Giving New Life To An Old Molecule

Well-known for 135 years as a biological stain and chemical indicator, the heterocyclic aromatic compound called methylene blue has also been used as a textile dye and an antimalarial drug. Medical researchers have investigated its effect against cancer, Alzheimer’s disease, and viral infections. The compound still is administered as a urinary antiseptic and as a dye for visualizing tissues during medical procedures. It even kills fungus in fish tanks.

Most critically, methylene blue, or meth­yl­thioninium chloride, is the single most documented and clinically used substance to treat a life-threatening condition called methemoglobinemia, according to the European Medicines Agency (EMA). When a high level of methemoglobin, the oxidized form of hemoglobin, forms as a result of certain drugs or toxins in the blood, meth­yl­ene blue is the antidote that converts it back to hemoglobin.

But the methylene blue that’s administered has been more dyestuff than pharmaceutical. Because producing a high-quality material is difficult and alternative treatments are limited, purity standards established in monographs by U.S. and European pharmacopoeial organizations have permitted high levels of toxic heavy metals in methylene blue.

Believing there’s a better option, a small French start-up, Provepharm, is close to bringing pharmaceutical-grade methylene blue to market. As produced by its partner Novasep, a contract manufacturing firm in Chasse-sur-Rhône, France, Provepharm’s Proveblue is made under current Good Manufacturing Practices (cGMP) and meets drug industry standards on residual metals and other impurities. The product has moved quickly through European regulatory review.

“Our goal is to reintroduce methylene blue as a new active pharmaceutical ingredient, where the dye becomes a drug,” explains Michel Féraud, Provepharm’s chief executive officer. Others have tried before to make a purer version but were stymied by the strong chelating nature of the molecule itself. Traditional synthesis methods that use metal-containing reagents leave behind contaminants, and organic impurities are hard to remove because of their similarity to the desired product itself.

Several years ago, Provepharm’s parent, the Marseille-based contract R&D firm Provence Technologies, decided to try its hand at purifying dyestuff-grade methylene blue using material already available on the market. But just cleaning up a raw material isn’t enough to satisfy regulators for a new marketing authorization. And a manufacturer must demonstrate a well-defined and controlled process of a few synthetic steps under cGMP conditions.

In the Provence Technologies process, low-purity methylene blue is chemically protected to veil its chelating nature. Contaminating metals are removed by filtering the material through a support that traps them. The protecting groups are then removed without using a metal-based oxidizing agent. Washing and crystallization further remove organic impurities.

After developing the route and protecting the process with a patent, Provence Technologies spun off Provepharm as a separate company. Provepharm began operations in 2008, and its goals were to scale up and validate the process and to produce material to support a filing with EMA for an injectable form of the antidote to treat methemoglobinemia.

By early 2009, Provepharm was preparing to file and needed a reliable manufacturing partner for the active pharmaceutical ingredient (API) to meet its timeline. “We are a start-up and wanted to get to the market fast,” says Provepharm R&D Director Jeff Kaufman. It turned to Novasep.

Novasep scientists first familiarized themselves with the Provence Technologies process and then worked on scaling it up from lab to pilot scale.

Nothing was unusual in this request except that lab scale had meant working in glassware. Making the strong chelating agent under entirely metal-free conditions at large scale would be more challenging. Novasep succeeded, says Johann Oviste, Novasep’s business manager in charge of the project. Within seven months, it had produced one demo and three process validation batches of Proveblue.

Novasep also assisted in creating a drug master file and the documentation to support a marketing authorization application for Proveblue’s use as a pharmaceutical. The company helped develop and validate new analytical methods because existing methods for the older and less pure material were out of date.

According to Babak Sayah, coinventor of the patented process, Novasep was able to meet several needs. “On the one hand, our process is in fairly dilute solution and the volumes are quite large. And on the other hand, we needed a company that wouldn’t be fazed by handling a blue product, which is inherently messy,” he says. “We also needed a partner that had the capabilities to support our regulatory group and could give us the whole package, not just say they could make the product. But most important to us was the speed at which they worked.”

About a month after producing the first batches, Novasep’s facility passed an inspection by French regulators, Oviste says. Working throughout 2010, the partners then began a development plan to expand the process to produce at commercial scale.

Just last month, an EMA committee recommended that the injectable formulation of Proveblue be approved for treating methemoglobinemia. The small company’s big bet on getting European Union-wide approval and being able to sell in all 27 countries looks close at hand. The first marketing authorization offers substantial commercial potential, with annual revenues forecast to reach $40 million to $50 million within five years, according to Féraud. The company is looking for distributors in the different markets covered by the approval.

Provepharm hopes that the product’s approval and its availability will renew interest in methylene blue-based drugs. For its part, the company wants to file for approval in other countries and for other disease indications, which may require research and clinical trials. “We closed a deal and have active discussions ongoing with potential licensees,” Kaufman adds, including with those in the U.S. and Japan.

Because methylene blue was already registered, the EU filing process was the same as for a generic drug. In other countries where no methylene blue products are registered, the regulatory path is somewhat unclear. Kaufman says Provepharm soon plans to discuss how to proceed with the U.S. Food & Drug Administration.

He thinks the company has a convincing case. Provepharm believes that it can now offer an API with a significantly improved risk-benefit profile. Metal levels in Proveblue are 1 to 2% of those specified in drug monographs. “We’ve been able to show that there is reduced toxicity when the heavy-metal impurities are removed,” Kaufman adds. Proveblue’s overall impurity profile meets international standards for pharma ingredients.

In addition to upgrading the purity, the partners say, they have increased the value of the API at least 10-fold. In fact, Proveblue may establish new standards for methylene blue purity and push less pure versions out of the human health market. This seems to be happening already. In November 2010, French regulators canceled the import license on product coming into the country and allowed Proveblue to be used before approval under “compassionate use” legislation.

Provepharm intends to petition the European Pharmacopoeia and the U.S. Pharmacopeia to update their monographs now that a purer methylene blue can be made. “The likelihood that there will be a harmonized specification is high,” Féraud contends.

The path from process development through scale-up, validation, production, and formulation and toward approval has taken only about two years. The partners anticipate eventually having to make ton quantities. To that end, Novasep is investing in the next stage of scale-up on its own, Oviste says, marking yet another step on the road to a pharmaceutical-grade meth­yl­ene blue.