membrane-embedded toxic waste pump.
Curing Mice with Breast Cancer
The Japanese group took this one step further. They injected their cancer cell lines into mice, where those cells went on to form tumors. In essence, they gave these mice breast cancer. Mice that were treated with RPN2 siRNA (knocking down RPN2 gene expression) and docetaxel did much better than mice that were treated with just docetaxel. Knocking out RPN2 in live mice produced the same effect observed in a petri dish.
What about Curing Humans?
The Japanese group has laid the groundwork for moving this treatment into humans. They have extensively tested their ideas in the lab, so what about the clinic? An immediate benefit is better prediction of docetaxel resistance - physicians can now know do a better job predicting which of their patients will benefit from docetaxel, and which should be spared the suffering caused by an ineffective drug.
But getting the siRNA treatment to work in humans is a more difficult problem (and the subject for another column). This kind of treatment is known as gene therapy. After a rocky start, gene therapy is making some slow strides forward. The advantage is that gene therapy can hit molecular targets that are tough to get at with drugs. Gene therapy is also more targeted towards specific genes, while drugs are more indiscriminate in their molecular attacks.
It is possible that targeting RPN2 won't work better than past drugs targeting P-glycoprotein directly. But two results of this Japanese study offer hope. First, it seems better to hit P-glycoprotein during the protein manufacturing process, rather than stopping it later on. The scientists, in addition to knocking down RPN2, tried hitting one particular P-glycoprotein with an siRNA treatment; that approach didn't work nearly as well as knocking down RPN2. And second, RPN2, like most genes, has multiple roles, one of which may include regulating cell suicide proteins. If you increase cell suicide in a tumor, your tumor shrinks. So by hitting RPN2, you may be hitting two birds with one stone: shutting down the toxic waste disposal system and promoting cell suicide.
Fighting cancer is a long slog with few magic bullets. But the bullets don't need to be magic, they just need to be more effective, and studies like this are going to steadily increase our anti-cancer arsenal. And just maybe, as gene therapy becomes more feasible, that arsenal may some day be stocked with a little magic.
Administering docetaxel to resistant patients obviously wastes time that could be spent on other treatments. It also causes needless suffering of side effects. But is there some way to predict in advance who is going to be resistant? Or better yet, is there something we can do to eliminate docetaxel resistance altogether?
A Japanese group from the Japanese National Cancer Research Institute set out to tackle this problem, and their encouraging results have been reported in Nature Medicine. These researchers discovered a gene that makes breast cancer cells resistant to docetaxel, and they used that knowledge to knock out the source of docetaxel resistance. Although this study was largely confined to petri dishes and mice, cancer researchers can now use this result to identify patients who won't respond to docetaxel, and they are ready to test this new therapy target in real human cancers.
Finding the Culprit Gene
What makes cancer cells drug resistant? We have an amazing arsenal of chemicals that attack all sorts of molecular processes inside cancer cells, and yet many tumors manage to evade the onslaught. They do this using a fantastic toxic waste disposal system made of up membrane pumps known as P-glycoproteins. P-glycoproteins sit embedded in the outer membrane of our cells and send toxic molecules packing back out into intercellular space. Oncologists have tried, with limited success, to reduce chemotherapy resistance by knocking out these pumps with more drugs, but the cell is able to stay ahead of the game by making more and more P-glycoprotein.
The Japanese researchers came to this problem hoping to find new drug-resistance genes that could be treatment targets. They started off by comparing breast cancer tissue samples from drug-resistant and non-resistant tumors, and they observed that drug-resistant breast cancer tumors express high levels of a gene called RPN2. Thus, RPN2 may have something to do with making breast cancers docetaxel resistant, but what do we do with this knowledge?
To dig deeper, the scientists moved from human tumor samples to petri dishes. Cells cultured in a petri dish offer a virtually unlimited supply of material for experiments aimed at unraveling the molecular details of cancer. The Japanese researchers used some widely available breast cancer cell lines - petri-dish cells obtained by taking tumor samples and coaxing them to proliferate indefinitely in the lab. Once such cell lines are established, they can be shared around the world, and researchers can treat these cells with drugs, break them up and analyze their genes, or even inject them into mice and study their tumor-forming properties.
