Melek Simsek

This study revealed that maintenance of corticosteroid-free clinical remission rates at 3, 6, and endpoint 12 months were 69%, 61%, and 45% respectively, and an 80% continuation rate until end of follow-up. Serious thioguanine related adverse events were observed in 13% and severe adverse events in 5% of patients. Myelo- and hepatotoxicity were both reported in 6%, infections in 10% and skin cancer and portal hypertension (diagnosed by ultrasound but without clinical or biochemical abnormalities) each in one patient. We concluded that thioguanine exhibits comparable effectiveness to conventional thiopurines, while offering a more favorable tolerability profile, as evidenced by a drug survival rate of 80%, consistent with previous studies involving thioguanine. These findings played a crucial role in the formal approval and licensing of thioguanine for IBD in the Netherlands as of 2022.

Patient-reported outcomes and experiences (PROMS and PREMS) play a crucial role in the evaluation of treatments, including thioguanine therapy for IBD. In Chapter 5 we describe our self-report survey among thioguanine treated patients with IBD. Questionnaires were distributed to members of the Dutch National Crohn’s and Colitis patient organization, employing the Treatment Satisfaction with Medicines Questionnaire (SATMED-Q) to capture insights into their satisfaction levels and the impact of thioguanine therapy on their disease and daily lives. A total of 173 organization members (73% female) participated in the survey and among them, 74% expressed satisfaction with the effectiveness of thioguanine, while 5% were dissatisfied. Moreover, 80% reported being content with the quality of care they received. In terms of the impact on daily life, 54% had a positive impact (good or excellent), while 40% a neutral perception, and 6% reported a negative impact. Regarding disease activity, 58% reported improvement, while it remained stable in 39% and worsened in 3%. In the evaluation of new or rediscovered therapies, patient-reported outcomes and experiences should be considered as a key instrument.

Part III of this thesis was dedicated to the safety of thioguanine, especially concerning hepatotoxicity and pregnancy. IBD frequently affects young adults during their reproductive years. As active disease has been linked to an increased risk of adverse perinatal outcomes, it is imperative to prioritize the maintenance of remission through appropriate therapies during conception and pregnancy. Both azathioprine and mercaptopurine are considered as safe options during conception and pregnancy. Regarding thioguanine, the available safety data are relatively limited. We gathered data from 117 thioguanine exposed pregnancies in 99 women, as described in Chapter 6. The majority (78%) had Crohn’s disease, and the mean age at delivery was 31 years. During these pregnancies, 18 cases (15%) experienced a flare-up of IBD. Ten pregnancies (8%) resulted in a first-trimester miscarriage, and one case ended in a stillbirth at 22 weeks gestational age. A total of 109 neonates were born from 101 singleton pregnancies and four twin pregnancies. Among the singleton pregnancies, 10 children were born prematurely, and 10 were classified as small for gestational age. Obstetric complications were observed in 15% of pregnancies and infectious complications were noted in 7%. Congenital abnormalities were identified in one induced abortion (trisomy 21) and one live-born child (cleft palate). Myelosuppression screening was conducted in sixteen neonates (15%), and two of them exhibited anemia in umbilical cord blood. Our thioguanine-related pregnancy outcomes were found to be comparable to the data involving the use of conventional thiopurines in pregnant IBD patients. Therefore, we believe that our data support that thioguanine may be safely used throughout the pregnancy in IBD patients.

The significance of identifying histopathological nodular regenerative hyperplasia (NRH), and monitoring its progression in asymptomatic patients, has been a matter of debate. As histopathological evaluation of the liver was standard practice during thioguanine therapy in IBD, the European TG working party no longer recommends routine liver ultrasound or biopsies during therapy, except in case of signs of liver disease. Still, the progression of NRH over time, especially when the causative agent has been eliminated, remains unknown. In Chapter 7, we delve into the clinical course of nodular regenerative hyperplasia (NRH) whereas Chapter 8 delves into the pathological evolution of NRH.

