Open Label, Study Of Efficacy and Safety Of AVR-RD-01 for Treatment-Naive Subjects With Classic Fabry Disease

The duration of each subject's participation in this study was approximately 64 weeks (or 1 year, 12 weeks), comprised of five study periods (Screening, Baseline, Pre-transplant, Transplant, and Post-transplant Follow-up). During the Screening Period (approximately 8 weeks), written informed consent (and assent, if applicable) was obtained and the subject had completed other Screening procedures to confirm study eligibility. Once study eligibility was confirmed, the subjects entered the Baseline Period (up to 3 days) during which time assessments would have been performed to establish a pre-transplant baseline. Once baseline assessments were completed, the subject entered the Pre-transplant Period (approximately 6 weeks) during which time mobilization, apheresis, AVR-RD-01 investigational drug product preparation and testing for release, and conditioning regimen administration to achieve myeloablation took place. Following completion of the Pre-transplant Period, the subject entered the Transplant Period (1 day) during which time AVR-RD-01 infusion took place. After AVR-RD-01 infusion, the subject entered the Post-transplant Follow-up Period (approximately 48 weeks), during which time periodic safety and efficacy assessments were performed to assess measures of engraftment, clinical response, and safety post-transplant.

In January 2022, the study was terminated early due to a decision by the study sponsor, to deprioritize its Fabry disease development program, and therefore, some subjects (n=5) did not complete the study (i.e., Week 48). Subsequently, in August 2023, the long-term follow-up study (AVRO-RD-01-LTF01), was also terminated early due to the decision by the sponsor to terminate the development program for Fabry disease, and therefore, no subjects completed the 15-year long-term follow-up study. This decision to terminate was not based on any safety or medical reasons.

• AVR-RD-01 (Drug): Single IV infusion of between 3 - 20 x 10\^6 CD34+ cells/kg.

Primary outcomes

• Incidence of and Severity of Adverse Events (AEs) and Serious Adverse Events (SAEs) [Baseline to Week 48 post gene therapy]: An AE was any untoward medical occurrence in a participant who received study drug without regard to possibility of causal relationship. An SAE is an AE resulting in any of the following outcomes or deemed significant for any other reason: death; initial or prolonged inpatient hospitalization; life threatening experience (immediate risk of dying); persistent or significant disability/incapacity; congenital anomaly. The AE/SAE are also inclusive of any abnormalities in Clinical Laboratory Tests, Vital Signs and in Electrocardiographs (ECGs). Of the 13 serious adverse events, no SAEs reported were considered related to AVR RD 01. Of the 354 adverse events, no AEs were considered related to AVR RD 01. The SAEs and AEs reported in the study were attributed to the conditioning agent used, the underlying disease, comorbid conditions, study procedures and concomitant medications.
• Change From Baseline in Immunogenicity of AVR-RD-01 [Baseline to Week 48 post gene therapy]: Number of subjects with changes in anti-AGA antibodies from Baseline to post infusion timepoints. Unite of measure: Number of subjects negative at baseline but positive at post-treatment timepoints. A negative or zero result (titer lower or unchanged at post-infusion timepoints compare to Baseline) indicates no immune response to the therapeutic protein.
• Presence of Replication Competent Lentivirus (RCL) [Baseline to Week 48 post gene therapy]: The "Presence of RCL" is a theoretical risk of lentiviral gene therapy treatment based on the theory that it may be possible for inadvertent generation of RCL caused either by recombination of the lentiviral vector plasmids during the vector production process or by mobilization of proviral DNA in vivo by infectious retroviruses (HIV). The absence of RCL is a positive indicator of safety.
• Evaluation of Aberrant Clonal Expansion [Baseline to Week 48 post gene therapy]: Integration Site Analysis (ISA) uses next generation sequencing to identify junction sites between the integrated therapeutic transgene and the host genome. Samples are analyzed for the emergence of clonality (defined as (a single clone accounting for greater than 20% of the population) and whether any integration site is within or near a known oncogene.
• Change From Baseline in the Average Number of Gb3 Inclusions (ie, Myelinosomes) Per Kidney Peritubular Capillary (PTC) Per Subject [Baseline to Week 48 post gene therapy]: Globotriaosylceramide (Gb3) Inclusions in Peritubular Capillaries (PTC) on Kidney Biopsy. Electron microscopic images of kidney biopsy samples were taken and read centrally by two independent renal pathologists, each of whom scored the average number of Gb3 inclusions per kidney PTC per subject using a quantification method. Healthy renal tissue would have no Gb3 inclusions. A reduction from baseline is desirable.

