Use of Rituximab to Attempt Recapture of Immune Tolerance to Pegloticase

Abstract

Patients suffering from erosive tophaceous gout who have failed oral urate-lowering therapies and pegloticase have few therapeutic options. Pegloticase failure (infusion reaction or lack of urate lowering) is primarily due to the development of anti-drug antibodies, and once present, retreating with pegloticase is mostly unsuccessful. We postulated that rituximab pre-treatment might permit recapture of pegloticase immune tolerance. We conducted an open label, safety and feasibility study to test this hypothesis. Patients with tophaceous gout and prior pegloticase failure were recruited to receive rituximab 1000 mg (twice), methotrexate 15 mg by mouth for at least 6 weeks, and then pegloticase per standard protocol. Patients were observed for infusion reaction and serum urate lowering in response to pegloticase infusions. Two patients completed induction and received at least 1 dose of pegloticase. Patient 1 had a moderate infusion reaction, requiring treatment with oral prednisone. Patient 2 had failure to lower urate treatment after 2 infusions. Based on these 2 outcomes, the trial was stopped. With either 6- or 12-week pre-pegloticase conditioning with rituximab, we were unable to recapture immune tolerance. Future trials considering use of rituximab might consider measuring anti-uricase antibodies in real time to guide the reintroduction of pegloticase.

1. Introduction

Patients with gout who fail standard oral urate-lowering therapies (ULTs) are at risk for developing chronic gouty arthritis and painful or unstable tophaceous deposits. These tophi lead to poor quality of life [1]. Successful treatment with uricase agents can be life-changing [2]. Pegloticase (PEG) is FDA-approved uricase for the management of gout, refractory to conventional therapy [3].

However, up to 62% of patients fail uricase therapy as manifested by failure to lower serum urate (SU) levels or infusion reactions, which are attributed to the development of anti-drug antibodies (either anti-uricase or anti-pegylated antibodies) (ADAs) [4,5,6]. Co-administration methotrexate (MTX) [4] or another immunomodulator [7] is now standard of care for those initiating PEG and, as reported in the MIRROR study, reduces the PEG failure rate to 29% [4].

Unfortunately, for patients who previously failed PEG, recapturing immune tolerance (for example with MTX pre-treatment) has had poor success. In a recent open-label trial of 11 patients with prior PEG failure, pre-treatment with MTX (25 mg/week, subcutaneous) was successful for only 1 patient (89% failure rate). Eight patients did not capture or lost SU response (prior to 6 months), and two patients experienced infusion reactions [8].

Rituximab (RTX) has been used to reduce antibody-mediated rejection after kidney transplant [9,10,11] and to desensitize patients prior to renal transplant [12,13]. Therefore, we postulated that for patients with prior PEG failure (and presumed ADA), pre-treatment with RTX (and MTX) would lower ADA sufficiently to recapture immune tolerance to PEG. To test this hypothesis, we obtained an FDA IND exemption for a proposed safety and feasibility open-label trial to use RTX pretreatment, followed by standard of care (SOC) methotrexate–pegloticase (MTX-PEG) to treat poorly controlled tophaceous gout in individuals with prior loss of response to PEG. We registered this protocol with ClinicalTrials.gov (NCT06186219). UCLA IRB (23-0270) approved the protocol. UCLA Health donors provided funding for this trial.

2. Materials and Methods

2.1. Eligible Patients

Patients with prior PEG failure (either loss of SU control or mild-to-moderate prior PEG-related infusion reactions) were recruited form the principal investigator’s university clinical practice. Patients with prior anaphylaxis or contraindication to RTX or MTX were excluded from participation.

2.2. Inclusion and Exclusion Criteria

After reviewing the study protocol, procedures, risks, and benefits, interested subjects provided informed consent to participate in this study. Standard safety labs for RTX, MTX and PEG were obtained. Patients with Chronic Kidney Disease stage 4 or worse, non-compensated congestive heart failure (within the preceding 3 months), unstable angina, uncontrolled hypertension, on treatment for non-skin cell cancer, on treatment for bacterial infection or severe chronic recurrent infections were excluded from study recruitment. Patients with organ transplant, or on immune-based organ transplant medications, history of Hepatitis B, Hepatitis C, or HIV positivity were excluded. Pregnant, planning conception, or breastfeeding patients were excluded. Also excluded were patients with PEG or RTX in the preceding 6 months, prior severe PEG infusion reactions, or with intolerance or contraindication to MTX or steroids. For a detailed list of inclusion and exclusion criteria, please refer to ClinicalTrials.gov (NCT06186219).

