ORIGIN-1 trial: study protocol for an organoid-guided N-of-1 trial evaluating CFTR modulator response in cystic fibrosis with rare non-F508del mutations

Abstract

Background

Cystic fibrosis (CF) is a genetic condition caused by mutation of the cystic fibrosis transmembrane regulator (CFTR) gene. Recently licensed modulator therapies target the defective CFTR protein and have transformed the formerly life-limiting trajectory of people with CF (pwCF). elexacaftor/tezacaftor/ivacaftor (ETI) has shown outstanding clinical efficacy in pwCF homozygous for F508del (approximately 50%), and those with a F508del mutation paired with any of the > 2000 other mutations identified in the CFTR gene (heterozygous pairing). However, clinical response to modulators is variable and people with rare CF mutations are often excluded from modulator access worldwide. The ORIGIN-1 trial aims to determine whether an organoid-guided n-of-1 trial can predict and demonstrate clinical response to ETI in pwCF with rare mutations.

Methods

Based on the in vitro testing, participants will be eligible to enter the interventional phase of the trial. Participants will be administered blinded study drug (i.e. ETI or placebo) for a 14-day treatment block followed by a 14-day washout period prior to beginning the alternate treatment, again for a 14-day period. A treatment cycle will be defined as one ETI treatment block, one placebo treatment block and the intervening washout period. Treatment blocks sequence will be randomly assigned. Participants will complete a minimum of two treatment cycles and up to four consecutive cycles depending on the results of planned interim analyses.

Discussion

The ORIGIN-1 trial is designed to demonstrate a precision medicine approach that targets CFTR modulator therapy to ensure the optimal use of these very high-cost therapies. This pathway may expand the application of CFTR modulators to include pwCF with rare mutations of the CFTR gene for whom CFTR modulators’ efficacy has not been (and are unlikely to ever be) proven by conventional, parallel-group clinical trials. We anticipate that a personalised organoid model that measures patient-specific CFTR responses to modulators in vitro will enable the reliable identification of pwCF with these rare CFTR mutations who are likely to respond to existing modulator therapy.

Trial registration

Australia New Zealand Clinical Trials Registry (ANZCTR) ID: ACTRN12623001136695 Registered on November 03, 2023.

Supplementary Information

The online version contains supplementary material available at 10.1186/s13063-026-09789-y.

Administrative information

Note: the numbers in curly brackets in this protocol refer to SPIRIT checklist item numbers. The order of the items has been modified to group similar items (see http://www.equator-network.org/reporting-guidelines/spirit-2013-statement-defining-standard-protocol-items-for-clinical-trials/).

Title {1}	Organoid-guided N-of-1 (ORIGIN-1) trial: a phase 4 study to investigate whether people with cystic fibrosis with rare non-F508del CFTR mutations who have an invitro response to elexacaftor/tezacaftor/ivacaftor also have a clinically meaningful response to elexacaftor/tezacaftor/ivacaftor versus placebo.
Trial registration {2a and 2b}	Australia New Zealand Clinical Trials Registry (ANZCTR)- ID: ACTRN12623001136695 Registered on November 03, 2023. https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=386544&isReview=true
Protocol version {3}	Version 8.0 dated 08 Apr 2025
Funding {4}	Funding is provided by a Medical Research Future Fund grant (GNT2014916)
Author details {5a}	Dr Lorena Rodrigues Sabino A/Prof Gerard Kaiko Dr Douglas Dorahy Dr Thomas Goodsall Prof Adam Jaffe Prof Andre Schultz Dr Shafagh Waters Dr Dane Cook Dr Harvey Kok Dr Koliarne Tong Mr. James Totterdell Prof James McGree Prof Tom Snelling Prof Peter Wark
Name and contact information for the trial sponsor {5b}	The University of Newcastle Research grants office: +61 2 4921 7733
Role of sponsor {5c}	As Sponsor, The University of Newcastle assumes ultimate responsibility for initiation, management and financing. Design, conduct, monitoring and reporting are delegated to the Steering Committee.

Introduction

Background and rationale {6a}

Cystic fibrosis (CF) has a population incidence of ~1 in 3000 in Australia [1]. It is a multisystem disease that causes recurring chest infections with a progressive loss of lung function, serious gastrointestinal pathologies and damage to the pancreas, and liver [2]. These effects impose on people with CF (pwCF) a need for lifelong intensive and expensive specialist clinical care. Despite improvements to clinical care, CF is still associated with a high burden of chronic illness and a median life expectancy of approximately 40 years [1].

CF is caused by genetic mutations on chromosome 7 that affect the chloride and carbonate ion transport protein [3]. However, CF is not a simple genetic disease, and a simplified interpretation underestimates the substantial diversity in individual disease trajectory and therapy responsiveness. An individual may have a pair of mutations that derive from any combination of the > 2000 known genetic mutations in the cystic fibrosis transmembrane regulator (CFTR) gene that contribute to its dysfunction. In addition, the impact of genetic variation in CFTR modifier genes, epigenetics, as well as the influence of environmental factors, can contribute to the ultimate phenotype of the disease and, importantly, the responsiveness to therapies [4, 5].