In order to investigate the long-term clinical course of NRH, we reached out to almost all hospitals in the Netherlands for data. A total of 72 hospitals reported 43 cases of NRH in thiopurine treated patients with IBD. Among these, 18 patients (42%) were exposed to thioguanine, and 90% of them previously received azathioprine and/or mercaptopurine. The cohort was characterized by a predominance of male sex (77%) and stricturing IBD disease phenotype (47%). At the time of NRH diagnosis, 17 patients (40%) were asymptomatic, while 26 patients (60%) presented with symptoms consistent with non-cirrhotic portal hypertension (NCPH). These symptoms included splenomegaly in 23 (88%), ascites in 4 (15%), and esophageal varices in 19 (73%), with variceal bleeding occurring in 11 patients (42%). Following a median follow-up period of 5.5 years (range: 2-13 years), we observed that liver enzymes and platelet counts improved in the entire study cohort. Also signs and symptoms of NCPH were observed to have resolved in 8 patients (31%), including improvement of thrombocytopenia and splenomegaly in all 8, and esophageal varices as well in 6 of them. Remarkably, the 17 patients who initially presented without clinical indications of liver disease at the time of diagnosis did not experience the onset of complications throughout the course of follow-up. None of the patients necessitated a liver transplantation, developed hepatocellular carcinoma or deceased during the extensive long-term follow-up.

The potential for reversibility of NCPH in NRH patients was also observed in our histopathological investigation, as described in Chapter 8 of this thesis. Within a cohort of 25 thioguanine using IBD patients (both 60% female and Crohn’s disease, mean age 44 years), sequential liver biopsies prior to, at time of, or after cessation of thioguanine treatment were performed with a median time between first and second biopsy of 25 months (range: 14 – 54 mo). In 19 patients (76%), both first and follow-up liver biopsies were performed at time of thioguanine treatment. In three patients (12%), the first liver specimen was obtained prior to thioguanine, and the follow-up specimen during thioguanine treatment. Another three patients (12%) underwent a first biopsy at time of thioguanine treatment and a following biopsy after thioguanine withdrawal.

In all but one case, the liver specimens exhibited various degrees of irregularities, encompassing inflammation, steatosis, fibrosis, and some vascular disturbances. Notably, the prevalence rates of perisinusoidal fibrosis (91%), sinusoidal dilatation (68%), and nodularity (18%) remained consistent between the first and second liver biopsies. A trend toward statistical significance was observed for phlebosclerosis, with 36% prevalence in the first biopsies and 68% in the second biopsies (p = 0.092). Importantly, the presence of histopathological liver abnormalities did not demonstrate a significant association with clinical outcomes. In two patients (out of 3) nodularity of the liver resolved upon thioguanine withdrawal. We concluded that vascular abnormalities of the liver were commonly observed in thioguanine treated IBD patients, although these did not exhibit a progressive course over time and retained limited clinical relevance.

Future perspectives
As research in the field of IBD continues to evolve, the potential role of thioguanine in IBD management presents several intriguing future perspectives. In our effectiveness studies, we have demonstrated that nearly half of the IBD patients maintain clinical remission with thioguanine over a 12-month duration, a finding of notable clinical significance (Chapter 3 and 4). Additionally, patient tolerance rate and patient satisfaction with thioguanine therapy have both been reported in 80% of patients who had experienced prior treatment failure with azathioprine or mercaptopurine (Chapter 3, 4 and 5).

Although current clinical practice in the Netherlands incorporates thioguanine as a viable treatment option for IBD, this is not yet standard in many other countries. Thioguanine, administered in oral tablets, is user-friendly and may be perceived as a cost-conscious therapy, particularly when compared to various biologic treatments, making it a potentially advantageous choice, especially in regions with constrained healthcare resources.

Prolonged safety
Given that thioguanine originated in the 1950s, its long-term safety profile has been well-documented. Additionally, the repositioning of thioguanine for IBD in the early 2000s prompted additional safety trials. We have seen that NRH, as initially reported in early IBD trials in patients with relatively high doses of thioguanine, occurs rarely when using the current, adequate dosages (0.3 mg/kg, not exceeding 25 mg/day) (Chapter 3, 4, 7 and 8). Our comprehensive long-term safety data suggests that the safety profile of thioguanine is quite comparable to azathioprine and mercaptopurine, with even fewer reported incidents of hepatotoxicity, myelotoxicity, and idiosyncratic adverse events, including pancreatitis (Chapter 3 and 4). There is no apparent increase in the risk of infections, malignancy, or adverse pregnancy outcomes (Chapter 6). Future research should focus on monitoring patients over extended periods to assess potential late-onset adverse effects to validate current findings.