2.3. Treatment Protocol

Our treatment protocol is summarized below (

Table 1. Scheduled Study medications (shaded) and as administered (not shaded).

		Wk −12	Wk −6	Wk 0	Pegloticase 8 mg IV q 2 Weeks				Wk 26
Rituximab 1000 mg IV, twice	Pt 1		↓↓	↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓
	Pt 2	↓↓		↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓ ↓
Methotrexate 15 mg po weekly	Pt 1		MTX
	Pt 2

). After informed consent and a screening exam, we started patients on MTX 15 mg by mouth weekly and RTX 1000 mg given twice, 2 weeks apart at least 6 weeks prior to the first planned PEG. (The protocol was modified to extend the RTX pre-treatment window to 12 weeks after Patient 1 had a moderate PEG infusion reaction.) Standard RTX and PEG pre-infusion medications were given as recommended on respective FDA package inserts [14,15].

2.4. Study Endpoints

Primary safety endpoints (and discontinuation) were either the occurrence of moderate to severe infusion reaction or failure to suppress SU < 6 mg/dL twice (prior to subsequent scheduled PEG infusions). Clinical endpoints were maintenance of SU < 6 mg/dL through week 26, reduction in largest tophus size or count of tophi.

3. Results

We enrolled two sequential patients before stopping the RTX-MTX-PEG trial due to successive treatment failures. Both patients met ACR/EULAR classification criteria for gout [16], and had severe, debilitating tophaceous gout. (Refer to

Table 2. Patients’ pegloticase pre-study history.

	2013	2014	2015	2016–2020	2021	2022	2023–2024	2025
Patient 1
PEG	↓ ↓ *
MTX-PEG					↓ ↓ †
RTX-MTX-PEG								↓ *, †
Patient 2
PEG			↓ ↓ *, †
MMF-PEG						↓… ↓ ↓ *, †
RTX-MTX-PEG								↓ ↓ †

MTX = methotrexate, MMF = mycophenolate mofetil, PEG = pegloticase. * Infusion reaction. ^† No urate-lowering benefit.

for detailed histories.) Both patients had previously failed PEG monotherapy, as well as rechallenge with either MTX-PEG or mycophenolate (MMF)-PEG.

Patient 1 had failed all available xanthine oxidase inhibitors (XOI) and uricosurics. He suffered with chronic gouty tophaceous arthritis, frequent polyarticular flares requiring oral prednisone or corticosteroid injections and several chronic open draining tophaceous lesions. He had prior surgical debulking of all four extremities. He was first treated with PEG (monotherapy) in 2013, tolerating the first infusion, but then developing chest tightness and sweating 15 min into the second infusion. There was no urate-lowering response. Patient 1 was rechallenged with MTX-PEG in 2021, again tolerating the first infusion, but without a urate-lowering response; the MTX-PEG rechallenge was discontinued. (See

Table 3. Patient characteristics.

	Patient 1	Patient 2
Age (years)	53 (current), 18 (age at onset)	43, (current), 20 (age at onset)
Prior Urate-Lowering Therapies	Allopurinol (GI intolerant at doses > 100 mg and lack of efficacy) Febuxostat (GI intolerant to 40 mg) Probenecid (lack of efficacy) Lesinurad (lack of efficacy)	Allopurinol up to 600 mg (ineffective SU response, lowest SU 6.8 mg/dL) Allopurinol/probenecid ineffective (lowest SU 9.3) Febuxostat 120 mg (ineffective SU response) Febuxostat/Lesinurad (ineffective and cost prohibitive)
Anti-inflammatory therapy	Prednisone 10 mg daily Anakinra 100 mg daily Colchicine 0.6 mg, twice daily	Prednisone 10 mg (up to 20 mg) Anakinra 100 mg daily Colchicine 0.6 mg, twice daily
Pre-treatment clinical images

SU = serum urate; GI = gastrointestinal.

for detail).

