• ClinicalTrials.gov Identifier: NCT02576795¹
• A Phase 1/2, Dose-Escalation, Safety, Tolerability and Efficacy Study of Valoctocogene Roxaparvovec, 
  an Adenovirus-Associated Virus Vector-Mediated gene transfer of human FVIII in Patients with severe Haemophilia A

• Ongoing development, Active, not recruiting
• Last Update Posted: August 06, 2024¹

• Male ≥18 years of age with base FVIII level ≤ 1 IU/dL
• Treated/exposed to FVIII concentrates or cryoprecipitate for a minimum of 150 Eds

• Detectable pre-existing immunity to the AAV5 capsid^a
• Prior treatment with any vector or gene transfer agent
• Use of systemic immunosuppressive agents or live vaccines within 30 days before the viral infusion

^a Cell-based in vitro transduction-inhibition assay and total antiAAV5 antibody assay

AAV5/baculovirus, Spodoptera frugiperda (Sf9) insect-cell production system²

    Codon-optimized expression cassette encoding the SQ variant of B-domain–deleted human FVIII²

    HLP³ is a 251-bp enhancer/promoter fragment consisting of:

• A 34-bp core enhancer from the human ApoE-HCR gene upstream of 
• A modified 217-bp gene promoter, which includes the distal X and the
   proximal A1B regulatory domains from SERPINA1 (α-1-antitrypsin)

The FVIII expression cassette also includes a synthetic polyA signal^{2 [supplementary material]}

    Systemic (peripheral vein infusion)²

• Co. 1: 6e12 (1)²
• Co. 2: 2e13 (1)²
• Co. 3: 6e13 (7)^2, 4, 5
• Co. 4: 4e13 (6) ^4, 5

    52 weeks², 3 years⁴, 4 years⁵ and 5 years⁶

• FVIII activity levels were assessed by both OS and CH clotting assays^2, 4, 10
• OS FVIII activity levels were approximately 1.6⁴ to 1.65² times as high as the FVIII levels from CH 

Data on FVIII activity levels up to week 52 for the low and intermediate cohorts (Co. 1 and Co. 2) have been reported in reference #2 and are summarized as follows:

FVIII activity at specific thresholds through week 52 in the high dose cohort was published in reference #2  and is summarized as follows:

Multiyear mean and median FVIII activity levels for Co. 3 (4e13 vg/kg) and Co. 4 (6e13 vg/kg) were published in references #4, #5, #6, #9 and #10 and are summarized as follows:

* Missing data from 2/7 pts. in Co. 3 (6e13 vg/kg) that resumed prophylaxis at year 7 are not imputed in the calculation of mean and median FVIII activities
† Missing data from 2/6 pts. in Co. 4 (4e13 vg/kg) are not imputed in the calculation of mean and median FVIII activities

• When including results prior to resuming prophylaxis, respective mean FVIII activity was 12.6 (median, 5.1) IU/dL (n = 7) 
  and 5.2 (median, 4.5) IU/dL (n = 6) for the 6e13 vg/kg and 4e13 vg/kg cohort¹⁰

Data concerning the annual absolute % change in mean FVIII activity per CSA were published in reference #10 are summarized below as follows:

As shown in Figure 1 in reference #4, the times to peak response are summarized as follows:

• 20 to 24 weeks in Co. 3 (6e13 vg/kg, n=7) and
• 44 weeks in Co. 4 (4e13 vg/kg, n=6)

Early follow-up efficacy data on ABR in Co. 1–Co. 3 were reported in reference #2 and are summarized below

• Co. 1: not applicable, as the only pt. in this cohort resumed FVIII prophylaxis after W16
• Co. 2: the ABR increased from 3 to 11 events in 1/1 pts. with stable but low FVIII activity level (1 to 3 IU/dL) through W54
• Co. 3: the median ABR dropped from 16 events per year before the study to 1event per year after gene transfer (mean reduction in rate, from 16 to 2 events)

