Identifying the trespasser: bladder cancer cells identified with AND gate genetic circuits

In 2015 it was estimated that about 1,620 people died of cancer every day. That meant that in that year alone, almost 590,000 were killed by cancer.

It is not easy to say that you are untouched by cancer in any way, in fact, it is almost impossible. There are constant ads flickering across screens of TVs, and billboard ads that ask for money to fund cancer research in both private labs and huge hospitals. In the case of the not so lucky, this “cancer touch” goes as far as reaching a loved one, friend, and in some cases, themselves.

Yet, it has been almost 250 years since the first cancer cause was discovered, and around 3,600 years since the first signs of the disease were identified (although it was not called cancer at the time). What’s more, although there has been significant progress in the diagnosis and treatment of the very often, fatal disease, these treatments often kill our healthy cells as well.

Is there no hope for creating a treatment for cancer without the unwanted side effects? … There is a chance!

Now more than ever treatments for cancer are being looked into from several different fields of emerging technology! That means that the chances of finding a cure, or at least a more efficient way of treating cancer has exponentially grown since humans first started combatting the disease. One of these technologies is Synthetic Biology, a field of science that mashes together basic engineering principles with the central dogma of biology — DNA is transcribed into RNA, and RNA is then translated into Proteins which carry out functions in the body.

The central dogma of biology

What does that have to do with cancer?

To put it simply, mutations in DNA or RNA during transcription or translation lead to a protein malfunction and expression. Not only does this cause a malfunction in the protein expressed by the particular gene, but the mutation can also cause the inhibition of apoptosis (programmed cell death) and the total shutting off of a cell’s ability to control its divisions. This means that cancer cells will divide uncontrollably, and they won’t die off.

If changing the protein expression of a cancer cell can inhibit its ability to divide and metastasize, then wouldn’t the first step to changing protein expression be to go back to the roots of the problem, the DNA?

To stop cancer at its roots, before the cancer cells become something fatal, they must be stopped from dividing and they have to be killed. By creating biological molecules that do this work for us, cancer treatments can become more specialized, more concentrated, and less deadly to the good cells in our body. That means that by implementing genetic synthesis of molecules there is a chance to get infinitely better treatments, and maybe even a potential cure for cancer.

What is the synthesis of Genetic Materials?

Synthesis of genetic materials is just a fancy way of saying making genetic molecules from scratch. That means that we as the human race are at a point in technology where we can not only cut and paste the fundamental building block of life (DNA), but we can make it from scratch!

Sure, we can cut and paste DNA and RNA, which most people refer to as genetic engineering, but synthetic biology is much more related to the engineering part of the term. That basically means that although we call genetic engineering, engineering, it is more like genetic editing, while synthetic biology combines a more accurate version of engineering with biological molecules that are only limited to the human imagination.

Not only can we synthesize new genetic molecules, but we can synthesize molecules so that they attach and bind to specific molecules already found in nature. Meaning that not only can we make molecules, but we can make molecules that track them or bind to them to create an output signal. That signal can manifest in different ways.

In the case of binding receptors, they can either create an output that inhibits expression of a gene or promotes the expression of a gene.

What does that mean for cancer?

It has been known that cancer cells differ from the usual healthy body cell, and that has further been proved by the different molecules that can be found on cancer cells. By creating a circuit that is able to bind to these molecules and then create an output for inhibiting cell division, scientists can create a way to stop the rapid growth of cancer cells!

Using this system of creating receptors for inhibiting molecules, Boolean Logic gates that control the expression of a gene were synthesized and tested. These weren’t regular logic gates, but instead, gates that were devised of genetic material encoded with a specific task to undertake.

But… what is a logic gate?

Traditionally, logic gates are usually found in electronics. They are devices that utilize the Boolean functions (OR, AND, or NOT) to produce an output. Simply meaning that combined with one of the functions, the devise used a logical operation to produce an output.

The functions themselves are easy to understand. The AND gate requires two inputs, both strong, for an output to be produced. The OR gate requires either molecule (when working with a binary system)but not both, to produce and output, and the NOT gate is responsive when there aren’t any inputs being put into the system.

When applied to genetics, logic gates can be made out of genetic materials and “programmed” to work the same way as logic gates in electronics do.

That was it. The key to identifying and stopping cancer cells was in genetic logic gates and circuits that bind to a compound to inhibit action! Using their knowledge of cancer cells and logic gates, a group of researchers from the Key Laboratory of Medical Reprogramming Technology devised a way to put together the characterization of cancer cells with logic gates and CRISPR to devise a way to reprogram a cancer cell into apoptosis (seen here).

The Logic gate, CRISPR Experiment

The scientists observed that cells usually emit small nanosized particles of DNA in very small increments. While taking a look at this, they also observed that cancer cells would release their DNA more rapidly and more frequently than their healthy cell counterparts. Using this knowledge of how cancer cells behaved, the scientists set out for a way to inhibit the cells from dividing uncontrollably using the patterns of releasing DNA they had found.

Using the Boolean logic gate model, the scientists created an AND gate genetic circuit design that combined with a CRISPR Cas-9 and sgRNA complex to detect and stop cancer cells in the bladder from gaining mass.

Using the hUP II and hTERT promoters as their inputs into the circuits, they were able to bind CRISPR and sgRNA together to produce a responsive signal in the gate. The hUP II promoter drove the transcription of the CRISPR Cas-9 RNA (which is a sequence of synthetic RNA that is used to identify a strip of complementary DNA and then cut using the Cas-9 protein), and the hTERT promoter was tied with the transcription of the sgRNA. The sgRNA is bound to the Cas-9 protein, and once together, are bound to the logic gate. The effect of the logic gate can only be expressed when both the protein and the RNA strand are present.

Once both these molecules were bound to the logic gate, the sgRNA targeted LacI (an operon used for the transport and regulation of lactose in the body and cell). This LacI controlled promoter drove the expression of the output genes, among which were hBAX and p21 (depending on which section of DNA was cleaved by the Cas-9 protein), who enabled apoptosis and growth arrest in the cancer cell.

But why the hUP I and hTERT promoters?

hUP II is bladder specific, which means that it is only found in the bladder, and the enzyme that binds to it is also only found in the bladder. The hTERT promoter is cancer-specific — this means only in bladder cancer are these two promoters together. This means that the logic gate does not attack healthy cells, only cells with the criteria for both promoters.

Cancer cured? No

Although this is a significant step into creating a better cure for cancer than alternative chemotherapy methods, it is still impractical.

For one, there has to be a new logic gate devised for every type of cancer that is out there (and that’s a lot). Furthermore, at the moment, both promoter regions must be strong, meaning that they must be able to bind their respective enzymes well, otherwise, the gate won’t work.

This work is still in the early stages of development, however, once logic gates and the detection of strands of DNA for different types of cancer becomes mainstream we might see a leap in genetic logic gate technology in the cancer field.

Takeaways:

  • cancer has been around for a long time, but now, faster than ever, we are coming up with new and faster techniques of fighting it.
  • Synthetic biology can offer a way into the future of fighting cancer.
  • Genetic logic gates can offer a new way of looking at cancer treatment.
  • Although these technologies aren’t mainstream today, and can’t be yet, with some time, a lot of tests, and many trials and errors, synthetic biology and logic gates could become the new norm for fighting off cancer.

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