In the last decade, small molecule CFTR modulators have been developed. These molecules specifically target the disease process, enhancing CFTR protein function and promise increased life expectancy [6]. Efficacy has been demonstrated in conventional randomised, blinded, placebo-controlled clinical trials, in which participant eligibility was based on specific genotypes. The first of these agents, ivacaftor, was shown to improve CFTR function in people with a class III/gating mutation found in approximately 5% of people with CF [7, 8]. Most recently, a triple combination agent, elexacaftor/tezacaftor/ivacaftor (ETI), demonstrated efficacy in pwCF homozygous for F508del (approximately half of pwCF), and in people with CF with one F508del mutation in combination with one of several mutations characterised by minimal function (MF) of the CFTR [9, 10].

Eligibility for these efficacy trials that compared response to placebo, targeted pwCF with the most common CF genotypes and mutations where the modulators were shown by high-throughput in vitro assay to be most sensitive to partial restoration of CFTR function. These trials demonstrated average group-level improvement in the surrogate markers of disease progression, lung function and exacerbations with outcomes measured for up to 24 weeks. The focus on average effects does not reflect the high degree of variation in individual’s clinical modulator response in the clinical trials [10]. This heterogeneity of response is true for pwCF sharing the homozygous F508del genotype and was substantially higher for genotypic pairings carrying one copy only of F508del [11, 12]. PwCF with F508del residual function mutations, gating mutations, and uncharacterised mutations were excluded from these studies [10]. There are more than 2000 mutations that can affect CFTR function and 800 of these have been definitively shown to cause disease [13]. Some mutations result in a complete failure to produce CFTR protein (Class I) and rescue therapy with a CFTR modulator would clearly be ineffective. For some class I mutations, clinical evidence of compromised function is empirically derived from direct testing but for others, functional interference is presumed from data derived from in silico studies. Thus, many mutations may respond to CFTR modulators that have not been clinically tested due to clinical trial design and regulatory restrictions rather than exclusion based on scientific evidence. For example, the mutation N1303K is the third most common mutation found worldwide after F508del and produces a dysfunctional CFTR protein which has reduced expression and gating function. In vitro evidence of an ETI response by N1303K suggested an improvement in CFTR function in human cell models, though not as pronounced as seen with F508del which led to this mutation being excluded from clinical trials [14]. Subsequent case series outcomes in Europe and the US demonstrated significant improvement in clinical outcomes in people who accessed ETI via uncontrolled compassionate access programmes [15]. Demonstrating reliable clinical responses using small case series is particularly challenging due to the very high cost of CFTR modulator therapies. Furthermore, establishing clinical benefit requires measurable outcomes, such as improvement in lung function. This creates a significant limitation for younger children, as they often need to develop irreversible disease before such changes can be detected and a trial of therapy can be justified.

Rationale for the present study: ORIGIN-1

While the efficacy of CFTR modulators has been established for most CFTR mutations, uncertainty remains for the rare but important minority mutations. An effective in vitro biomarker that can be used to demonstrate CFTR response to modulators and can predict clinical response is needed to address the inequity. A demonstration of modulator efficacy using a biomarker has been accepted by international regulatory agencies as a reasonable proxy for pwCF with mutations that have not been tested by clinical trial, though the magnitude of a biomarker and its relationship to clinical benefit and threshold of CFTR responsiveness remain a challenge. The most easily available biomarker of CFTR response is by measurement of sweat chloride. Changes in sweat chloride have been shown to correlate with lung function changes at a population level. However, these analyses do not reliably predict individual clinical response to modulators [16, 17]. While an untargeted approach to treatment for groups of patients with rare diseases may be acceptable for a lower-cost medicine with a low-risk safety profile, the on-market CFTR modulators are high-cost medicines with safety risks. Therefore, an untargeted approach is not reasonable. These therapies must utilise tailored precision medicine to ensure quality use of medicines (QUM) and patient safety. For pwCF with rare or unique CFTR gene mutations, a new strategy for evaluating treatments using personalised targeting and individualised clinical assessment could offer a more equitable, sustainable and scientifically sound approach.

We hypothesise that a personalised autologous organoid system, which mimics the patient’s tissues by using an individual’s colon-derived stem cells, provides a sensitive and specific assessment of CFTR response to modulator treatments.

Objectives {7}

The primary objective of the ORIGIN-1 trial is to determine whether an organoid-guided N-of-1 approach can identify pwCF who have rare, non-F508del mutations and whether an n-of-1 placebo multi-crossover trial can demonstrate a clinically meaningful improvement in lung function following treatment with elexacaftor/tezacaftor/ivacaftor (ETI). Participants will have a colorectal biopsy to collect epithelial tissue and derive participant-specific cells in the laboratory. Individual organoids developed from the biopsy will have endogenous CFTR function assessed and the response to ETI measured in vitro.

Trial design {8}

ORIGIN-1 is a Bayesian, placebo-controlled, within-subject, double-blinded, n-of-1 multi-crossover trial with 5 parallel treatment sequences, each comprising two to four consecutive cycles. Each cycle comprises both a 2-week ETI treatment block followed by a 2-week washout period, and a 2-week placebo (PBO) treatment block followed by a further 2-week washout period. Participants are randomly assigned to one of the five sequences of 4 treatment cycles as shown in Fig. 1.

Fig. 1.

