A Progress Report from the Celiac Sprue Research Foundation

Celiac.com 07/23/2022 - With the aim of harnessing science to improve the quality of life of celiac sprue patients, in 2002 my colleagues and I started the non-profit Celiac Sprue Research Foundation. The first challenge the Foundation undertook was arguably the most ambitious—to translate available knowledge about celiac sprue pathogenesis into a comprehensive plan for developing therapeutic alternatives to the complete and lifelong gluten exclusion diet. Two years later this remains the primary goal of the Celiac Sprue Research Foundation, one that we continue to pursue with vigor and commitment.

This article will review the current status of the Foundation’s frontrunning drug development project aimed at developing an oral enzyme pill to protect a celiac from the toxic effects of gluten. This effort was initiated based on results reported in two publications in 2002 (Hausch et al, Am. J. Physiol. 283, G996- G1003, 2002; and Shan et al, Science 297, 2275-2279, 2002). In essence those papers proposed that:

• (i) Among dietary proteins, gluten is unusual because it is especially difficult to break down completely into nutritionally useful amino acids. Indeed, some incompletely digested products of gluten persist in the human gut for an exceptionally long time;
• (ii) In most humans, this does not present a problem. However, in celiac sprue patients, digestive resistance of gluten is problematic because the residual products induce an inflammatory response; and
• (iii) A specific type of enzyme called prolyl endopeptidase (PEP) can mitigate this problem by accelerating the destruction of toxic products generated as a result of incomplete gluten digestion.

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Since the publication of those initial findings, a series of increasingly complex studies have been performed to assess the validity of the proposals described above. The findings of these studies have either been recently published or are being prepared for publication in peer-reviewed scientific journals. Below I summarize key results from three recently published papers that, in aggregate, lend considerable support to the hypothesis that a PEP pill may indeed be an effective way to treat celiac sprue.

1) Piper, J.L., Gray, G.M., and Khosla, C. Effect of prolyl endopeptidase on digestive-resistant gliadin peptides in vivo. Journal of Pharmacology and Experimental Therapeutics. In press. (E-Publication available online through PubMed).

This work was part of the doctoral thesis of my former student Justin Piper at Stanford University. Using a challenging procedure pioneered by Gary Gray, Justin investigated the digestive resistance of a panel of gluten peptides in live rats (Yes, when it comes to the digestive system, rats are indeed very similar to us!). This panel of gluten peptides was chosen to reflect the diversity of opinions different scientists have regarding how gluten triggers the disease. Remarkably, but perhaps not surprisingly, nearly all of these potentially problematic peptides were highly resistant to breakdown in the rat gut. By contrast, the most stable product of myoglobin (a nutritionally important protein from meat) was rapidly and completely digested. Justin then went on to investigate the effect of PEP on the breakdown of one of the least digestible gluten peptides. He observed that, when therapeutically reasonable quantities of PEP were dosed in the rat gut, virtually all of this toxic peptide could be destroyed in less than 10 minutes.

2) Shan, L., Marti, T., Sollid, L.M., Gray, G.M., and Khosla, C. Comparative biochemical analysis of three bacterial prolyl endopeptidases: Implications for celiac sprue. The Biochemical Journal In press. (E-Publication available online through PubMed)

This work was pursued in collaboration between Lu Shan, a doctoral student in my lab at Stanford University, and Thomas Marti, Maribeth Evelyn Lynn Research Fellow at the Celiac Sprue Research Foundation. Most of our studies thus far have utilized a PEP from the bacterium Flavobacterium meningosepticum. While this PEP has many attractive features, it did not evolve to break down gluten, and is by no means a perfect solution for the problem at hand. As we learn more about how gluten triggers celiac sprue, we believe we can do a much better job of designing an optimal enzyme drug. With this in mind Lu and Thomas set about comparing the properties of three naturally occurring PEPs. They found many similarities and differences, suggesting that, in the long term, it may be possible to recombine the good features from different enzymes into a single mousetrap. In the near term, however, they were able to work around a major problem with the Flavobacterium enzyme—its high cost. Specifically, they identified a PEP from an unrelated bacterium, Myxococcus Xanthus, that appears to be comparable to the Flavobacterium PEP in all respects but is substantially cheaper to produce. Assuming further studies can confirm this equivalence, we expect to use the Myxococcus enzyme in future animal and human testing.

