New standards may help push more drugmakers to use a synthetic product for critical drug safety Testing, instead of one made from horseshoe crab blood that has long drawn criticism.

The U.S. Pharmacopeia (USP) announced a draft chapter on Endotoxin Testing that provides techniques for using “non-animal-derived reagents.” Its comment period will run from November 1 through Jan. 31, 2024.

“This approach advances USP’s commitment to transition methods from using animal-derived materials to synthetic and recombinant materials,” the group said in a statement.

“I think it’s an important step forward,” Larry Niles, PhD, a wildlife biologist in New Jersey who has long fought for greater protections for horseshoe crabs, told MedPage Today. “Now our goal will be to get companies to switch over.”

Jay Bolden, a senior biologist and director of quality at Eli Lilly in Indianapolis, who has championed his company’s leadership in adopting an animal-free endotoxin testing alternative, called it a “monumental, positive change.”

Endotoxin testing is an essential component of ensuring the safety of pharmaceutical products, especially injectable drugs and implants. It has been dominated since the 1990s by limulus amoebocyte lysate (LAL), which is made from the blood of horseshoe crabs, a species estimated to be more than 400 million years old.

Critics have called attention to the process for making LAL, which involves taking horseshoe crabs from estuaries along the Atlantic coast and bleeding them out for their bright blue blood. That blood contains the LAL enzyme that coagulates in the presence of endotoxins, a byproduct of bacteria.

LAL has been the global standard of safety testing for decades and is involved in the production of nearly every injectable pharmaceutical product and implantable device on the market, estimated to be used in some 70 million tests per year.

But critics have noted that the true mortality rates for horseshoe crabs after bleeding are unknown, and could be as high as 30%. The horseshoe crab population is considered to have declined substantially — they have also been overfished for bait — and its eggs provide necessary food for migratory birds that are endangered or threatened.

In response, a synthetic alternative to LAL, known as recombinant Factor C (rFC), was developed in the late 1990s by a single company and became commercially available in the early 2000s. Industry was slow to make the switch, at first in part because of limited data, supply issues, and also because regulatory guidance didn’t exist.

In 2012, however, the FDA issued guidance for incorporating rFC into endotoxin testing, Bolden said. Around the same time, a second rFC producer came online, he added. And as an avid birder, Bolden also knew that the threatened Red Knot relied on horseshoe crab eggs to fuel its long annual migratory journey — nearly 10,000 miles — from Tierra del Fuego in South America to its Arctic breeding grounds.

With the confluence of those factors, Bolden began looking more closely at rFC, and over time helped build a substantial body of data showing rFC was just as good as — if not better than — LAL.

By 2016, Eli Lilly was able to declare that all new products, along with key components, would be tested using rFC instead of LAL. The company brought the first-ever rFC-tested drug to market — its migraine drug, galcanezumab (Emgality) — and it currently has eight products approved using the synthetic alternative. Bolden says the company is now 80% converted to rFC instead of LAL.

“We thought it was better from a quality perspective, and the ethical piece lines up with our corporate policy on the care and use of animals, which says that if there’s an alternative to animal testing that’s acceptable to regulators and backed by the science, that’s something Lilly should be looking at,” Bolden told MedPage Today, noting that it’s also cost-effective.

Bolden added that when LAL was developed, it was an improvement over prior endotoxin testing, which involved injecting rabbits and waiting to see if they’d get a fever.

“LAL was great, it replaced a full-on animal test, and it protected patient safety for more than 30 years,” he said. “But now we have an alternative test reagent that … takes the animal part out of the equation.”

Bolden said peer companies, including Sanofi, Pfizer, and Roche, have made similar commitments to prioritize the synthetic alternative over LAL, and one small medical device company already has an approved product that was tested using rFC.

In addition to rFC, there’s another synthetic product for endotoxin testing called recombinant cascade reagent, or rCR, which involves three recombinant proteins rather than just one.

Bolden says rFC and rCR have similar efficacy: “From a scientific perspective, I think they’re equally fine and they’re both better than using an animal source test reagent,” he said.

All four rCR manufacturers are historical makers of LAL, Bolden said: Charles River Laboratories, Lonza, Associates of Cape Cod, and Wako.

In an email to MedPage Today, Samantha Jorgensen, a spokesperson for Charles River Laboratories, said the company is “pleased to see the USP is considering the proposal, as it is important to the future of the industry,” but noted that much “remains to be determined.”

“Recombinant technology will allow the industry to optimize resources and continue its mission to reduce reliance on the horseshoe crab by innovating formulations and microfluidics technology to enrich the LAL assay. However, it must do so without compromising patient safety,” Jorgensen said.

“Current recombinant technologies may not be suitable for all products produced by the biopharmaceutical industry,” she added. “As such, LAL will remain necessary to support safety testing programs.”

USP’s European counterpart, the European Pharmacopeia Commission, endorsed the use of rFC in 2019. USP had previously drafted a chapter in support of the use of rFC in endotoxin testing, but it was tabled after the comment period in 2020.

Bolden noted that USP’s current draft chapter includes use of both rFC and rCR.

“It’s pretty inclusive, so it’s hard to think that there will be negative comments,” he said, “but you never know.”