Patient 2 was similarly burdened with tophaceous deposits and failed all oral XOI and uricosurics. He was mostly non-ambulatory due to painful deposits in the soles of his feet. He had previous amputation of the distal phalanx of his right third finger and resection of large tophus from the left first metatarsal phalange region. Patient 2 was first treated with PEG (monotherapy) in 2015, also tolerating the first infusion, but then developing deep low back kidney pain. The infusion was slowed, and the patient was able to complete, but there was no urate-lowering benefit and the treatment was discontinued. The patient was rechallenged with MMF-PEG in 2022. He tolerated several doses with a marginal urate-lowering benefit, but he eventually developed a rash and lip swelling. The MMF-PEG rechallenge was discontinued for infusion reaction and incomplete urate-lowering benefits. (See Table 3 for detail).

Both patients tolerated the RTX 1000 mg infusions and 15 mg of oral MTX weekly. Both patients responded with suppression of their CD19 cells to 0 prior to the first scheduled PEG infusion. (See

Table 4. Patient treatment and response.

Patient 1	Date	2 May 2024 16 May 2024	13 June 2024 (Week 6)	27 June 2024
		RTX 1000 mg ×2	PEG #1, 8 mg
		Methotrexate 15 mg weekly
	Urate (mg/dL)	9.8	9.0	9.9
	CD19	148	0
Patient 2	Date	9 August 2024 30 August 2024		8 November 2024 (Week 12)	22 November 2024	5 December 2024
		RTX 1000 mg ×2		PEG #1, 8 mg	PEG #2, 8 mg
		Methotrexate 15 mg weekly
	Urate (mg/dL)	11.0		11.1	7.5	9.8
	CD19	Not Administered		0

Patient 1 experienced infusion reaction after first infusion resulting in discontinuation from the trial. Patient 2 had 2 SU levels > 6 mg/dL resulting in discontinuation from the trial. Owing to these 2 failures, we terminated our RTX-MTX-PEG clinical trial.

).

Patient 1 reported nausea, a rapid heart rate, and low back pain during the first PEG infusion. Symptoms resolved after pausing the infusion, slowing the rate on restart, allowing for completion of the first infusion. Post-infusion, Patient 1 complained of fatigue and “flu-like” symptoms and presented for medical evaluation on post-infusion day 8 with fevers (37.9 C), orthostatic light-headedness, nausea, night sweats, and bloating. Patient 1 reported that these symptoms were similar to his prior infusion reactions. His course was further complicated by a flare of his right shoulder. Ultrasound showed a positive power Doppler signal, consistent with inflamed tophaceous deposit. All symptoms resolved with a seven days of prednisone 20 mg daily, tapered to 10 mg. Prior to his planned second PEG infusion, his urate level had increased to 9.9 mg/dL (from 9.0 mg/dL). This lab and events were reviewed by our allergy consultant and DSMB physician, and the decision was made to discontinue the patient from the trial for moderate infusion reaction and failure of urate response. With this initial failure, we modified the protocol for Patient 2 to administer first PEG infusion 12 weeks after the first dose of RTX.

Patient 2 similarly tolerated RTX pretreatment and MTX. However, 2 weeks prior to first PEG, the patient presented to the outside Emergency Room with increased swelling of the R third finger with concern for infected or inflamed tophus. He was treated with intravenous ceftriaxone (two doses) and underwent amputation of his right third finger. He was discharged off antibiotics. He had been continued on methotrexate, anakinra, and prednisone uninterrupted. The study team was not notified of this event until the patient presented for his first PEG infusion.