Schematic of study design. Participants will be randomised to one of 5 possible sequences of 4 treatment cycles. Each cycle consists of 2 blocks of 14 days of treatment with ETI and PBO, with 14 days of washout between blocks

In an ETI block, a participant will receive ETI (elexacaftor 200 mg/tezacaftor 100 mg/ivacaftor 150 mg) in the morning, and ivacaftor 150 mg in the evening for 14 consecutive days; in a PBO block, they will receive an ETI-matched placebo morning and evening for 14 consecutive days. During a 2-week washout period, no study medication will be taken. Participants will continue to receive study treatment according to the treatment sequence to which they have been randomised (Fig. 1).

Methods: participants, interventions and outcomes

Study setting {9}

Participants will be recruited from cystic fibrosis clinics in Australia. Only centres that have fulfilled all the duties regarding study selection and training will be added as participating sites. As of August 2025, two Australian sites are active: The John Hunter Hospital in Newcastle, NSW; and The Alfred in Melbourne, Victoria.

Eligibility criteria {10}

Cystic fibrosis patients with rare mutations (non-F508del), aged 6 years or older and with lung function (ppFEV1) between 40 and 90% are eligible for inclusion in the study. A comprehensive list of inclusion and exclusion criteria is summarised in Table 1. Written informed consent will be obtained from the participant or authorised surrogate before inclusion in the study.

Table 1.

Origin-1 trial inclusion and exclusion criteria

Inclusion criteria

Exclusion criteria

1. A confirmed clinical diagnosis of CF as determined by the investigator 2. Age ≥ 6 years old and weighing ≥ 30 kg 3. Informed consent obtained from the participant or a legally responsible guardian 4. Subject has two non-F508del CFTR mutations. Two genetic variants affecting the cystic fibrosis transmembrane regulator, that are listed on the CFTR2 website as CF causing, variants of varying clinical significance or variants of unknown significance 5. Subject has provided or is willing to provide a colonic/rectal biopsy for 3D organoid culture to asses response to ETIin vitro 6. Forced expiratory volume in 1 s (ppFEV1) ≥ 40% and ≤ 90% of predicted mean for age, sex and height using the Global Lung Initiative (GLI) at the screening visit 7. Stable CF disease as judged by the investigator in the absence of any illness or clinical condition that in the judgement of the investigator may pose an additional risk in administering the study medication to the subject 8. Willing and able to adhere to all trial procedures

1. History of lung transplantation 2. Any of the following abnormal laboratory values at screening: ▪ Haemoglobin < 10 g/dL ▪ Total bilirubin ≥ 2 × upper limit of normal (ULN) ▪ Aspartate transaminase (AST), alanine transaminase (ALT), gamma-glutamyl transferase (GGT), or alkaline phosphatase (ALP) ≥ 3 × ULN 3. An acute upper or lower respiratory infection, pulmonary exacerbation(s) (PEx), or changes in therapy (including antibiotics) for sinopulmonary disease within 28 days before the first dose of study medication in the run-in period (day −28) 4. An established contraindication the study medication, i.e. ETI 5. Pregnant or nursing females. All female subjects must have a negative pregnancy test at Screening (urine test) 6. A life expectancy of less than 12 months

Who will obtain informed consent? {26a}

Written informed consent will be obtained from eligible patients by signing the consent form after detailed information is provided by a trained and delegated physician or senior researcher. The principal investigator (PI) retains overall responsibility for the informed consent of participants at their site and ensures that any person with delegated responsibility to participate in the informed consent process is duly authorised, trained and competent.

Additional consent provisions for collection and use of participant data and biological specimens {26b}

By signing the informed consent form, study participants agree with the collection of clinical data and biological specimens as described in the protocol.

Interventions

Explanation for the choice of comparators {6b}

The study is an n-of-1 multi-crossover trial following the treatment block sequence they assigned during randomisation. Participants will self-administer an oral placebo (manufactured by Syntro Pty Ltd, Australia) visually and physically matching the investigational product. Participants will be blinded to whether they are taking ETI or placebo throughout the treatment cycles.

Intervention description {11a}

Eligible participants will be included and randomised to 1 of 5 parallel sequences of treatment, each comprising of up to 4 cycles. Each cycle lasts for 8 weeks and is comprised of a 14 day treatment block with ETI or placebo followed by a 14 day washout period, and a 14 day treatment block with ETI or placebo followed by a 14 day washout period. The order of the ETI and placebo blocks in each cycle varies according to the assigned sequence. Given the uncertainty about an appropriate washout period for ETI, only 5 of the 16 possible sequences of treatment were selected to ensure there is a minimum of 6 weeks between consecutive ETI treatment blocks.

In an ETI block, a participant will receive ETI (Elexacaftor 200 mg/Tezacaftor 100 mg/Ivacaftor 150 mg morning, and Ivacaftor 150 mg in the evening for 14 consecutive days); in a PBO block they will receive an ETI-matched placebo for 14 consecutive days. During a 14 day washout period following treatment blocks no study medication will be taken.

Criteria for discontinuing or modifying allocated interventions {11b}

Participants who develop a generalised rash will be monitored closely. Study medication dosing should be interrupted if a participant develops a generalised rash of Grade 3 or higher, or a rash that is considered a serious adverse event (SAE).

Participants with new treatment-emergent alanine aminotransferase (ALT) or aspartate aminotransferase (AST) elevations of > 3 × upper limit of normal (ULN) and total bilirubin > 2 × ULN will be followed closely, including repeat confirmatory testing performed within 48 to 72 h of the initial finding and subsequent close monitoring of ALT and AST levels, as clinically indicated. If levels remain elevated, the situation will be discussed with the site PI and participant. If needed, this will be referred to the Independent Safety Committee (ISC) and Trial Management Committee (TMC) to arbitrate if the intervention should be discontinued.

