Researchers identify mutation in gene capable of regulating pain

Worldwide, 1.5 billion people suffer from pain. Even with several drug options available, not all forms are treatable and, when they are, there may be adverse effects, such as the development of dependence or tolerance – especially in the case of morphine and other opioids.

In search of new options for analgesics, researchers from the Special Laboratory of Pain and Signaling (LEDS) at the Butantan Institute studied a cell receptor called TRPV1, responsible for capturing harmful heat stimuli and the burning sensation of pepper. They found a mutation in the gene that encodes this protein that can cause loss of sensitivity to pain.

The survey results were published in the journal The Journal of Clinical Investigation. The study was supported by the State of São Paulo Research Foundation (FAPESP).

Analysis details

In partnership with Stanford and Emory universities, both in the United States, and the University Hospital of Münster, in Germany, the Brazilian group analyzed a series of mutations in humans. The authors also benefited from existing knowledge about birds – animals that are not very sensitive to harmful stimuli and indifferent to spicy foods precisely because of a mutation in the TRPV1 gene.

“There are more than a thousand mutations for the TRPV1 receptor in humans and it is not new to try to switch it off to relieve pain, but until today these attempts have not been successful”, says Vanessa O. Zambelli, researcher at LEDS and first author of the study alongside Stanford’s Shufang He.

“First, because many drugs resulting from this process interfere with body temperature and second, because, as it is an important channel for the sensation of harmful thermal stimuli, completely altering its activity nullifies the physiological pain, interfering with the sensation of heat, which has protective function”, he adds.

The first step for the researchers was to investigate a genome database to understand what the genetic sequence of the avian TRPV1 receptor was like and compare it with the genomic analysis of humans.

Through computational studies, five mutations were identified in birds that would be related to resistance to pain. Electron cryomicroscopy analyzes (in which samples preserved at low temperatures are observed under a microscope) showed that they are located in an amino acid residue (K710, one of the components that form the TRPV1 protein) that controls the opening and closing of the channel.

Although they can also exist naturally in humans, they are very rare. Scientists then asked themselves: what would happen if these mutations were transposed to mammals?

In studies of cells, it was discovered that, in this case, there was indeed a change in the function of the channel. Tests in mice with the mutation induced through the CRISPR/Cas9 gene editing technique confirmed the absence of nociceptive behavior (the term “pain” is not used for animals due to the difficulty of measuring the subjective and emotional component) after injection of capsaicin (compound active ingredient of pepper) in the paw and ingestion of feed (spicy) for birds, differently from what happened with wild mice.

Next, the cell body of the mouse sensory neuron was removed and, in cell culture, stimulated with capsaicin. Again, neurons reduced their ability to transmit nociception, which was also repeated in a more robust model of chronic pain, through sciatic nerve injury. All this without losing the ability to feel the harmful temperature.

In addition to modulating pain, TRPV1 also plays an important role in protecting against other stimuli. Previous studies show, for example, that its activation controls cytotoxicity induced by excess glucose. Additional tests carried out in this work, with removal and stimulation of cardiac cells (cardiomyocytes) by harmful agents, such as hydrogen peroxide and high glucose, in a model of cerebral ischemia, confirmed that the protective effect was maintained even with the mutation.

translational work

The second part of the study involved trying to reduce receptor function pharmacologically. For this, the researchers developed a peptide named V1-cal, which acts selectively in the region of residue K710.

Treated with this compound, animals that received capsaicin showed less nociceptive behavior. There was also a decrease in the release of neuropeptides that cause inflammation and swelling, without changes in temperature. Finally, chronic pain also improved considerably.

“Now, we want to add value to this study by validating it under good laboratory practice conditions. [requeridas pelas agências regulatórias] and to identify, in addition to the peptide, other small molecules that are easier to synthesize to advance in pre-clinical studies and, if all goes well, enter a clinical trial”, concludes Zambelli.