At presentation for his first PEG infusion, surgical wounds from his recent amputation were well healed. There was no sign of infection (off antibiotics), and inflammatory markers were normal (ESR 9, CRP 1.0 mg/dL). He reported missing his most recent dose of MTX due to GI intolerance. The decision was made to proceed with his first PEG infusion, which he tolerated well except for moderate worsening of his hypertension. His baseline urate decreased from 11.1 mg/dL to 7.5 mg/dL (obtained prior to his second scheduled PEG infusion). Unfortunately, after second infusion, his urate went back up to 9.8 mg/dL (prior to his third planned infusion). With 2 post-PEG SU > 6 mg/dL, the third PEG infusion was cancelled in accordance with the study protocol stopping rule.

4. Discussion

For patients with severe tophaceous gout who have previously failed PEG and all other prior ULT agents, there are no meaningful medical debulking therapeutics. While there is good rationale for the use of RTX to recapture immune tolerance to PEG, these two decisive failures did not support continued recruitment. Given the recently reported failed effort to recapture immune tolerance with MTX 25 mg subcutaneous [8] and our failed effort with RTX, the importance of preventing development of ADA with MTX or other established immunosuppressant to limit development of ADA should be a priority for all providers starting patients on PEG.

Analyses of ADA from clinical trials describe antibodies against the pegylated portion as well as the porcine uricase portion of the pegloticase drug with the anti-pegylated antibodies being stronger drug-neutralizing antibodies [17]. Patient 1 showed no evidence of recapturing immune tolerance (with either the 2021 MTX rechallenge or the 2025 MTX-RTX rechallenge). Patient 2 may have shown some partial immune tolerance recapture to MMF in 2022, where several infusions were tolerated with modest urate suppression before infusion reaction and urate failure, and failure to MTX-RTX challenge in this study. While a single case does not establish theory, it should be noted that MMF has been used to decrease circulating alloantibody levels in heart transplant recipients (greater than Azathioprine) [18] and has been the primary immune modulating treatment for patients with Systemic Lupus Erythematosus.

Clinicians should probably be cautious about considering calcineurin inhibitors (CNI) to recapture immune tolerance based on recent publications. CNI are effective immune modulators but also inhibit regulatory T cells and therefore limit induction or recapture of immune tolerance [19], although low-dose CNI may promote regulatory T cell development.

Despite our failure, the concept of RTX B cell depletion has merit to recapture immune tolerance through reduction in ADA titers. As noted above, RTX has been used as pre-conditioning for organ transplant. The known risks of RTX (HBV reactivation [20], blunted vaccination responses [21,22], hypogammaglobulinemia) further suggest potential efficacy of RTX on ADA suppression.

One option to consider going forward would be the real-time evaluation of ADA suppression post-RTX to determine optimal timing of PEG rechallenge. We did not have access to PEG-ADA assays. Higher ADA titers are more likely to be associated with PEG infusion reactions [6]. Although the recent multicenter MTX rechallenge study failed to achieve endpoints (only 1 of 11 subjects responded), they did report that the single responder had the highest baseline ADA that fell to “near zero” and had the lowest ADA titer of all subjects by week 10. The authors suggest that this demonstrated “immune tolerance to pegloticase, presumably due to MTX cotherapy. However, … this seems to be a rare occurrence.” [8] Given the treatment need for patients with prior PEG failure, a RTX induction study guided by ADA titer reduction prior to PEG rechallenge could have tremendous clinical impact.

While new uricase therapies are being evaluated (Clinical Trials NCT01021241 [23]), PEG failure remains an exclusion criteria. Other non-XOI urate-lowering interventions (e.g., losartan [24], Sodium-Glucose Transport Protein 2 (SGLT2) Inhibitors [25,26]) may help patients with refractory hyperuricemia but are unlikely to be sufficient in patients with severe tophaceous gout.

Our early failures (both prior to third infusion) were similar to the outcomes reported in the recent open-label multicenter MTX rechallenge study where 8 of the 10 cases failed within the first three infusions [8].

5. Conclusions

Despite these failures, the concept of recapturing immune tolerance remains intriguing. Ideally, a clinical trial designed to recapture PEG immune tolerance would include concurrent ADA evaluations to guide informed decisions about timing and whether to rechallenge with PEG. We did not have access to real-time ADA assays for treatment decisions and based on this small experience, caution against empiric use of RTX pretreatment that did not include concurrent ADA evaluation.