Study medication will be ceased immediately if LFTs are > 8 × ULN or clinical symptoms of liver damage become evident. Participants will not recommence ETI/PBO and will be withdrawn from the study.

Strategies to improve adherence to interventions {11c}

At each visit, participants will be given a schedule for their treatment cycle, and site personnel will review the participant’s self-reported compliance and remind the participant/caregiver of study medication dosing requirements.

Relevant concomitant care permitted or prohibited during the trial {11d}

At each study visit (including unscheduled visits such as hospital admissions), concomitant medications will be checked and documented. The use of any CFTR modulator, investigational or approved, is not allowed after the initiation of study treatment or until the completion of study participation.

Moderate and strong CYP3A inducers or CYP3A inhibitors (except ciprofloxacin^a) are not allowed within 14 days before the first dose of study medication and through the completion of study participation.

Provisions for post-trial care {30}

We do not anticipate severe harm during the trial. All participants will have an end of study visit 2 weeks after the last dose of study treatment.

Outcomes {12}

The primary outcome—short-term improvement of lung function (ppFEV1)—will be measured at baseline, randomisation and daily using home spirometry throughout trial participation. Lung function improvement will be expressed as absolute difference in ppFEV1 (ΔppFEV1) after approximately 14 days of ETI therapy, compared to ppFEV1 after approximately 14 days of PBO, after at least 2 and no more than 4 complete cycles. These responses will also be compared to baseline ppFEV1.

The ppFEV1 is an established correlate of survival in CF and was used as the basis of registration of all CFTR modulators. Trials of CFTR modulators (including the Phase 3 and post-approval trials of ETI) have shown that ppFEV1 rises significantly from baseline by day 15 [18]. The time it takes for the effect of ETI to washout is not clear but the mean terminal half-lives of elexacaftor, tezacaftor and ivacaftor are approximately 24.7, 60.3 and 13.1 h, respectively [18]. It is anticipated that most of the loss-of-effect of ETI on ppFEV1 will occur within 14 days and should result in a substantial decrease in ppFEV1 back to pre-treatment baseline. In addition, the 5 randomised sequences were selected such that the minimum period between any two rounds of ETI is at least 6 weeks consisting of a 14-day PBO block between two 14-day washout periods (Fig. 1).

The secondary outcomes will be measured at baseline, randomisation and study visits and are summarised on Table 2.

Table 2.

Secondary outcomes and associated measurements

Secondary outcome	Secondary endpoint
The short-term improvement in lung function	Absolute difference in ppFVC (ΔppFVC) after approximately 14 days of ETI therapy, compared to the ΔppFVC after approximately 14 days of PBO therapy, after at least 2 and no more than 4 complete cycles of ETI and matched PBO
	Absolute difference in self-reported Cystic Fibrosis Respiratory Symptom Diary (CFRSD)-Chronic Respiratory Infection Symptom Score (CRISS) after approximately 14 days of ETI therapy, compared to the CFRSD after approximately 14 days of PBO therapy, after at least 2 and no more than 4 complete cycles of ETI and matched PBO
The short-term improvement in CF-related symptoms	Absolute difference in self-reported sinonasal outcome test (SNOT-22) after approximately 14 days of ETI therapy, compared to the SNOT-22 after approximately 14 days of PBO therapy, after at least 2 and no more than 4 complete cycles of ETI and matched PBO
	Absolute difference in self-reported Cystic Fibrosis Abdominal symptom (CFAbd) score after approximately 14 days of ETI therapy, compared to the CFAbd after approximately 14 days of PBO therapy, after at least 2 and no more than 4 complete cycles of ETI and matched PBO
	Absolute difference in the risk of pulmonary exacerbation in the first 14 days of ETI therapy, compared to the risk in the first 14 days of PBO therapy, after at least 2 and no more than 4 complete cycles of ETI and matched PBO
	Absolute difference in the risk of serious and non-serious adverse events, and the risk of serious and non-serious adverse reactions, in the first 14 days of ETI therapy, compared to the risk in the first 14 days of PBO therapy, after at least 2 and no more than 4 complete cycles of ETI and matched PBO

Participant timeline {13}

The screening period will begin up to 28 days before the first study dose. Participants will be randomly assigned to a treatment sequence in a double-blinded manner, with both the participant and site investigator unaware of the allocation. Each cycle will include an ETI and a placebo treatment block, with study visits—conducted face-to-face or via telehealth—scheduled at the end of each treatment and washout period (e.g. day 14, day 28, and day 42). Lung function will be monitored daily by the participant at home using a portable spirometer, while adverse events and patient-reported outcome measures (CFRSD-CRISS, SNOT-22, CFAbd) will be assessed during study visits. Safety blood tests will be performed every 4 weeks following randomisation. Study medication will be dispensed at key time points and made available for collection at a participant-nominated community pharmacy.

Trial participation will last between 16 and 32 weeks, excluding the 28-day screening period. A minimum study duration of 16 weeks will permit completion of 2 full treatment cycles. Based on a planned interim analysis following the completion of Cycle 2 and Cycle 3, a participant may not be required to complete any further cycles (i.e. Cycles 3 and/or 4) if they have already met the criteria to be classified as either a clinical responder or non-responder to ETI at the completion of Cycle 2 or 3 (see the ‘Statistical analysis’ section).