Tailored treatment
Future advancements in pharmacogenomics and prediction tools may allow for more precise selection of patients who are likely to benefit from thiopurine therapy, including thioguanine. Tailoring treatment based on an individual’s genetic profile and drug metabolism can potentially enhance its effectiveness while minimizing side effects. Regarding therapeutic drug monitoring (TDM) of thioguanine, it is not yet common practice to measure concentrations of active metabolites, such as 6-TGN, as is routinely done with azathioprine or mercaptopurine. Nevertheless, one of our studies demonstrated that 6-TGN concentrations exceeding 682 pmol/8×10^8 RBC were associated with effectiveness (Chapter 4). In the future, a rounded cut-off value of 700 pmol/8×10^8 RBC may be established for TDM of thioguanine, but further research is necessary to substantiate this.

Co-therapy
The exploration of combination therapies involving thioguanine may be a promising avenue. Immunomodulators like azathioprine, mercaptopurine and methotrexate are currently used alongside biologics to reduce immunogenicity and enhance efficacy. Although not yet extensively studied, thioguanine may offer similar benefits with potentially fewer anticipated side effects. Within our extensive cohort of 274 IBD patients treated with thioguanine, 21 individuals (7%) used combination therapy with anti-tumor necrosis factor-alpha (anti-TNF) (Chapter 3). Although this study did not reveal significant changes in clinical effectiveness or toxicity in this subgroup, its size limited our ability to draw firm conclusions. Presently, studies are underway to further investigate this approach of combination therapy with thioguanine and biologics.

Targeted drug delivery
Moreover, advancements in the drug delivery system for thioguanine hold the potential to achieve more targeted and controlled release, thereby reducing undesirable side effects and optimizing its therapeutic benefits. Recent ‘in vitro’ studies have demonstrated that the gut microbiome possesses the capability to metabolize thioguanine into active metabolites, the 6-TGN, which was not the case for mercaptopurine. Additionally, investigations have revealed that when applied rectally, thioguanine exerts local effects and effectively treats distal colitis in Winnie mice. In a recent case series, we presented our findings regarding thioguanine enemas and suppositories in 16 patients suffering from distal ulcerative colitis. Preliminary results are indeed promising; nevertheless, further research is imperative to validate and substantiate these outcomes.

Expanding treatment indications
The therapeutic applications of thioguanine extend beyond IBD. While not as a first choice, thioguanine still holds a place in the management of leukemia (acute lymphoblastic leukemia, acute myeloid leukemia and chronic myeloid leukemia). In the fields of dermatology and rheumatology, thioguanine is used off-label to treat moderate to severe psoriasis and other difficult-to-treat dermatitis and vasculitis conditions. In the field of gastroenterology and hepatology, thioguanine has shown promising results, at various degrees, in the management of autoimmune hepatitis, collagenous sprue, refractory coeliac disease, in addition to its applications in IBD. Additional research and clinical experience are required to comprehensively investigate the potential benefits of thioguanine in these, and perhaps potentially in other inflammatory and immune-mediated diseases.

Drug rediscovery
Finally, the rediscovery of thioguanine for IBD treatment was not without its challenges. Drug rediscovery, in general, is a valuable pursuit as it expands therapeutic options and reduces drug development-associated costs. However, the absence of a formal regulatory approach and limited economic interest from pharmaceutical companies pose significant challenges. Thiosix®, the presently conditionally registered thioguanine compound, received official approval for IBD treatment in the Netherlands as of early 2022.

We believe that regulatory reform and market regulation in Europe are necessary to ensure the full utilization of every generic drug and to prevent excessive pricing by pharmaceutical companies.

CONCLUSION

Thioguanine, initially originated as chemotherapeutic drug in 1950’s, can be described as a hidden treasure among older pharmaceutical agents. Currently, thioguanine emerges as a promising therapeutic option for treating IBD, as evidenced by a notable patient tolerance rate of 80% and clinical effectiveness observed in almost half of the patients for over a year. Extended safety studies have further exposed that hepatotoxicity, and especially occurrence of clinical relevant NRH, occurs rarely during thioguanine application at 20 mg per day for IBD.

Despite the challenges encountered during the rediscovery of thioguanine for IBD, including issues related to off-label prescribing and concerns about hepatotoxicity, it has now become a formally registered medication for IBD in the Netherlands (as of early 2022). This repositioning of thioguanine for IBD emphasizes the value of exploring existing therapeutic agents, which may hold untapped potential for the benefit of patients.