Several decades from now we hope to have sophisticated medical nanorobots, produced by molecular manufacturing, that can enter cells, analyze the state of the cell, and initiate appropriate therapy, such as killing cancer cells. A team of scientists from Harvard University, MIT, and the ETH in Zurich, Switzerland has taken an important step in that direction by demonstrating a synthetic circuit that, when incorporated into a cell, detects the presence or absence of five specific small RNA molecules,processes that information, and then, based upon that result, either kills or does not kill the cell. ScienceDaily reprints an ETH news story written by Peter Rüegg “Profiler at the cellular level“
Researchers led by ETH professor Yaakov Benenson and MIT professor Ron Weiss have successfully incorporated a diagnostic biological “computer” network in human cells. This network recognizes certain cancer cells using logic combinations of five cancer-specific molecular factors, triggering cancer cells destruction.
Yaakov (Kobi) Benenson, Professor of Synthetic Biology at D-BSSE, has spent a large part of his career developing biological computers that operate in living cells. His long-term goal is to construct biocomputers that detect molecules carrying important information about cell wellbeing and process this information to direct appropriate therapeutic response if the cell is found to be abnormal. Now, together with MIT professor Ron Weiss and a team of scientists including postdoctoral scholars Zhen Xie and Liliana Wroblewska, and a doctoral student Laura Prochazka, they made a major step towards reaching this goal. In a study that has just been published in [Science abstract], they describe a multi-gene synthetic “circuit” whose task is to distinguish between cancer and healthy cells and subsequently target cancer cells for destruction. This circuit works by sampling and integrating five intracellular cancer-specific molecular factors and their concentration. The circuit makes a positive identification only when all factors are present in the cell, resulting in a highly-precise process. Researchers hope that it can serve a basis for very specific anti-cancer treatments.
Selective destruction of cancer cells
The scientists tested the gene network in cultured human cells: cervical cancer cells, called HeLa cells, and normal cells. When the genetic biocomputer was introduced into the different cell types, only HeLa cells, but not the healthy ones, were destroyed.
Extensive groundwork was required to achieve this result. Benenson and his team had to first find out which combinations of molecules are unique to HeLa cells. They looked among the molecules that belong to the class of compounds known as microRNA (miRNA). The researchers had identified one miRNA combination, or profile, that was typical of a HeLa cell.
This was a challenging task. In the body there are about 250 different healthy cell types. In addition, there are numerous variants of cancer cells, of which hundreds can be grown in the laboratory. The diversity of miRNA is ever greater: between 500 to 1000 different species have been described in human cells. “Each cell type, healthy or diseased, has different miRNA molecules switched on or off,” says Benenson.
… It turned out that a combination of only five specific miRNAs, some present at high levels and some present al very low levels, is enough to identify a HeLa cell among all healthy cells.
“The miRNA factors are subjected to Boolean calculations. They are combined using logic operations such as AND and NOT, and the network only generates the required outcome, namely cell death, when the entire calculation with all the factors results in a logical TRUE value”, says Benenson.
The researchers were able to demonstrate that the network works very reliably in living cells. It correctly combines all the intracellular factors using a prescribed molecular “program” and gives the right diagnosis. This, according to Benenson, represents a significant achievement in the field.
… “We are still very far from a fully functional treatment method for humans. This work, however, is an important first step that demonstrates feasibility of such a selective diagnostic method at a single cell level,” said Benenson.
The researchers constructed what they describe as a “classifier” gene circuit that is transiently expressed inside a cell and then integrates information from five molecular markers to determine the state of the cell, and then produces a protein that sets off the cellular suicide cascade if the cell is determined to be cancerous. The DNA circuit they constructed contains numerous control sequences chosen from standard genetic engineering toolkits that respond to specific miRNAs such that only the combination that identifies the particular cancer cell line used in the experiments activates the circuit and triggers the onset of cellular suicide. The results presented do show some false positives and some false negatives, so further optimization of the genetic circuit would be needed. Nevertheless, the results are impressive. Also, in principle, this method could be adapted to different cell types by choosing the combination of miRNAs appropriate to distinguish that cancerous cell from neighboring cells. Perhaps the biggest problem to be faced in gearing up for animal tests is finding a way to deliver the genetic circuit in animals. These experiments were all done in cell culture, and transient expression involves treating the cells with chemicals to induce the cells to take up and briefly express foreign DNA. To use such a genetic circuit in animals (or humans) the DNA circuit must be protected from degradation in the blood stream and it must be inserted into cells in a way that it ends up being efficiently expressed inside the cell rather than being rapidly degraded by the cell. Over the past decade the biotechnology industry has investigated many different types of nanoparticles for use in genetic engineering, so perhaps one or more of these will turn out to be useful. For an additional level of specificity, such nanoparticles could be outfitted with molecules that preferentially attach to cancer cells.