A participant who cannot be classified as a responder or non-responder will complete the remaining cycle/s extending the overall treatment period for up to 32 weeks.

A final end-of-treatment (EOT) visit, and 2-week follow-up will conclude study participation. All study assessments are specified in Table 3.

Table 3.

Participant timeline: schedule of enrolment, interventions, and assessments

			Treatment cycle
	Screening		ETI/PBO		Washout		PBO/ETI		Washout
	Day −27 to −1	Day 0 (Randomisation day)	Day 1–13	Day 14	Day 15–27	Day 28	Day 29–41	Day 42	Day 43–55	Day 56/(day 0 for next cycle)	EOTf
Study visit#	X	X		X		X		X		X	X
Informed consent	X
Inclusion/exclusion	X	X
Demography	X
Medical History	X
ConMedsa	X	X		X		X		X		X	X
BP, pulse, temp., pulse oximetry, resp. rate, weight, height	X
Physical examinationb	X	X		X				X			X
Daily home or in-clinic spirometryc	X	X	X	X	X	X	X	X	X	X	X
Safety blood—LFTsd (4 mls) every 4 weeks after randomisation	X	X				X				X	X
Sweat chloride test	X
Urine pregnancy test—females	X
Randomisation		X
Dispense Study medication		X				X				X
ETI or PBO Administration			X	X			X	X
Return treatment cycle meds				X				X
Set-up Smart phone app	X
CFRSD-CRISSe	X	X	X	X		X		X		X
SNOT-22e	X	X		X		X		X		X
CFAbd scoree	X	X		X		X		X		X
Adverse Events	X	X	X	X	X	X	X	X	X	X	X

^#Screening visit, Day 0 and EOT visits are planned to occur face to face. For participants who live a distance from the study site or where travel to the site is difficult for that person, study visits will take place virtually. In such cases eConsent may be used to obtain consent. To minimise disruption for participants but to provide accurate and close monitoring and support the other visits are planned to occur virtually or online. Spirometry will be performed at home, questionnaires will be made available using a RedCap link to the participant’s mobile phone and/or email address; and where a participant is randomised and chooses not to attend the clinic for study visits, distribution of study drug will be from the JHH Pharmacy to a nominated community pharmacy located convenient to the participant. If after a telehealth study visit a clinical intervention is advised, remote participants will be referred to their local CF clinic, GP or hospital emergency department for any TEAEs

^aConcomitant medications will be recorded from 30 days prior to the screening period until the end of study for each participant

^bPhysical examination will include chest auscultation and/or symptom-directed assessments

^cSpirometry data will be entered by participants into Spirohome Cloud app via their own personal SmartPhone. All spirometry data will be derived from the subject-issued spirometer

^dAll safety blood analyses will be done at the site’s local pathology service or using a pathology service convenient for the participant if study visits are being done via telehealth. A blood volume of 28 mls is required if 2 cycles are completed and 44 mls if 4 cycles are required

^eCFRSD-CRISS Questionnaire will be completed weekly during each treatment cycle; SNOT-22 and CFAbd questionnaires will be completed at each visit

^fA follow-up interaction will take place 2 weeks after the participant has completed a final treatment cycle. If all treatment cycles are completed, this will be the 22nd study interaction over a 36-week period from initial screening to safety follow-up

Sample size {14}

To understand the trial operating characteristics, a trial simulation was implemented across a range of sample sizes with different numbers of treatment cycles, and assuming plausible scenarios in terms of treatment effect and variability of outcomes within and between participants. Within each simulation, a primary analysis model was implemented for data generation and analysis, and the decision rules (as specified below) were evaluated for all planned interim analyses with participants exiting or continuing in the trial, as appropriate. In addition, the decision rules were evaluated at the end of the trial simulation to assess whether each participant was classified a responder or non-responder. Throughout all simulations, 10% of the daily data was assumed missing completely at random. With conservative estimates of within and between participant variability (e.g. a participant’s response generally varied within ± 8% (in absolute terms) of their average ppFEV1 value and average ppFEV1 values for participants generally varied between 50 and 90% at day 14 when on ETI), it was found that there was a greater than 80% chance of declaring a participant a responder if their underlying increase in ppFEV1 when on active treatment was ≥ 4.5 percentage points. It was also found that, for a given participant, if their underlying decrease in ppFEV1 when on active treatment was ≤ − 3.5 percentage points, then there was a greater than 80% chance of declaring them a non-responder. In addition, there was a low probability (i.e. < 5%) of declaring someone a responder when ETI had no effect. This suggests that the trial design as described above has a high probability of declaring a participant a responder if there is a moderate increase in their lung function when on active treatment and a high chance of declaring a participant a non-responder if there is at least a moderate decrease in lung function when on active treatment.

Recruitment {15}

Up to 20 participants will be recruited from cystic fibrosis treatment centres within Australia. The initial approach for pre-screening potential patients will be done by a member of the patient’s existing clinical care team with referral to an ORIGIN-1 investigator. The ORIGIN-1 investigator will confirm the eligibility of the patient and start the screening process after a written informed consent form is obtained.

