Some long non-coding RNAs may also affect later developmental stages. We met the HOX genes in Chapter 4. These are the genes that are important for correct patterning of body parts. They’re the ones where mutations in fruit flies can lead to bizarre effects such as legs on the head. HOX genes are found in clusters in the genome, and these regions are extraordinarily rich in long non-coding RNAs. This is in contrast to their lack of ancient viral repeats. Scientists were keen to investigate if the long non-coding RNAs influenced the activity of the HOX genes in the same place in the genome. To test this, researchers used a technique to decrease the expression of a specific long non-coding RNA from the HOX gene region in chick embryos. When they did this, limb development went wrong. The bones towards the ends of the limbs were abnormally short.{146} Similarly, knocking out expression of another long non-coding RNA from this genome region in mice resulted in animals with malformations of the bones of the spine and wrists.{147} Both sets of data are consistent with the long non-coding RNAs being important regulators of HOX gene expression, and consequently of limb development.
Cancer can in some ways be thought of as the flip side of development. One of the problems in cancer is that mature cells may change and revert to having some of the characteristics of less specialised cells, with a higher capacity to divide uncontrollably. Given that long non-coding RNAs are important in pluripotency and in development, it’s perhaps not surprising that some have now been implicated in cancer.
One large study analysed the expression of long non-coding RNAs in over 1,300 individual tumours from four different cancer types (prostate, ovarian, a type of brain tumour called glioblastoma and a specific form of lung cancer). There were about 100 long non-coding RNAs where high levels of expression were most commonly found in patients who died quickly from the disease. Nine of these long non-coding RNAs showed this association no matter the class of cancer that was assessed, which suggests they may be useful as more general markers for predicting survival chances in a patient.{148}
For three of the cancer types (prostate cancer was the exception), the same study reported that they could detect long non-coding RNAs that differentiated one sub-class of tumour from another. Although we refer to ovarian cancer, for example, there are different types of ovarian cancer depending on the cell types involved, and this affects the natural history of the tumour in a patient. This in turn can have implications for the disease prognosis and the treatment that a patient should receive. Analysing the expression of specific long non-coding RNAs in a tumour sample may help clinicians in the future to select the most appropriate therapies for an individual patient.
The number of studies that report associations between long non-coding RNA expression and cancer are growing all the time. Intriguing data are also emerging from genetic studies of cancers. Some cancers are caused by a single really strong mutation which is passed on within a family. Probably the best-known example is the mutated BRCA1 gene which puts women at very high risk of aggressive breast cancer. It was knowing that she had a mutation in this gene that led the actress Angelina Jolie to elect for a double mastectomy in 2013. Such very strong single gene mutations are pretty rare in cancer. But studies have shown that quite a number of cancers do have a genetic component. The problem has been that when scientists mapped where the genetic variations were that were associated with cancer risk, they were frequently in regions of the genome where there were no protein-coding genes. Of just over 300 genetic variations linked to cancer, only 3.3 per cent changed amino acids in a protein, and over 40 per cent were located in regions between classical protein-coding genes. In these situations the variations may be affecting not protein-coding genes but long non-coding RNAs. Recent studies have confirmed this is the case for some of these variations in at least two cancer types (papillary thyroid cancer and prostate cancer).{149}
Encouragingly, we are also beginning to gather functional data that shows in some cases that these relationships are more than just associations, that the long non-coding RNAs are themselves causing alterations in the behaviour of the cancer cells.
There is a long non-coding RNA whose expression is increased in prostate cancer. This over-expression causes decreased expression of key proteins that normally hold cells back from proliferating too fast.{150},{151} Over-expression of this long non-coding RNA is therefore essentially like releasing the handbrake on a car parked facing down a hill. The long non-coding RNA that causes skeletal deformations when it is knocked out in developing mice is over-expressed in a variety of cancers including liver,{152} colorectal,{153} pancreatic{154} and breast{155} and its over-expression is associated with poor prognosis for the patients. Studies using cancer cells in culture in the lab suggest that the over-expression of this long non-coding RNA may make the cells more likely to migrate and invade other parts of the body.
Some of the strongest data confirming that long non-coding RNAs are actively involved in cancer, rather than just carried along for the ride, come from prostate cancer. When prostate cancer begins to develop, its growth depends on the male hormone, testosterone. Testosterone binds to a receptor and this leads to activation of various genes that promote cell proliferation. Testosterone binding to its receptor is like you putting your foot down on the accelerator pedal of your car. Prostate cancer is initially treated using drugs that stop the hormone binding to its receptor. This is like having something between your foot and the accelerator pedal, so that you can’t press down on it to make the car go faster.
But over time, the cancer cell frequently finds a way around this. The hormone receptor finds ways of activating genes irrespective of whether there is testosterone around or not. It’s as if someone has put a bag of sugar on top of the accelerator. The pedal is always pressed down and speeding up the car, even if you have your feet on the dashboard. Two long non-coding RNAs that are highly over-expressed in aggressive prostate cancer have been shown to play a critical role in this process. They assist the receptor, driving gene expression even when there is no hormone around, and accelerating cell proliferation. They play the role of the bag of sugar in the car simile. If expression of these specific long non-coding RNAs is knocked down in cancer models, the tumours show a really dramatic decrease in growth, supporting the critical role of these molecules.{156}
Another long non-coding RNA has also been implicated in prostate cancer. The higher the levels of this long non-coding RNA, the more aggressive the cancer, the shorter the recurrence time after treatment and the greater the risk of death. Knockdown of this long non-coding RNA has a similar protective effect in cancer models to that described above, but in this case the effects do not seem to be due to interactions with the testosterone receptor.{157} This indicates that long non-coding RNAs may affect cancer progression in different ways, even in one tumour type.