Secondary outcomes

• Average Vector Copy Number (VCN) in Peripheral Blood Leukocytes as Assessed by Quantitative Polymerase Chain Reaction (qPCR) and/or Droplet Digital Polymerase Chain Reaction (ddPCR) [At Week 24 and Week 48 post gene therapy]: Vector Copy Number (VCN) is a measurement of the number of copies of the therapeutic transgene found in a sample, relative to copies of a reference gene in the human genome. This is an estimate of the number of integration sites per cell (on average). A VCN of 1 would signify that a sample of cells evaluated contains on average at least one \[working\] copy of the therapeutic transgene per cell.
• Change From Baseline (CFB) in AGA Enzyme Activity Level in Plasma and Peripheral Blood Leukocytes (PBLs) [Baseline to Week 24 and Week 48 post gene therapy]: Treatment-naïve Fabry patients are deficient in alpha-galactosidase A (AGA) enzyme activity due to mutations in the GLA gene. Any therapeutic option offered should aim to increase the amount of available AGA enzyme. This assay measured the AGA enzyme activity levels in plasma and PBLs. It should be noted that the measurement in plasma reflects the amount of "free" AGA enzyme that has been released from cells into the extracellular space and is therefore considered a more indirect measure of AGA enzyme activity, compared to the result in PBLs which is more of a direct measure of enzyme within cells. In both cases, enzyme activity is expected to increase from Baseline to the post-infusion timepoints.
• Change From Baseline in Globotriaosylceramide (Gb3) Biomarkers for Fabry Disease in Plasma [Baseline to Week 24 and Week 48 post gene therapy]: Globotriaosylceramide (Gb3) is the substrate that accumulates in the lysosomes of patients affected by Fabry Disease as a result of deficiencies in AGA enzyme activity. Treatment-naive patients are expected to have high levels of Gb3 in their lysosomes and correspondingly elevated levels in plasma. Treatment with AVR-RD-01 is intended to replace the missing AGA enzymatic activity, which allows degradation of accumulated Gb3 substrate in the lysosomes and reductions in the levels of circulating Gb3 in plasma.
• Change From Baseline in Globotriaosylceramide (Gb3) Biomarkers for Fabry Disease in Urine [Baseline to Week 24 and Week 48 post gene therapy]: Globotriaosylceramide (Gb3) is the substrate that accumulates in the lysosomes of patients affected by Fabry Disease as a result of deficiencies in AGA enzyme activity. Treatment-naive patients are expected to have high levels of Gb3 in their lysosomes and correspondingly elevated levels in urine. Treatment with AVR-RD-01 is intended to replace the missing AGA enzymatic activity, which allows degradation of accumulated Gb3 substrate in the lysosomes and reductions in the levels of excreted Gb3 in urine.
• Change From Baseline in Substrate (i.e. Gb3) in Skin Biopsy [Baseline to Week 24 and Week 48 post gene therapy]: Globotriaosylceramide (Gb3) is the substrate that accumulates in the lysosomes of patients affected by Fabry Disease as a result of deficiencies in AGA enzyme activity. Treatment-naive patients are expected to have high levels of Gb3 in their lysosomes and correspondingly elevated levels in tissue samples. Treatment with AVR-RD-01 is intended to replace the missing AGA enzymatic activity, which allows degradation of accumulated Gb3 substrate in the lysosomes and reductions in the levels of measured Gb3 in tissues.
• Change From Baseline in Renal Function as Assessed by Measured Glomerular Filtration Rate (mGFR) [Baseline to Week 48 post gene therapy]: mGFR is a measure of the time the kidney takes to filter products that the body does not naturally produce.
• Change From Baseline in Renal Function as Assessed by Estimated Glomerular Filtration Rate (eGFR) [Baseline to Week 24 and Week 48 post gene therapy]: eGFR is the measure to evaluate kidney function. It is the estimated amount of blood that is filtered through all glomeruli in a given time.
• Change From Baseline in Renal Function as Assessed by Urine Total Protein Levels [Baseline to Week 24 and Week 48 post gene therapy]
• Change From Baseline in Renal Function as Assessed by Urine Albumin Levels [Baseline to Week 24 and Week 48 post gene therapy]: A healthy kidney only allows very small amounts of albumin to pass from the blood into the urine. An increased level of albumin in urine (albuminuria) is a marker of renal damage.
