Helminthic Therapy
Drugs

Luca Allois
Pier Paolo Bocchino

For as long as modern humans have existed, they have carried parasitic worms. There are two groups of worms that inhabit the interior of humans: the nematodes (roundworms) and the platyhelminths (flatworms). They have distinct evolutionary histories and whereas some reside harmlessly in the gut others can cause problems. Most of these worms, however, fail to sustain human colonization, presumably because of genetic differences in the human host, and have no association with human illness (e.g., Trichuris suis).

Helminths have preferences for living in various locations of their host like the intestinal lumen, bile ducts, blood stream, lungs, and elsewhere. In order to accomplish this, they must evade and control the host’s immune system. This has been achieved through millions of years of coevolution allowing time for both the parasite and its host to gradually adjust to this relationship.

Many worms seem invincible to human defense. Some worms produce factors and receptors with homology to molecules of the human immune system as revealed by recently decoded worm genomes, for example schistosomula can acquire surface molecules from the host, including blood group determinants and MHC molecules, or they can produce cytokine mimics. It is likely that helminths sense hostile changes in the local host environment and take action to quell these responses. Many worms also have a peculiar outer integument that can absorb host molecules and which may assist in cloaking the parasitic organism from host detection.

Mechanisms of immunomodulation

Since the death of the host causes the death of the worm, a 'good' parasite, in order to survive, is an organism which imparts some advantage to its host. Clearly, after thousands of years of co-evolution, the human immune system has evolved to handle the presence of most parasitic worms, which have, in turn, developed adaptations that enable them to live for years in a human host.

The cloning of the genomes of some helminths has allowed a systematic search for substances with potential immunomodulatory activity. Analysis of the Trichuris suis’ transcriptome has led to the identification of more than a hundred proteins containing conserved domains or signatures relevant to inflammation and immunity, like proteases, protease inhibitors, kinases and other proteins. Whether these proteins are synthesized and what their activities are remain to be investigated.These molecules seem to have three major effects on the immune system (Autoimmunity: The worm returns, 2012).

First, they seem to cause changes that activate regulatory T cells such as Treg. These cells dampen immune responses and curb autoimmunity — by, for example, increasing the production of regulatory molecules such as interleukin-10 (IL-10) and Transforming Growth Factor-β (TGF-β).

Second, helminths seem to act on other cells, like regulatory dendritic cells and macrophages. These prevent the activation of dangerous effector T cells, which normally lead to inflammation and disease.

Both of these effects seem to protect from autoimmunity in a redundant fashion. This redundancy may be one of the reasons why helminths are so effective in controlling these immune-mediated diseases.

Third, worms seem to alter the bacterial composition of intestinal flora. Research in mice suggests that helminths promote the growth of gut microorganisms typically considered to be 'probiotic', which help to maintain intestinal health.

However, the pathways are very complicated and they are not yet fully understood; this is mainly due to the astonishing number of different molecules and cells activated or inhibited in response to the helminthic infection (Chronic helminth infections induce immunomodulation: consequences and mechanisms, 2007).

Infections with helminth parasites generally induce a strong polarization towards a Th2 type response, leading to the hallmark features of high Th2 cytokine production, eosinophilia, mastocytosis, goblet cell hyperplasia and high levels of IgE antibodies. Despite the successful generation of the Th2 response, worms are often able to survive in the host for long periods of time, causing chronic infections. This long-term survival within an immunocompetent host is facilitated by the induction of immunoregulatory mechanisms, including regulatory T cells (T reg), capable of secreting immunosuppressive cytokines such as IL-10 and/or TGF-β, resulting in an anti-inflammatory environment. Even though this regulatory environment prevents the elimination of worms it also protects the host against excessive inflammation. In worm-infected mice, blocking the IL-10 receptor partly restores IFN-γ and IL-12 production in lamina propria mononuclear cells. Mice that cannot make IL-10 develop a chronic colitis driven by cytokines associated with Th1/Th17 responses (e.g., IFN-γ, IL-17, IL-12, IL-23), attesting to the importance of this cytokine for mucosal immune homeostasis.

Helminth parasites stimulate a similar type of immune response to that induced by allergens (Th2 type response), but they also induce strong regulatory responses and in recent years there has been considerable interest in the potential role of helminth infections in modulating/regulating allergy risk in helminth-endemic areas. The underlying mechanisms are suggested to involve the regulatory presence of parasite-induced IL-10.

Worm infection inhibits mucosal IL-23 and IL-17 secretion, suggesting that regulation of IL-17 is an important mechanism of colitis control. How helminths limit IL-17 (Th17) responses is not understood completely. However, blockade of IL-23 secretion and stimulation of IL-4 production at the mucosal surface plays a partial role.

