By Anette Breindl
Science Editor

Their company may be young and, for the time being, mainly virtual, but the folks at Carolus Therapeutics do not shy away from thinking big. It’s named after Carolus Magnus, or Charlemagne, who in the first century A.D. founded an empire that eventually encompassed much of Europe. Carolus’ scientific co-founders are located in Aachen, Germany, the main residence of the original Carolus. The company’s business operations are in the New World, and headquartered in San Diego. Carolus was founded in 2007 with seed funding from Avalon Ventures, where Carolus CEO Jay Lichter also is managing partner.

The company, Lichter told BioWorld Today, is fundamentally about “a novel approach to cytokines” that is based on “a more complex view of how cytokines work.” It currently is testing out that view on two programs, both of them, for the time being, preclinical.

The typical approach to targeting cytokines, Lichter said, has been to take a single cytokine or a single receptor and agonize or antagonize it – an approach that is often too simplistic.

“There have been a lot of failures on that space,” he added.

The company’s first program targets macrophage migration inhibiting factor. That cytokine usually helps activate immune responses by suppressing the anti-inflammatory effects of glucocorticoids. But it is elevated in several diseases, including rheumatoid arthritis and atherosclerosis.

Macrophage inhibiting factor was discovered in the 1960s and has been “well studied” in basic research, Carolus Chief Scientific Officer Joshua Schultz told BioWorld Today. But for a long time, those basic findings did not translate into making it a great target “because no one knew what the receptor was,” he said.

In findings published in Nature Medicine in 2007, the Technical University Aachen researchers identified two receptors, CXCR2 and CXCR4, that both bind macrophage inhibiting factor. When MIF’s interactions with both receptors were simultaneously inhibited, plaques regressed in mice with advanced atherosclerosis, and the remaining plaques had fewer monocytes and T cells in them, suggesting less inflammation within them.

Based on those and other results, Carolus is developing a dual antagonist for both receptors – an unusual approach.

“No one that I’m aware of is making a dual antagonist,” Lichter said, though he added that, of course, many early stage drug development programs are not public knowledge.

Nevertheless, the general strategy in drug development is to find a compound with maximal specificity for one receptor. But in this case, perhaps “you don’t want to be as clean as possible.”

In the Jan 4, 2009, issue of Nature Medicine, researchers from the Technical University Aachen reported new findings that are the basis of Carolus’ second program.

By targeting the interaction of two different chemokines called PF4 and RANTES, they were able to slow the formation of atherosclerotic plaque in susceptible mice.

PF4 and RANTES are secreted by platelets; when they bind to each other, they recruit immune system cells to the arterial wall, causing the inflammation that is characteristic of atherosclerosis.

The researchers designed a series of peptides to disrupt the interactions between RANTES and PF4. They then selected one of those peptides, termed CKEY2 for further study in vitro and in vivo.

In vitro, CKEY2 inhibited the recruitment of monocytes to endothelial cells. In in vivo experiments, they found that mice that are genetically predisposed to develop atherosclerosis when fed a high fat diet did not develop chronic arterial inflammation when injected with the mouse version of the CKEY2 peptide.

Attempts are being make by other groups to inhibit RANTES function in general; besides forming complexes with PF4, different RANTES molecules also bind to each other to regulate immune responses. In animal studies by other groups, blocking RANTES directly directly also has been successful in preventing diet-induced atherosclerosis.

But in their Nature Medicine paper, the Technical University Aachen scientists noted that “CCL5 antagonism may have therapeutic liabilities, as suggested by studies using Ccl5-deficient mice, which demonstrate that a complete absence of CCL5 compromise systemic immune responses, delay macrophage-mediated viral clearance and impair normal T-cell functions.”

Schultz noted that because knockout animals completely lack RANTES signaling, and therapeutic receptor blockage presumably would still allow a low level of signaling, the effects seen in knockouts “may or may not” be a good indicator of what to expect with a pharmacological RANTES receptor blocker drug.

But, he added, “animals treated with RANTES receptor blockers have another problem. Some animals treated with RANTES receptor blockers develop kidney inflammation, specifically a condition known as glomerulonephritis. The effect may be due to systemic immune reactions, including the removal of apoptotic cells and inducible nitric oxide synthetase expression. Whatever the cause, it highlights potential issues with direct antagonism at the CCR5 receptor,” he said.

In contrast, “at present, we have seen no indications that CKEY2 (or it’s murine ortholog, MKEY) interferes with systemic immune responses and/or functions. . . . Thus, we believe selective disruption of RANTES-PF4 heteromers has a therapeutic advantage, e.g. over direct RANTES receptor antagonism, in that it should permit normal immune defense and surveillance mechanisms.”

Both papers deal with cardiovascular indications. But, Lichter noted, cardiovascular trials are large and complicated, making them difficult for a start-up.

For that reason, the Carolus team is planning to start development of their compounds in other indications, with the hope that once another indication has delivered a proof of concept the company will be able to find a bigger partner for cardiovascular trials.

As for what those initial indications will be, that depends on the data now being generated in preclinical experiments; the company is testing its compounds in several animal models of inflammatory disease, both cardiovascular and others.

“As we generate the data, we will decide” which indications are most promising, Schultz said, with Lichter adding that the goal is to identify a good combination of activity with a “feasible development plan” for a small company like Carolus.