Assignment of interventions: allocation

Sequence generation {16a}

At the completion of screening period, suitable participants will be randomly assigned to one of 5 alternative sequences of ETI and placebo and will complete up to 4 × 8-week cycles (Fig. 1). Assignment to one of the 5 sequences will be performed using centralised computer-generated block randomisation, with up to 5 blocks of 5 assignments.

Concealment mechanism {16b}

The randomisation schedule will be generated in R for the whole trial by Prof James McGree, Queensland University of Technology. The randomisation schedule and seed will be held in restricted access folders that are only accessible by the trial pharmacist and unblinded statisticians.

Implementation {16c}

The allocation sequence was generated by Prof McGree.

Assignment of interventions: blinding

Who will be blinded {17a}

All clinical site investigators, care providers, site and sponsor staff, participants and data administrators will be blinded.

Procedure for unblinding if needed {17b}

Individual participant treatment assignments will not be unblinded during the study unless a patient safety issue arises in which unblinding is necessary to ensure optimal patient management. Individual participant unblinding during a treatment cycle will only occur after approval from the coordinating principal investigator (CPI). On receipt of the treatment allocation details, the PI or treating health care professional will continue to deal with a participant’s medical emergency as appropriate.

The breaking of the blinding code and the reasons for doing so will be documented in the participant’s medical notes and the study CRF. Unblinded data will be kept strictly confidential until the trial is unblinded and will not be accessible by study staff.

Once study participation is completed, the database is locked, and individual analysis is completed, treatment assignments will be unblinded.

Data collection and management

Plans for assessment and collection of outcomes {18a}

Individual patient data recorded in an electronic case report form (eCRF) will be handled in compliance with all applicable laws and regulations.

Study data will be transcribed from the source documents on to an eCRF by study staff within five working days of the participant’s visit. Worksheets may be used for the capture of some final visit to facilitate completion of the eCRF. Any worksheets will become part of the subject’s source documentation.

All eCRF entries, corrections, and alterations must be made by the investigator and/or authorised study-site personnel.

Spirometry will be measured every day during the treatment and washout periods and performed according to the American Thoracic Society Guidelines/European Respiratory Society. The spirometry parameters listed below will be normalised using the standards from the Global Lung Function Initiative (GLI):

• FEV1 (L)

• Forced vital capacity (FVC) (L)

All spirometry will be performed using the portable spirometer (Spirohome) issued to the participant for home-based spirometry measurement and captured directly onto the Spirocloud—a web-based cloud connected to the portable spirometers.

CFRSD-CRISS questionnaire

Cystic Fibrosis Respiratory Symptom Diary (CFRSD) and Chronic Respiratory Symptom Score (CRISS) are validated and responsive tools for assessing respiratory symptoms in people with CF. This will be collected at each visit conducted. The change from baseline at the commencement of each cycle will be assessed.

SNOT-22

The sinonasal outcome test (SNOT-22) is a patient-reported outcome that measures nasal and sinus symptoms and their impact on quality of life [19]. This will be collected at each visit conducted. The change from baseline at the commencement of each cycle will be assessed.

CFAbd

The CFAbd is a patient-symptom instrument that has been validated for use in assessing abdominal symptoms in people with CF [20]. This will be collected at each visit conducted. The change from baseline at the commencement of each cycle will be assessed.

Plans to promote participant retention and complete follow-up {18b}

Participants will be encouraged to remain on the assigned treatment and will be closely monitored for the duration of the study. However, at any time during the study and without giving reasons, participants may withdraw from the study at their own request. They will not suffer any disadvantage as a result of their withdrawal or discontinuation. The withdrawal or discontinuation will be recorded in the eCRF and in the participant’s medical records.

Data management {19}

All collected study data will be recorded and stored in a REDCap electronic database, as part of a secured clinical trial management system. The project manager, or delegate, will perform regular validation of data, queries and corrections.

The data will be stored for at least 15 years and the sponsor will be the data custodian.

Confidentiality {27}

Personal information will be collected, kept secure and maintained in accordance with Australian guidelines for protection of privacy. Participant data are identified by a unique study code on all study documentation, including the REDCap database, to ensure confidentiality. Minimum contact data, including email address and mobile phone, will be collected to ensure follow-up can be completed. Consent data, which are identifiable, will be stored separately to all other study documentation in a secure location.

Plans for collection, laboratory evaluation and storage of biological specimens for genetic or molecular analysis in this trial/future use {33}

Rectal biopsies will be recovered by qualified gastroenterologists at the lead site following a fleet enema. Samples will be processed for expansion of cells as organoids for the in vitro testing of CFTR function following treatment with ETI. Biological specimens will be cryopreserved in liquid nitrogen.

Statistical methods

Statistical methods for primary and secondary outcomes {20a}

To estimate the primary outcome of difference in day 14 ppFEV1 after initiating ETI compared to placebo, the daily ppFEV1 measurements will be analysed using Bayesian normal mixed effects linear regression models. The effect of treatment on the mean ppFEV1 response profile is not expected to be constant throughout a treatment period. Instead, a rapid initial increase may occur during the first week of active treatment, with a plateau during the second week. Therefore, a piecewise linear model is assumed for the mean response with a changepoint after day 8. The model includes subject specific random intercepts and coefficients on the piecewise-linear terms and a fixed effect of ETI on each piece. The model also adjusts for random effects of treatment cycle shared between the ETI and placebo periods within the cycle. Weakly informative priors are assumed on model parameters.