Gram-negative bacteria frequently express lipopolysaccharides (LPS). LPS interacts with receptors of the innate immune system (Toll-Like Receptor 4) on host cells, driving production of protective proinflammatory molecules. The normal intestinal mucosa usually is relatively unresponsive to LPS, since the normal intestinal flora produces large amounts of this molecule and uncontrolled responses to LPS would be dangerous for the mucosa. H. polygyrus infection induces display of TLR4 on a subset of mucosal T cells. Engagement of this receptor promotes production of regulatory (TGF-β, IL-10) rather than proinflammatory molecules. Thus, under the influence of helminths, damage to the mucosa allowing LPS penetration may induce activation of IL-10/TGF-β producing regulatory T cells, helping to quell excessive adaptive immunity.

Trials

Despite the potential dangers of some helminths, there appears to be a connection between the decrease in parasitic infections and the rise of autoimmunity. This is especially true considering the so-called 'Hygiene Hypothesis' (Helminths and the IBD hygiene hypothesis, 2008), according to which abnormal high levels of sanitation in the environment of the developed world may contribute to disordered immunoregulation in some autoimmune disorders (Trichuris suis ova: testing a helminth-based therapy as an extension of the hygiene hypothesis, 2012)

There is expanding epidemiological data indicating a protective role for helminth infections in various immunological diseases. A case-control study in Ethiopia showed that people infected with hookworm have a low frequency of asthma; this observation was later supported by a similar study conducted in Vietnam.

Multiple sclerosis is another disease displaying a North-South gradient with an increasing prevalence in developed countries. This rise in multiple sclerosis correlates with diminished carriage of T. trichiura (whipworm) in various locales. Multiple sclerosis patients who developed helminthic infection during the course of their disease displayed fewer disease exacerbations and developed fewer new brain lesions, as shown by magnetic resonance imaging, compared to the uninfected multiple sclerosis control group.

It was not until the mid-1990s that investigators seriously considered the possibility that live parasitic worms might be intentionally administered to humans in an attempt to treat autoimmunity. In recent years there has also been considerable interest in the potential role of helminth infections in modulating/ regulating allergy risk in helminth-endemic areas, due to the fact that, even though helminth parasites stimulate a similar type of immune response to that induced by allergens (Th2 type response), they also induce strong regulatory responses. On this basis, the possibility of treating some disorders with live helminths or helminth products has been explored in animal models, natural human infections and phase 1 clinical trials.

In general, the effects of helminths on autoimmune diseases are consistent, showing that these parasites can protect a host from developing auto-immune disease and/or can relieve symptoms ofestablished autoimmune inflammation.

Two conclusions can be drawn from the studies performed with TSO. One is that TSO is safe, and the other is that the efficacy of TSO should be tested in diseases in which there is an unmet need, such as food allergy and autism, or when available therapies have significant side effects, such as IBDs. The infrequent adverse events of TSO allow to performed clinical trials in children. This is obvious for food allergies because the effect of the disease is most severe in children. In addition, it is likely that TSO will be more efficient in young children with a maturing immune system than in adults.

Trichuris suis

Trichuris suis is a helminth of the nematode family. When pigs, the preferred host, ingest embryonated eggs, larvae are released in the intestine and colonize the superficial portion of the cecal and colonic mucosa. No systemic invasion occurs, as the worms mature in the intestinal lumen. In the pig T. suis’ infection is mainly asymptomatic, except in piglets, in which it can cause diarrhea and prevent growth.

Although human subjects are not the natural host for T. suis, it can survive for a few months in people without causing illness.

Trichuris suis ova (TSO) were orally administered to patients with inflammatory bowel disease (Helminths and our immune system: friend or foe?, 2009). In all studies no adverse events were reported.

Evidence of safety is also available from a study performed in patients with multiple sclerosis. One concern about the use of TSO as a therapy in humans is the possibility that egg preparations could contain other infectious agents often present in pigs, which are pathogenic in humans, such as hepatitis E virus. The TSO preparations used in all the recent clinical trials were systematically tested for multiple infectious agents according to procedures and processes approved by the FDA. This guarantees their microbiological safety. Another element contributing to the overall safety of using TSO as a therapy is the availability of anthelmintic drugs, such as albendazole or mebendazole, which are active against T. suis and are generally considered safe, including in children. These drugs could be used if a subject receiving TSO were to experience a TSO-related adverse event considered severe enough to require specific treatment.Overall, the evidence of safety of TSO is strong. The side effects have been absent o mild and spontaneously resolving.

Because of the safety of T. Suis, Trichuris Suis Ova (TSO) were chosen as the therapeutic agent to start experimenting with. Pigs were infected with T. suis eggs, then adult worms were isolated from the infected animals and cultured in vitro. The worms survived long enough to produce eggs that could be harvested and cleaned for clinical use.

