December 9, 2009
Hi-Tech Microscopes Make Androgen Therapy ‘Personal’
On rare occasions, an infant is born with outward appearance of a female but the XY chromosomes of a male. If the child has a normal Y chromosome -- the chromosome responsible for testicular development -- the condition is known as androgen insensitivity syndrome.
Experts estimate such births occur in about one in 20,000 infants. Other children are born with a partial form of the condition that can affect their genitalia and/or fertility, but how many is not known.The cause is a wide range of androgen receptor (AR) mutations that fail to perceive the presence of the male hormones testosterone and dihydrotestosterone to differing degrees. How to overcome the problem remained a mystery until Baylor College of Medicine and Michael E. DeBakey Veterans Affairs Medical Center experts used a high throughput, automated microscopy technique called high content analysis to solve the puzzle. A report of their findings appears in the current issue of PLoS One, an open access journal.
Reverse effect of mutation
They not only identified the functional abnormality of the AR, but also used high content analysis to "personalize" a treatment that reverses the effects of that mutation.
"With this microscopy technique, we have been able to quantify how the receptor moves and functions inside cells taken from children with normal receptors and in those with the mutation," said Dr. Marco Marcelli, professor of medicine-endocrinology at BCM and a physician at the Michael E. DeBakey Veterans Affairs Medical Center. He and Dr. Michael Mancini, associate professor of molecular and cellular biology at BCM, and director of its Integrated Microscopy Core, are senior authors of the report.
Androgen insensitivity syndrome
They used banked cells taken from patients "“ both those with the mutation and those without "“ to study the action of the receptor in cell cultures grown in the laboratory. Dr. Michael J. McPhaul, a collaborator on the study and a professor of internal medicine"”endocrinology at The University of Texas Southwestern School of Medicine Dallas, provided the samples from patients with androgen insensitivity syndrome.
"We did this on a cell-by-cell basis, using high content analysis," said Mancini. "It is a proof-of-principle study carried out as though we had a patient and a library of hormones. We tried to find the perfect hormone for the mutation through high-speed collection of dozens of measurements from thousands of cells."
"In two of the three specimens we tested, we found we were able to reverse the activity of the mutated receptor to almost normal," said Marcelli.
In one patient, large doses of the male hormone dihydrotestosterone were sufficient. In another, they used a synthetic androgen that also activated the receptor.
These approaches overcame the central problem "“ the mutation changes the shape of the receptor and prevents it from maintaining normal contact with the hormone. It is as though a key is bent and can no longer turn the tumblers in a lock. In these cases, the hormone is designed to go into a pocket created by the receptor. When the pocket is changed by the mutation, the hormone is unable to establish good contact.
"Large amounts of testosterone may create more stable contact," said Marcelli. "The synthetic androgen may have a conformation that establishes better contact."
In the future, scientists may be able to screen large banks of such compounds to find a "superandrogen" that may be even more efficient.
"We might be able to use this technique to create a personalized medicine test," said Mancini.
Similar techniques might be used to screen drugs for treatment of different cancers, particularly those in which the androgen receptor is responsible for cancer progression. This study proves that the concept is valid providing quantitative information collected quickly on numerous measurements normally requiring separate biochemical tests and huge numbers of cells.
Marcelli said they have yet to use this kind of technique in patients, and such studies will require careful preparation, and go through a variety of approvals before it can be used in clinics. He also said it would be used only in individuals with the partial form of the syndrome. Finding well-matched hormones to defective androgen receptors through screening of thousands of compounds from available libraries could be one of the future developments of this technique.
The paper's first author, Dr. Adam T. Szafran, an M.D./Ph.D. student who worked in Mancini's laboratory, championed these studies, said Mancini. Szafran is now finishing the clinical part of his studies at BCM.
Others who took part in this study include Drs. Sean Hartig and Ivan P. Uray, Maria Szwarc, Jennifer Bell, Huiying Sun, Yuqing Shen and Sanjay N. Mediwala, all of BCM. Sun, Shen and Mediwala are also of the MEDVAMC.
Funding for this work came from the National Institute of Diabetes and Digestive and Kidney Diseases, the John S. Dunn Foundation and the Veterans Administration.
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