The secondary outcome of ppFVC will be analysed similarly to the primary outcome given ppFVC is also measured daily throughout each treatment period.

For the other secondary outcomes which are measured only at day 14 of each treatment period, response-appropriate mixed-effects models for the day 14 measure will be assumed. For Likert scale outcomes, cumulative logistic regression models will be assumed and for binary outcomes, logistic regression models. Model appropriate transformations will be used to obtain mean differences in day 14 outcomes.

Interim analyses {21b}

Analyses are scheduled to occur as each participant completes a second, third or fourth treatment cycle. Participants who have completed at least one cycle will contribute data to the statistical model, and participants who have completed at least two cycles will be assessed for their response to treatment.

At each interim analysis, the primary model will be estimated and pre-specified decision rules used to determine whether a participant may be considered a responder or non-responder to ETI. If they have completed at least two cycles, a participant-specific estimate (marginal posterior density) of the difference in day 14 ppFEV1 for ETI compared to PBO will be assessed against the decision rules. A participant will be classified as a responder if the posterior median difference is greater than 2.5 percentage points and the posterior probability that the difference exceeds zero is at least 0.95. If both conditions are satisfied, the participant will be classified a responder to ETI and will not undergo any additional treatment cycles. A participant will be classified a non-responder if the posterior probability that their ppFEV1 difference exceeds 2.5 percentage points is less than 0.05. If either condition is satisfied, the participant will not undergo any additional treatment cycles. If neither is satisfied, then the participant will progress to another cycle (up to the maximum of 4 cycles).

Methods for additional analyses (e.g. subgroup analyses) {20b}

Association of organoid response with clinical response to treatment will be assessed initially by plotting the organoid response against participant estimates of treatment effect so that the nature of any association can be assessed. The piecewise linear model will be extended by including an interaction between the organoid response and ETI treatment, with the plot being used to inform what may be an appropriate functional relationship for including organoid response into the linear predictor. Once this model is fitted, the posterior distribution of the associated fixed effect parameter will then be assessed to determine the magnitude of association. To assess the predictive performance of the model with organoid response, leave-one-group-out cross validation will be used to compare against the reduced model which does not use organoid response.

Methods in analysis to handle protocol non-adherence and any statistical methods to handle missing data {20c}

The primary analysis is based on the assigned treatment policy and will not adjust for non-adherence to the intended treatment. The analysis will assume that missing ppFEV1 measurements are missing at random under the assumed model and only available measurements will be used for each participant. Sensitivity analyses may investigate other missingness assumptions using multiple imputation and the influence of any non-adherence to treatment.

Plans to give access to the full protocol, participant-level data and statistical code {31c}

The ORIGIN-1 Trial is registered on ANZCTR ID: ACTRN12623001136695ID and participant-level data may be available upon request.

Oversight and monitoring

Composition of the coordinating centre and trial steering committee {5d}

The Origin-1 trial is coordinated by a core management team based at the Hunter Medical Research Institute and The University of Newcastle in partnership with the University of Sydney, and Queensland University of Technology. It is composed of a coordinating principal investigator (CPI), co-investigators, clinical trial manager, lead pharmacist, data managers and biostatisticians who are responsible for operational management of trial including recruitment strategy, randomisation, monitoring, safety reporting, protocol adherence, data integrity and communication with participating sites. CPI, co-investigators and clinical trial manager will meet bimonthly (in person or via videoconference), and sub-groups will meet more frequently as required.

An independent safety monitor (ISM) is charged with providing independent safety oversight and adherence to the prespecified safety measures and project decision-making processes. The ISM is responsible for reviewing safety reports for serious adverse events (SAEs) and suspected unexpected serious adverse reactions (SUSARs), identifying potential threats to project safety and providing independent safety advice if the need arises.

Composition of the data monitoring committee, its role and reporting structure {21a}

The ORIGIN-1 trial has an Independent Statistical Monitoring Committee (ISMC) composed of three independent statisticians with expertise in clinical research and Bayesian adaptive methods. The committee is responsible for reviewing interim statistical reports to ensure alignment with the pre-specified Statistical Analysis Plan, confirming the appropriate implementation of trial adaptations, identifying potential threats to the scientific integrity of the project, and providing independent statistical advice if required. The ISMC operates entirely independently of the trial investigators and sponsor and does not make decisions on operational or management aspects of the study. Meetings are held on an ad hoc basis during recruitment and treatment phases, typically in response to scheduled interim analyses. Any concerns or recommendations identified by the ISMC are reported to the Trial Manager and CPI, who retain full decision-making authority for the design and conduct of the trial.

Adverse event reporting and harms {22}

Safety evaluations will include adverse events (AEs), urine pregnancy testing and blood biochemistry, i.e. liver function testing (ALT, AST, direct bilirubin, and total bilirubin). Participants will be questioned about adverse events at each visit, and reported adverse events will be documented. The severity of adverse events will be graded as outlined by the National Institute of Health (NIH), and the severity of AEs and causality will be assigned by the PI at each site.

All serious adverse events (SAEs), regardless of causality, will be reported to the coordinating investigator within 24 h of an awareness of the event; corrections and additions are also required to be submitted within 24 h as they become available.

Unexpected drug-related SAEs as determined by the local PI and the coordinating investigator or authorised person qualify for expedited reporting and will be reported to the EC, regulatory authorities and participating investigators.