Experiments in animals were eventually followed by pilot safety trials in humans in inflammatory bowel disease and allergic rhinitis. Subsequently, observational studies and exploratory clinical trials have been extended to multiple sclerosis.

In murine models of autoimmune diseases, the presence of helminth infections has been shown to have a protective effect. In addition, helminths were found to be protective in several models of inflammatory bowel disease (IBD).

Inflammatory Bowel Disease (IBD)

The trial with a patient suffering from IBD began with a dose of 2,500 eggs. One investigator in an earlier study had given themselves a similar dose to prove that this organism could colonize the human intestine, and reported no clinical symptoms. After six weeks, which is the time it takes for T. suis eggs to mature into adult worms, he reported no adverse events and showed improvement in disease symptoms that lasted for several months.

In another trial, three more patients with Crohn's disease and another three with ulcerative colitis all reported substantial improvements (or complete remission), with no side effects. Eventually, live eggs were administered every two weeks for 24 weeks to 29 patients with Crohn's disease. By week 24, nearly 80% of them reported a decrease in symptoms, and 72% were in remission - more than one would expect from a placebo effect. None reported side effects.

In a third trial of 54 patients with ulcerative colitis, about half of whom were given placebo, 43% of helminth-treated patients improved after 12 weeks, compared with only 17% of those given a placebo.

Several pharmaceutical companies have since taken on the task of developing T. suis eggs as a drug. Both the US Food and Drug Administration and the European Medicines Agency have formally approved the manufacturing process and allowed further testing.

Overall, there is substantial preliminary evidence of efficacy for TSO in patients with IBDs.

Multiple Sclerosis (MS)

If, according to the hygiene hypothesis, microbial deprivation causes abnormal immunoregulation and if, as demonstrated, helminths are able to promote normal immunoregulation, the question naturally arises as to whether helminth replacement may be therapeutic in MS and related conditions. Although the mechanisms of helminth therapy may overlap to some extent with the mechanisms of therapeutic oral tolerance, there are different reasons to employ the two treatments. Oral tolerance typically refers to the administration of a known, specific antigen or substance, generally in an attempt to alter immune responses to the specific antigen(s). By contrast, during treatment with antigenically complex live helminths (or molecules derived from them) in a disease such as MS, in which the pathogenic antigen(s) is unknown, the aim is to produce generalised immunomodulation, i.e. to alter responsiveness to many antigens or determinants (Helminth therapy and multiple sclerosis, 2013).

In 2007, Correale and Farez reported on a remarkable series of observations which illuminated the relationship between parasitic infections and MS (Correale and Farez, 2007). This prospective investigation compared two demographically-matched cohorts of relapsing-remitting MS subjects. The first cohort consisted of 12 control MS subjects without parasitism, and the second cohort consisted of 12 MS subjects identified at the onset of asymptomatic gastrointestinal parasitism contracted naturally.

During 4.6 years of follow up, MS activity virtually ceased in the infected cohort; the reductions in clinical attacks and new MS lesions relative to the control group, which was not infected with helminths, were greater than 95% on both measures. While neurological disability continued to worsen in the uninfected control cohort, in the stable, helminth-infected cohort there was essentially no worsening in functional status.

The first phase 1 clinical trial of helminth therapy in MS was the HINT 1 study (Helminth-induced immunomodulation therapy).

In the HINT 1 study, five newly-diagnosed, treatment-naïve MS subjects underwent brain MRI investigations at presentation before treatment (baseline phase), during each of the following 3 months, in which they were given 2,500 live TSO orally every 2 weeks (treatment phase), and 2 months after the end of TSO treatment (post-treatment phase). For the group of five subjects in HINT 1, before TSO treatment the mean number of new active brain MS lesions was 6.6 at baseline; this value fell to a mean of 2.0 after 3 months of TSO administration; and it rose again to a mean of 5.8 at 2 months after TSO was stopped.

Although the MRI results in this exploratory study are promising, particularly as they speak against an early, major anti-therapeutic effect of TSO in MS, they should be interpreted with caution, given the small number of subjects and the short period of observation. The favourable MRI results could be due to chance or regression to the mean; on the other hand, the “rebound” or return to the level of baseline activities after TSO administration was stopped suggests that a significant biological effect may be operative.

To date, all phase 1 studies of helminth therapy in MS have shown excellent safety and some, but not all, have been encouraging with regard to effects on clinical outcomes, brain MRI measures and immune status. Nevertheless, it is essential to recall that the data from exploratory studies to date are very preliminary.