Now that the scientists had identified RPN2 as a possible source of drug resistance, they knocked out RPN2 in their petri dish cells and looked at what happened. They took a line of docetaxel-resistant breast cancer cells and treated it with a gene-knockdown agent called siRNA. An siRNA is a little snippet of nucleic acid that is able eliminate the RNA template necessary for making a particular protein. In this case, RPN2 siRNA eliminated any RPN2 template floating around, thus preventing any Rpn2 protein from being made. And as the researchers had hoped, once RPN2 was knocked down, the petri dish cells were now sensitive to docetaxel. Knocking out RPN2 somehow let docetaxel get around the toxic waste disposal system.
How Does It Work?
This result is exciting, but how does it work? The function of RPN2 offered the researchers a clue: RPN2 puts sugars on proteins. P-glycoprotein is one protein that is heavily decorated by sugars. These sugars play two major roles: they are involved in quality control, and they help make sure P-glycoprotein ends up in the right place in the cell, on the outer membrane. If the sugars on P-glycoprotein aren't right, then P-glycoprotein doesn't make it out to the membrane, and it also gets scrapped by the quality control process of the protein manufacturing system. When the scientists knocked out RPN2, they found that P-glycoprotein wasn't making it to the membrane.
And this isn't just true in a petri dish. The researchers also checked P-glycoprotein in their tumor tissue samples. Tumor cells that were docetaxel resistant, which had high levels of RPN2, also had a lot of P-glycoprotein in the membrane. Tumor cells that were sensitive to docetaxel had low levels of RPN2, and P-glycoprotein was not found in the membrane. This correlation is explained by the results found in the petri dish cell lines - knock out RPN2, and you prevent P-glycoprotein from doing its job as a membrane-embedded toxic waste pump.
Curing Mice with Breast Cancer
The Japanese group took this one step further. They injected their cancer cell lines into mice, where those cells went on to form tumors. In essence, they gave these mice breast cancer. Mice that were treated with RPN2 siRNA (knocking down RPN2 gene expression) and docetaxel did much better than mice that were treated with just docetaxel. Knocking out RPN2 in live mice produced the same effect observed in a petri dish.
What about Curing Humans?
The Japanese group has laid the groundwork for moving this treatment into humans. They have extensively tested their ideas in the lab, so what about the clinic? An immediate benefit is better prediction of docetaxel resistance - physicians can now know do a better job predicting which of their patients will benefit from docetaxel, and which should be spared the suffering caused by an ineffective drug.
But getting the siRNA treatment to work in humans is a more difficult problem (and the subject for another column). This kind of treatment is known as gene therapy. After a rocky start, gene therapy is making some slow strides forward. The advantage is that gene therapy can hit molecular targets that are tough to get at with drugs. Gene therapy is also more targeted towards specific genes, while drugs are more indiscriminate in their molecular attacks.
It is possible that targeting RPN2 won't work better than past drugs targeting P-glycoprotein directly. But two results of this Japanese study offer hope. First, it seems better to hit P-glycoprotein during the protein manufacturing process, rather than stopping it later on. The scientists, in addition to knocking down RPN2, tried hitting one particular P-glycoprotein with an siRNA treatment; that approach didn't work nearly as well as knocking down RPN2. And second, RPN2, like most genes, has multiple roles, one of which may include regulating cell suicide proteins. If you increase cell suicide in a tumor, your tumor shrinks. So by hitting RPN2, you may be hitting two birds with one stone: shutting down the toxic waste disposal system and promoting cell suicide.
Fighting cancer is a long slog with few magic bullets. But the bullets don't need to be magic, they just need to be more effective, and studies like this are going to steadily increase our anti-cancer arsenal. And just maybe, as gene therapy becomes more feasible, that arsenal may some day be stocked with a little magic.
