With its first microarray product scheduled to hit the market later this year, Alopex of Kulmbach, Germany, hopes to grab a piece of the expanding in vitro fertilization market.
Founded in 2001, the 15-employee company has been developing technology for the genetic analysis of single cells. With its first DNA chip — to be used in preimplantation diagnostics — Alopex aims to help improve the low odds of successful pregnancy and childbirth after IVF procedures. “The baby take-home rate should be increased,” said Wolfgang Mann, a company co-founder and CEO of Alopex, although he and his co-workers don’t know yet by how much.
Each of the six co-founders of the company brought in at least five years’ worth of experience at a chip company or molecular diagnostics firm, according to Mann, who himself introduced chip technology to the Max Planck Institute for Molecular Genetics in Berlin before joining MWG Biotech. After heading up the biochip department at MWG, Mann was looking for a promising chip-based application that would form the foundation of a company.
“With a small company of 15 people, it is not possible to compete with big pharma, big industry, or with IVD companies that … already produce things,” said Mann. What he and his colleagues sought, he said, was an application that yields easily interpretable results that —unlike expression profiling — can be linked to a definite disease outcome.
What they came up with is a preimplantation diagnostic application, which counts the chromosomes of polar bodies. The microarray contains probes for the sequence of different chromosomes on the polar bodies.
Polar bodies are attached to the egg, or oocyte, and represent “a mirror image” of the egg’s chromosomal content, according to Mann. Counting these chromosomes allows researchers to deduce chromosomal abnormalities of the fertilized egg, such as monosomies or trisomies, which can result in miscarriage, non-implantation, or disorders such as Down’s syndrome. Embryo biopsy, another form of pre-implantation diagnosis, is prohibited in Germany, Mann said, making polar-body analysis the only alternative. The analysis is permitted only during a short window of approximately 12 hours after fertilization, when the nuclei of sperm and egg have not yet fused, thus requiring a rapid diagnostic method.
To amplify the genetic material from the polar body, Alopex uses PCR and “related amplification techniques,” Mann said. To make the microarrays, scientists currently apply oligo probes to glass-slide-based chips by contact printing at the moment, but are “also testing other formats,” he said. Readout is fluorescence-based and is done using standard scanners.
Mann said he is not aware of any other company using a chip-based approach to polar body chromosomal analysis.
The first commercial version of the chip will cover about half of the 23 chromosomes, but Alopex is currently working on probes for the other half. The company has already been validating the technology in-house but has contracted about half a dozen IVF centers in Germany for a second validation phase, scheduled to start this summer.
This technology is an alternative to fluorescent in situ hybridization, or FISH, which is currently used for polar-body analysis but is not widespread in Germany yet. While the number of chromosomes possible to study in a single FISH assay is limited by the number of available fluorescent dyes, Alopex’s chip would eventually be able to interrogate all of the chromosomes, Mann said. And although most of the known disorders resulting from monosomies and trisomies occur on a small number of chromosomes, it would be better to cover all chromosomes: It is not that the other chromosome number abnormalities do not occur, “but you don’t find them because these oocytes might never start to divide and develop,” Mann said.
Frank Dechent, a human geneticist and head of a laboratory for cytogenetics and molecular biology at a private IVF center in Germany, agreed that a test covering all chromosomes would be useful. But he cautioned that data on whether polar body analysis actually increases pregnancy rate and baby-take-home-rate, and decreases miscarriage rate as well, is still controversial.
According to Dechent, pregnancy rates after implantation of in vitro fertilized eggs hover between 10 and 35 percent, depending on the institute performing the implantation.
Alopex’s chip-based test would cost €100 ($122) or less, Mann said. This pricetag is comparable to that of FISH analysis, according to Dechent.
The market could be sizable: In 2002, 70,000 IVF cycles were initiated in Germany, up from 60,000 the year before, according to a report by the German IVF Register. Assuming that each cycle results in about 10 oocytes, and 10 percent of these were to undergo polar body diag-nosis, Mann estimated, the German market volume for Alopex’s test could reach up to €7 million a year.
Initially, Alopex plans to market its chips to German IVF centers, which number less than 200, and expand to other European countries about two years later. Mann is not worried about insurance reimbursement for the test since IVF patients have to cover a variety of procedures out of pocket already, and do so readily, he said. “If you can tell a patient that the chance to get pregnant is doubled, or even more, … simply because of better diagnostics, they will pay everything.”
Dechent agreed that polar body analysis would become more widespread if its usefulness were actually proven. Right now, he said, it is only performed at a small number of centers in Germany. “If you show the [spontaneous] abortion rate will decrease, or the baby-take-home rate will increase, maybe even the pregnancy rate will be increased, then I think there will be a chance it will be performed quite often,” he said. According to Dechent, the analysis is especially promising for women over 38 who undergo IVF because among this group, up to 80 percent of the eggs can have an abnormal number of chromosomes.
Longer term, Alopex is considering other applications for its chip-based cell analysis technology, including sperm-typing and the analysis of mutations in primary tumors.
So far, the company has raised a total of €3.5 million in venture capital funding, from S-ReFit of Regensburg, Bayern Kapital of Landshut, and Sparkasse Kulmbach. In addition, it has won €700,000 in grant money from the German federal government, part of a €3.8 million project with Advalytix of Brunnthal and NMI of Reutlingen to develop an integrated microfluidics system for DNA amplification, hybridization, and detection.