It is not known how the helminths ingested may influence the central nervous system (CNS) in MS. An interesting observation was made by van der Valk and colleagues, who have provided compelling evidence that the first or earliest lesion of MS – before T cell invasion, breakdown of the blood-brain barrier and demyelination – is a small cluster of activated microglia, which they have called the pre-active lesion (PAL) of MS (van Noort et al., 2011 and Kipp et al., 2012). Furthermore, these investigators have shown that PAL are numerous and that their fate is variable; over a period of months some PAL will progress to classic plaques of inflammation and tissue damage, while others apparently will remain stable or even regress (van Horssen et al., 2012). These findings, if confirmed, would be of enormous importance, since they indicate that control of PAL fate may be the key to control of MS itself. Potentially, PAL are an attractive target for helminth immunomodulation mediated through the gut-systemic-CNS axis, e.g., by means of helminth-induced anti-inflammatory cytokines, other molecules, Tregs or additional cell types (Banks and Erickson, 2010) which may provide crucial modulating signals to microglial cells. Quite possibly, if immunomodulation by helminth or other agents favourably modifies CNS microglial PAL, MS itself will become quiescent.

Case study in autism

The story of a boy with autism has raised much interest among parents of autistic children or children with food allergy. This boy had severe autism characterized by self-abuse, agitation, aggression, anxiety, obsessive/compulsive behavior, behavioral rigidity, impulsivity, ‘‘stimming’’ behavior, and extreme sensitivity to external stimuli. He was also allergic to pecan nuts and presented with seasonal allergic rhinitis. At the age of 15 years, he was started on TSO at a dose of 1000 ova every 3 weeks for 26 weeks. No clear effect was observed. The dose was increased to 2500 ova every 2 weeks. After 10 weeks at the higherdose, most of his autism symptoms had improved substantially. His seasonal and food allergies were gone, and he was able to eat pecan cookies without having any reaction. After 2 years, the dose of TSO was reduced to 1600 ova every 2 weeks, and the autism symptoms reappeared. The dose was increased back to 2500 ova every 2 weeks, and the autism symptom immproved again. This case suggests a role of autoimmunity in autism onset and it suggests a possible way to relieve autistic symptoms.

Peanut/tree nut allergy

A study was performed in 6 adults aged 26 to 59 years with peanut or tree nut allergy. Peanut or tree nut allergy was chosen because it is the most common food allergy in the United States. After the treatment, no significant change in allergen-specific serum IgE levels were observed. There was no change in skin prick test reactivity, except in one subject who had a general decrease in reactivity and lost reactivity to peanut, which was the clinically dominant allergen for this subject. Four subjects reported a decrease in seasonal allergies while receiving TSO.

Helminths and vaccination

Many modern vaccines are designed to induce a Th1 response for optimal efficiency. Since helminth infections in general induce either a Th2 or a regulatory response, or both, there has been much discussionand speculation on whether concurrent helminth infections are responsible for reducing vaccine efficacy.

Clear evidence exists from murine models that helminth infections can alter the nature of immune responses to non-parasite or other antigens via the skewing of the cytokine response towards a Th2 response. Several human studies on cholera, BCG (Bacillus Calmette–Guérin) and tetanus vaccination report immune skewing during helminth infection. In general these studies suggest reduced Th1 responses to the vaccine, and impaired antibody titers, and with de-worming treatment significantly improving vaccine-specific responses. Maternal helminth infection may reduce the efficacy of BCG vaccination of newborns.

Helminths and Malaria

Much interest has focused on a potential link between helminth infections and other infectious diseases, in particular malaria. Since both helminth and malaria infections are prevalent throughout many areas of the tropics, co-infections with helminth and malaria parasites are frequently observed. The immune response to malaria, similarly to many other infectious diseases, needs to be carefully balanced so as not to result in an excessive inflammatory response that may harm the host, while retaining the ability to control parasite growth. This balance may be disrupted by helminth co-infections. Different studies report a wide variation of results. This depends on the fact that the intensity, and even the species, of the helminth infections are not always taken into account although it is increasingly clear that the outcome of co-infections are strongly influenced by the species and the intensity of helminth infection, and the age of the study population.

CONCLUSIONS

There is now little doubt that helminth infections really do have the ability to influence the development of immune responses. Both animal and human studies have demonstrated this repeatedly and convincingly.

However, what remains to be established is if this ability is of any real public health significance.

The stage of infection is likely to play a key role with respect to concurrent immune responses during a primary infection. Very little is known about the immune response during repeated helminth infections and even less of how such responses may affect concurrent immune responses.

The wide, and sometimes contradictory, range of results reported from human as well as animal studies clearly suggest that the impact of helminth co-infections on the immune response is complicated and there is little knowledge regarding the interactions between these parasites and the immune system.The wide range of results reported from human co-infection studies probably depend on basic differences in study design, choice of statistics and differences in statistical power, the choice of drugs and intervention strategies as well as the type of assays and other immunological investigations.

But it is also clear from animal studies that helminth species, timing and intensity of infections play major roles in the outcome of co-infections, and these factors have only recently begun to be considered in human studies.