• Change From Baseline in Left Ventricular Mass Index (LVMI) as Assessed by Cardiac Magnetic Resonance Imaging (MRI) [Baseline to Week 48 post gene therapy]: LVMI is a surrogate of left ventricular hypertrophy. An increase in LVMI is an independent risk factor for cardiovascular morbidity and mortality.
• Change From Baseline in Abdominal Pain and Stool Consistency as Assessed by the Diary for Irritable Bowel Syndrome Symptoms-Diarrhea (DIBSS-D) [Baseline to Week 24 and Week 48 post gene therapy]: DIBSS-D assesses bowel habits and abdominal symptoms over a period of time. It was administered daily for 14 days commencing at each study visit. Stool Consistency scale has been converted to a numeric rating scale for ease of analysis, where 1=Very hard; 2=Hard; 3=Neither too hard nor too soft; 4=Loose but not lumpy; 5=Very loose and watery. The median of each 14-day period was derived per patient, per visit before deriving the group median. Group median at Baseline (pre-treatment) was 3.540 (n=9). Change from baseline (CFB) is presented below. An increase CFB indicates softening of the stools, and a decrease CFB indicates hardening of the stools. Abdominal Pain measure asked the patient to rate the worst level of pain within the past 24hrs (0=no pain; 10=worst possible pain). A mean score was derived for each 14-day period per patient, per visit, and these means used to derive a group mean. An increase CFB indicates more abdominal pain; a decrease CFB indicates less abdominal pain.
• Change From Baseline in Brief Pain Inventory-Short Form (BPI-SF) Questionnaire Scores [Baseline to Week 24 and Week 48 post gene therapy]: The short version of the BPI (Short form) includes 9 items: Q1 - Q9, Question 9 includes 7 sub-items (Q9a - Q9g). It uses a 0 to 10 numeric rating scales for item rating. The pain severity score is calculated as the average of questions answered: Q3 (worst pain), Q4 (least pain), Q5 (average pain) and Q6 (current pain). The pain interference score is calculated as the average of the answered Q9 sub-items, which represents pain interference with general activity (Q9a), mood (Q9b), walking ability (Q9c), normal work (Q9d), relations with other people (Q9e), sleep (Q9f), and enjoyment of life (Q9g). A reduction in score from baseline indicates less pain.
• Change From Baseline in Physical and Mental Functioning as Assessed by the Short Form 36 (SF-36) Physical Component Summary (PCS) and Mental Component Summary (MCS) Scores [Baseline to Week 48 post gene therapy]: The original version of the SF-36 was administered to the participants and consisted of eight subscales (Vitality, Physical Functioning, Bodily Pain, General Health Perceptions, Physical Role Functioning, Emotional Role Functioning, Social Role Functioning and Mental Health) each scored from 0 (worst health) to 100 (best heath). These scores were normalized (re-scaled) against mean scores obtained in the US general population (Mean=50, Standard deviation 10). The summary health components PCS and MCS are derived from the eight subscales mentioned above and summarize information from all eight subscales but with different weights. For PCS, highest weights are given to the physical subscales while some mental subscales are given negative weights. For MCS, highest weights are given to the mental subscales while some physical subscales are given negative weights. An increase in the normalized score from baseline indicates improvement in physical and mental functioning.
• Average Vector Copy Number (VCN) in Bone Marrow / Progenitor Cells as Assessed by Quantitative Polymerase Chain Reaction (qPCR) and/or Droplet Digital Polymerase Chain Reaction (ddPCR) [At Week 48 post gene therapy]: VCN is defined as the average number of copies of the therapeutic gene (transgene) in a sample of cells and is a measurement of the number of copies of the vector found in a sample, relative to copies of a reference gene in the human genome. This is an estimate of the number of integration sites per cell (on average). A VCN of 1 would signify that a sample of cells evaluated contains on average at least one \[working\] copy of the therapeutic transgene per cell. This measurement was for VCN in a sample of Bone marrow progenitor cells obtained from an aspirate.

