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King's College Spinoff Using MRM-MS for Hemoglobin Screening, Aims to Offer DTC Proteomics Testing

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By Adam Bonislawski

Scientists at King’s College London and Guy’s and St Thomas’ NHS Foundation Trust have launched SpotOn Clinical Diagnostics, a firm specializing in mass spec-based diagnosis of hemoglobin disorders.

Based on multiple-reaction monitoring assays, SpotOn is able to identify a range of hemoglobinopathies, including sickle cell anemia and thalassemia, from dried blood spot samples, Neil Dalton, professor of pediatric biochemistry at King's College and one of the company’s founders, told ProteoMonitor.

SpotOn, Dalton said, plans to offer the assays as an alternative to the HPLC- and isoelectric focusing-based assays currently used to screen newborns for hemoglobin disorders. Roughly 750,000 babies are born in the UK each year, all of which are tested for hemoglobinopathies.

The company has also developed MRM-MS assays for a variety of proteins linked to nephrotoxicity, and plans to offer them to support Phase I drug trials. SpotOn has also developed assays to measure albumin/creatinine to track patients’ renal and cardiovascular health, and hopes to eventually offer those as direct-to-consumer products.

In pursuing these indications, SpotOn is tackling the “low-hanging fruit” of clinical proteomics, Dalton said. Much proteomics research – cancer biomarker work, for instance – has focused on identifying and measuring low-abundance proteins, a goal that has proven quite challenging due to factors like biological and experimental variability and instrument limitations.

SpotOn, Dalton said, has chosen instead to focus on high-abundance clinically relevant proteins like hemoglobin that can be quantified quickly and reliably with a minimal amount of sample prep.

“If you’ve got a blood spot sample, [hemoglobin] is one of the highest abundance proteins that you’ve got,” he said. “That’s the beauty of it.”

“We’re using a very simple sample prep protocol,” he said. “In essence we’re taking the blood spot; we put in a stable isotope internal standard; we add the trypsin, incubate for half an hour to an hour, dilute, and run [on the mass spec].”

The simplicity of the workflow also increases its throughput, another issue that clinical proteomics and protein biomarker work, in particular, has struggled with (PM 2/11/2011). According to Dalton, his lab can process up to 700 samples per hour for analysis by SpotOn’s MRM-MS assays. Mass spec acquisition time is roughly one minute per sample.

In 2008, Dalton, along with his SpotOn co-founder and King’s College researcher Charles Turner, used their MRM-MS assays to screen 40,000 newborns for hemoglobinopathies, comparing the method to conventional HPLC and IEF techniques.

Using an AB Sciex 4000 QTRAP, the researchers ran MRM assays for HbS, HbC, HbE, HbDPunjab, HbOArab, as well as normal beta, alpha, gamma, and delta hemoglobin chains, identifying sickle cell disorders and thalassemia major. According to a poster detailing the results, the mass spec-based workflow performed as well as conventional methods and offers the prospect of savings through lower consumables costs and the potential for multiplexing with newborn metabolic tests currently done via mass spec.

The researchers’ mass spec-based hemoglobinopathy assays received an EU patent last year, Dalton said, calling it one of the main drivers behind the decision to launch SpotOn. The company is also pursuing patents on the stable isotope internal standards used in the assays and on multiplexing the measurements of proteins, enzymatic activity, and metabolites from a single blood or urine spot.

According to Dalton, SpotOn has hemoglobinopathy screening kits ready to sell, and is awaiting regulatory approval. In the meantime, he said, the company has provided the assays to an unnamed UK customer who plans to use them for smaller-scale hemoglobinopathy screening efforts comprising around 36,000 patients per year.

Provided the test receives approval, Dalton said the company plans to start with the UK’s 750,000 newborn-per-year market and rely on that business to fund expansion into the rest of Europe, Asia, and the US. Thus far, he said, SpotOn has declined to seek outside funding, relying instead on money from King’s College and Guy’s and St Thomas’ NHS Foundation Trust. The company also has revenues from research use of its assays, Dalton noted.

“We’ve been given some pump-priming money from [King’s College and Guy’s and St Thomas’], and the income stream from the research services we offer [is] also put into the company,” he said. “So we immediately from day one had a significant income stream that is being used to make sure patent costs are met and development proceeds.”

Dalton and SpotOn aren’t alone in focusing on more straightforward applications of proteomics to the clinic. In a talk at the Human Proteome Organization's 10th annual meeting in September, Denis Hochstrasser, chairman of Geneva University Hospital's Genetic & Laboratory Medicine Department and founder of the Swiss Institute for Bioinformatics and GeneBio, similarly suggested that in concentrating on difficult questions like cancer biomarkers, the field might have overlooked more immediate clinical uses for proteomics.

In a later interview with ProteoMonitor, Hochstrasser cited hemoglobinopathy screening as among the most obvious clinical uses for mass spec-based proteomics (PM 10-28-2011).

Because hemoglobin is present in the blood “in such a large concentration, you don't have to worry about any sensitivity issues,” Hochstrasser said at the time. “You can inject the red blood cells directly into the mass spec – you don't even need chromatography up front.”

He also noted that while current assays for quantitative disorders like thalassemias and qualitative disorders like sickle cell require two different tests, these assays could be combined into one using mass spec.

“With one mass spec test we can do several tests faster and cheaper than we are currently doing in clinical practice,” Hochstrasser said, adding that his lab was working on assays for hemoglobinopathy screening and hoped to have them in the clinic by early 2012.

Beyond moving mass spec into the clinic for hemoglobinopathy screening, SpotOn also has designs on the direct-to-consumer market, Dalton said. In particular, he noted, assays measuring things like albumin/creatinine ratios to monitor renal and cardiovascular health or glycated hemoglobin to monitor diabetes, could be well suited to such a format.

SpotOn’s use of dried blood spot samples makes its assays particularly amenable to a DTC model because of the stability and durability of these samples. As Stephen Johnson, a researcher at Arizona State University's Biodesign Institute, told ProteoMonitor during a February interview discussing his lab’s interest in offering DTC proteomics testing, patients could send samples they provide via fingerpricks through the mail for analysis (PM 2/4/2011).

"Our big idea is that this is what healthy people will be sending in on a regular basis for diagnostic purposes," Johnson said at the time. "The idea is that people could send a drop of blood through the mail and we could do the [assay] on that."

Dalton, likewise, advocates such an approach. “We are working on direct-to-consumer,” he said. “Because that is personalized medicine. If you look at all the efforts that have been made trying to diagnose renal disease early, the problem is that you are always compared against the normal range.”

“These markers have very low individual variability but relatively high variation across a normal population,” Dalton said. “Therefore you miss the changes. The key is to track an individual over time. You might have a [glycated hemoglobin level] in the normal range, but if it’s risen, then you know you’re developing diabetes. So you might be able to pick up the disease ten to 15 years before it’s clinically present.”

“We’re not interested in [DTC testing] in relation to hemoglobinopathy, but we’re certainly interested in it in relation to kidney disease, diabetes, anyone with a family history of cardiovascular disease, and so on,” he said.


Have topics you'd like to see covered in ProteoMonitor? Contact the editor at abonislawski [at] genomeweb [.] com.

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