3) Marti, T., Molberg, O., Li, Q., Gray, G.M., Khosla, C., and Sollid, L.M. Prolyl endopeptidase mediated destruction of T cell epitopes in whole glutenChemical and immunological characterization. Journal of Pharmacology and Experimental Therapeutics In press. (E-Publication available online through PubMed)

This study was a collaborative effort between scientists at the Celiac Sprue Research Foundation and the University of Oslo. As is well known, grocery store gluten is an extremely complex substance consisting of literally hundreds of proteins. Prior to this study, all of our investigations focused on individual proteins from gluten or smaller pieces (peptides) derived from individual proteins. Consequently, demonstrating that our key conclusions also apply to grocery store gluten represented a major milestone. To do so, Thomas Marti and Qing Li at the Celiac Sprue Research Foundation developed a procedure that reliably mimics in a test-tube the effects of the digestive system on grocery store gluten. These test-tube procedures were then used to analyze the composition of residual gluten before and after PEP treatment. They showed that nearly all of the toxic peptides that could be identified in untreated gluten were eliminated from PEPtreated gluten. To verify the relevance of this observation, they teamed up with Oyvind Molberg at the University of Oslo, who had isolated a panel of gluten-responsive T cells from the small intestines of 14 celiac sprue patients. In a pivotal experiment, Molberg showed that PEP-treated gluten samples had dramatically reduced toxicity towards T cells from 12 out of 14 patients. Thus, to the extent the Oslo procedure accurately reflects the primary cause of gluten toxicity in celiac sprue patients, it appears that PEP may indeed be able to render grocery store gluten safe for celiac sprue patients.

In summary, a considerable body of additional laboratory evidence has emerged over the past two years in support of the potential utility of an oral PEP enzyme as a therapeutic alternative to total gluten exclusion for celiac sprue patients. For those of you who are inclined, I encourage you to download the above papers, and assess the data for yourself. Regardless of your scientific inclinations however, it is also important to understand what has not yet been accomplished and what remains to be done.

Thus far, to our knowledge, no studies of PEP have been conducted in humans. Doctors at the Celiac Sprue Research Foundation have recently completed a preliminary study aimed at determining whether gluten can be rendered harmless for celiac sprue patients by treatment with PEP outside the body. The results from that study, which could further bolster the case for enzyme therapy, are being analyzed, and will be published in due course. However, even if those results are encouraging, it is not the same as dosing a patient orally with the PEP enzyme with the intention of detoxifying ingested gluten. An initial study of that nature requires several hundreds of thousands of dollars (primarily for production of clinical-grade enzyme pills and for testing the safety of these pills in laboratory animals). A funding source for this effort has not yet been identified.

Meanwhile, more could be done to improve the quality of the enzyme pill, so as to maximize its chances for success in clinical trials. Finally, estimating the maximum dose of gluten that could be safely tolerated by a celiac in conjunction with an enzyme pill is also a major unanswered question that lies ahead of us. While celiacs may fantasize about going to a pizza parlor armed with a wonder drug, a more realistic goal for a first-generation therapeutic agent would be to take the worry out of a celiac’s daily life by enabling the patient to cope with modest quantities of gluten that seem to be all around us without the blatant garb of a hamburger bun or pizza slice.

I hope that the above progress report helps you understand that, while much remains to be done to develop a safe and effective therapy for celiac sprue, excellent progress is being made. My sincere thanks to all of you who have donated to the Celiac Sprue Research Foundation, without which none of this would have been possible. I trust that, thus far, we have kept the faith.