Sex: Male, ages: 16 Years to 50 Years

Inclusion Criteria:

1. Subject was male, 16 years of age or older (18 years of age or older in the US), and post pubertal,(minimum age by region)
2. Subject had a confirmed diagnosis of classic Fabry disease based on deficient AGA enzyme activity (defined as < 1% of normal).

Exclusion Criteria:

1. Subject had a galactosidase alpha (GLA) gene mutation associated with late-onset cardiac variant Fabry disease.
2. Subject had previously received ERT and/or chaperone therapy within 3 years for treatment of Fabry disease.
3. Subject had tested positive for anti-AGA antibodies at the time of screening.
4. Subject had eGFR < 60 mL/min/1.73 m² (ie, chronic kidney disease \[CKD\] stage ≥ 3) at Screening.
5. Subject had a prior history of myocardial infarction (MI).
6. Subject had a history of coronary artery disease (CAD) with angina requiring percutaneous transluminal coronary angioplasty (with or without stent placement) and/or coronary artery bypass graft (CABG).
7. Subject had a history of moderate to severe valvular heart disease requiring valve replacement.
8. Subject had a history of heart failure, moderate to severe diastolic dysfunction, and/or left ventricular ejection fraction (LVEF) ≤ 45% on echocardiogram (ECHO) performed at rest at Screening.
9. Subject had a history of clinically significant cardiac arrhythmia (eg, heart block \[second or third degree\], atrial fibrillation requiring therapy, ventricular fibrillation, ventricular tachycardia, supraventricular tachycardia, or cardiac arrest).

   Note \[history of intermittent atrial fibrillation not requiring treatment was allowed\].
10. Subject had a prior history of stroke and/or transient ischemic attack (TIA).
11. Subject had aspartate aminotransferase (AST) and/or alanine aminotransferase (ALT) ≥ 3 times the upper limit of normal (ULN) at Screening.
12. Subject had a prior history of (or current) malignancy; the one exception is a prior history of resected basal cell carcinoma.
13. Subject had previously received treatment with AVR-RD-01 or any other gene therapy.

Other inclusion/exclusion criteria apply.