SAEs, including those that do not meet the requirements for expedited reporting, and all other AEs will be reported to regulatory agencies as appropriate.

Frequency and plans for auditing trial conduct {23}

An audit may be carried out at any time, and may apply to all stages of the research, from protocol development to publication of results and classification of data used or produced in the course of the study.

Plans for communicating important protocol amendments to relevant parties (e.g. trial participants, ethical committees) {25}

Amendments to the protocol will not be implemented until approval from all relevant ethical and regulatory review bodies. Amendments will be communicated to participant sites and all correspondence with the EC will be retained in the Investigator Site File.

Dissemination plans {31a}

Upon completion of the trial, the final study report, including the results of the study, will be made available at ANZCTR.gov.au. The results will also be communicated by presentation and publication in peer-reviewed journals. All the investigators will have the opportunity to review publications prior to submission.

Discussion

The ORIGIN-1 trial aims to demonstrate that among pwCF who show an in vitro response to modulator therapy, an n-of-1 randomised placebo-ETI multi-crossover trial can be used to assess a clinically meaningful response. This information is expected to help identify pwCF who may benefit from CFTR modulator treatment but who are currently not eligible for modulator therapy.

Patient-derived organoids are a relevant model for studying CFTR function and response to modulators at both the mutation and individual levels. This allows a more personalised assessment of the CFTR modulator response, complementing traditional mutation-based approaches to identifying responsive patients. While the use of patient-derived cells to assess the CFTR modulator response has expanded significantly in recent years, clinical validation and the potential to predict patient outcomes using in vitro methods remain limited [21–23].

A recent observational study examined the compassionate use of ETI in pwCF with rarer mutations. Although the study involved a small sample size and lacked control groups, the findings contributed to the growing body of evidence suggesting that pwCF with rare mutations may benefit from ETI treatment and highlight the need for improved pre-clinical assessment tools [24].

To our knowledge, the ORIGIN-1 trial will be the first n-of-1 placebo-controlled clinical trial to evaluate the clinical efficacy of ETI in pwCF with rare mutations based on patient cell-derived responses to modulators. Despite the high cost of treatment, the trial aims to enrol up to 20 pwCF with rarer mutations to ensure that this trial platform and its findings can be translated into clinical practice and improve health outcomes.

Trial status

The current protocol version 8.0 dated 08 April 2025 was approved on 03 July 2025. Recruitment started in November 2023 and will be completed in mid-2026.

Abbreviations

CF

Cystic fibrosis

CFTR

Cystic fibrosis transmembrane regulator

MF

Minimal function

ETI

Elexacaftor/tezacaftor/ivacaftor

PBO

Placebo

FEV1

Forced expiratory volume in 1 s

TGA

Therapeutic Goods Administration

PBS

Pharmaceutical Benefits Scheme

QUM

Quality use of medicines

Pex

Pulmonary exacerbation

CPI

Coordinating principal investigator

PI

Principal investigator

LFTs

Liver function transaminases

ALT

Alanine aminotransferase

AST

Aspartate aminotransferase

ULN

Upper limit normal

TMC

Trial Management Committee

CFRSD-CRISS

Cystic Fibrosis Respiratory Symptom Diary – Chronic Respiratory Infection Symptom Score

SNOT-22

Sinonasal outcome test

CFAbd

Cystic Fibrosis Abdominal symptom score

eCRF

Electronic case report file

FVC

Forced vital capacity

DSMB

Data Safety Monitoring Board

MCAR

Missing data completely at random

MNAR

Missing data not at random

AE

Adverse events

SAE

Serious adverse events

NIH

National Institute of Health

IRB/EC

Independent Review Board/Ethics Committee

Authors’ contributions {31b}

PABW is the lead clinical principal investigator; he conceived the study, led the proposal and protocol development. GEK conceived the study, contributed to protocol development and study design. LRS and DD contributed to protocol development and study design. AJ, AS, and SW contributed to proposal and study design. DC, HK, KT, and TG contributed to protocol development. JT and JM led the statistical study design. TS is the lead trial methodologist and conceived the study. All authors read and approved the final manuscript.

Funding {4}

Funding is provided by the Australian Federal Government’s Medical Research Future Fund (MRFF) Clinical Trials Activity – Rare Cancers, Rare Diseases and Unmet Need grant (GNT2014916). The current funding period ends in 2026. The funding body will have no role in the design of the study or the collection, analysis, or interpretation or reporting of data. Funding allocation to parties contributing to the study will be administered by the University of Newcastle and will be in accordance with a research collaboration agreement.

Data availability {29}

Access to data will be granted to authorised representatives from the steering committee, sponsor, host institution and regulatory authorities to enable trial-related monitoring, audits and inspections. Site investigators will be allowed access to the full dataset if a formal request describing their plans is approved by the steering committee.

Declarations

Ethics approval and consent to participate {24}

This study has been approved by the Hunter New England Human Research Ethics Committee, registration number 2023/STE01015 on May 16th 2023. Written informed consent is obtained from all participants.

Consent for publication {32}

Not applicable. No identifying images or other personal or identifiable clinical information of participants are presented here or will be presented in reports of the trial results.

Competing interests {28}

GK has a patent on an RNA therapy for CFTR, and shareholding in Akte Therapeutics a gene therapy company. TMG has received speaker fees and research support from Johnson & Johnson and speaker and advisory fees from Abbvie.

The other authors declare that they have no competing interests.