Participant flow

Outcome results

• Incidence of and Severity of Adverse Events (AEs) and Serious Adverse Events (SAEs) (Primary), Number, Number of events: Single Assignment AVR-RD-01 354
• Change From Baseline in Immunogenicity of AVR-RD-01 (Primary), Number, Number of positive subjects: Single Assignment AVR-RD-01 4
• Presence of Replication Competent Lentivirus (RCL) (Primary), Number, Number of positive subjects: Single Assignment AVR-RD-01 0
• Evaluation of Aberrant Clonal Expansion (Primary), Number, number of abnormal clonal proliferations: Single Assignment AVR-RD-01 0
• Change From Baseline in the Average Number of Gb3 Inclusions (ie, Myelinosomes) Per Kidney Peritubular Capillary (PTC) Per Subject (Primary), Mean, Number of inclusions: Single Assignment AVR-RD-01 -3.555 (-4.020 to -3.090)
• Average Vector Copy Number (VCN) in Peripheral Blood Leukocytes as Assessed by Quantitative Polymerase Chain Reaction (qPCR) and/or Droplet Digital Polymerase Chain Reaction (ddPCR) (Secondary), Mean, copies/diploid genome: Single Assignment AVR-RD-01 0.383 +/- 0.4628
• Change From Baseline (CFB) in AGA Enzyme Activity Level in Plasma and Peripheral Blood Leukocytes (PBLs) (Secondary), Mean, nmol/h/mg: Single Assignment AVR-RD-01 1.973 +/- 2.585
• Change From Baseline in Globotriaosylceramide (Gb3) Biomarkers for Fabry Disease in Plasma (Secondary), Mean, nM: Single Assignment AVR-RD-01 -3334.0 +/- 3846.8
• Change From Baseline in Globotriaosylceramide (Gb3) Biomarkers for Fabry Disease in Urine (Secondary), Mean, nmol/mmol creatinine: Single Assignment AVR-RD-01 -11.73 +/- 309.78
• Change From Baseline in Substrate (i.e. Gb3) in Skin Biopsy (Secondary), Mean, number of myelinosomes: Single Assignment AVR-RD-01 -0.4 +/- 1.3
• Change From Baseline in Renal Function as Assessed by Measured Glomerular Filtration Rate (mGFR) (Secondary), Mean, mL/min/1.73m^2: Single Assignment AVR-RD-01 -2.358 +/- 14.502
• Change From Baseline in Renal Function as Assessed by Estimated Glomerular Filtration Rate (eGFR) (Secondary), Mean, mL/min/1.73 m^2: Single Assignment AVR-RD-01 -6.386 +/- 9.699
• Change From Baseline in Renal Function as Assessed by Urine Total Protein Levels (Secondary)
• Change From Baseline in Renal Function as Assessed by Urine Albumin Levels (Secondary), Mean, mg/dL: Single Assignment AVR-RD-01 0.915 +/- 6.704
• Change From Baseline in Left Ventricular Mass Index (LVMI) as Assessed by Cardiac Magnetic Resonance Imaging (MRI) (Secondary), Mean, g/m^2: Single Assignment AVR-RD-01 7.270 +/- 6.055
• Change From Baseline in Abdominal Pain and Stool Consistency as Assessed by the Diary for Irritable Bowel Syndrome Symptoms-Diarrhea (DIBSS-D) (Secondary), Mean, units on a scale: Single Assignment AVR-RD-01 -0.445 +/- 0.583
• Change From Baseline in Brief Pain Inventory-Short Form (BPI-SF) Questionnaire Scores (Secondary), Mean, score on a scale: Single Assignment AVR-RD-01 -0.188 +/- 0.874
• Change From Baseline in Physical and Mental Functioning as Assessed by the Short Form 36 (SF-36) Physical Component Summary (PCS) and Mental Component Summary (MCS) Scores (Secondary), Mean, score on a scale: Single Assignment AVR-RD-01 3.235 +/- 4.810
• Average Vector Copy Number (VCN) in Bone Marrow / Progenitor Cells as Assessed by Quantitative Polymerase Chain Reaction (qPCR) and/or Droplet Digital Polymerase Chain Reaction (ddPCR) (Secondary), Mean, copies/diploid genome: Single Assignment AVR-RD-01 1.14 (1.14 to 1.14)

Adverse events

Time frame: Up to week 48

Serious adverse events by term

Other adverse events by term

89 more terms not shown.

Limitations and caveats: SF-36 data at W24 was inadvertently not collected due to omission in the Protocol Schedule of Assessments. Invasive sampling, such as bone marrow aspirate, was only provided by one subject at Week 48. Central laboratory only reported albumin levels in urine and not urine total protein data.

• Study Protocol (PDF, 2020-04-01)
• Statistical Analysis Plan (PDF, 2023-07-26)

• Inderpal Panesar, MRPharmS, Study director, AVROBIO, Inc