The most recent multiyear follow-up of treated ABRs was reported in reference #10 and is summarized as follows:

Summary of reported reductions from baseline in treated bleeds per year over the course of up to 7 years, as shown below:

* Six of the seven participants were receiving regular FVIII prophylaxis at baseline (one participant was receiving on-demand FVIII prophylaxis
and was excluded). Baseline (n = 6) ABR mean and median were 16.3 and 16.5 bleeds/year, and the mean ABR over the entire study was
0.77 bleeds/year, representing a 95% decrease from baseline.

Early follow-up efficacy data related to AIR in Co. 1 – Co. 2 were reported here² and are summarized as follows:

The most recent multiyear follow-up of AIRs before and after treatment was reported here¹⁰ for in both high dose cohorts and is summarized as follows:

* Six of the seven participants were receiving regular FVIII prophylaxis at baseline (one participant was receiving on-demand FVIII prophylaxis 
and was excluded). Baseline (n = 6) ABR mean and median were 16.3 and 16.5 bleeds/year, and the mean ABR over the entire study was
0.77 bleeds/year, representing a 95% decrease from baseline.

In Co. 3 (6e13 vg/kg), 3/7 (42.9%) pts. and in Co. 4, 4/6 pts. (66.7%) were affected by IRRs⁶

The latest summary of the incidence of AEs in each year by cohort across a span of up to seven years was reported here¹⁰ and is summarized as follows:

Note: Data are presented as n (%).
Abbreviations: AE, adverse event; ALT, alanine aminotransferase; MedDRA, Medical Dictionary for Regulatory Activities; SAE, serious AE; ULN, upper limit of normal; Y, year.
^a Pyrexia on study day 2.
^b Defined as ALT ≥1.5× ULN or ALT ≥1.5× baseline.
^c Identified with a MedDRA search strategy using the high-level term “liver function analyses.”

AEs and SAEs at the five years follow-up were reported here⁶ and are summarized as follows:

Abbreviations: AE, adverse event; ALT, alanine aminotransferase; AST, aspartate aminotransferase; SAE, serious adverse event; ULN, upper limit of normal; Y, year. Data are presented as n (%).
^a Percentages were calculated using the total number of participants (N) in each analysis population as the denominator. Participants with more than one AE of the same category were counted only once for that category.
^b Relationship to study drug was assessed by the investigator.
^c A Grade 1 AE of ALT increase (1.4 × ULN) occurred on study day 1389. The AE was deemed not related to the study drug by the investigator and resolved by the next study visit on day 1431 without treatment.
^d A Grade 1 AE of ALT increase (1.3 × ULN) occurred on study day 1626. The AE was deemed not related to the study drug by the investigator and resolved by study day 1631.
^e A Grade 1 AE of ALT increase (2.1 × ULN) occurred on study day 1100. The AE was deemed not related to the study drug by the investigator and resolved by the next study visit on day 1135 without treatment.
^f A Grade 1 AE of AST increase (1.7 × ULN) occurred on study day 1100. The AE was deemed not related to the study drug by the investigator and resolved by the next study visit on day 1135 without treatment.

AEs and SAEs including also the event numbers at the two years follow-up were reported here⁴ and are summarized as follows:

AE: adverse event , SAE: serious adverse event
* Adverse events were graded in accordance with the Common Terminology Criteria for Adverse Events, version 4.0.3 (CTCAE)
† Treated ALT elevations includes all ALT elevations that resulted in initiation or modification of glucocorticoid treatment

Further statements related to AEs and SAEs were reported here^{4, 6} and summarized as follows:

• Most common AEs associated with valoctocogene roxaparvovec were transient, asymptomatic, and mild-to-moderate ALT elevations
• Treatment related SAE of grade 2 pyrexia developed along with myalgia and headache in 1/1 pt. (P13, Co. 4) within 24 hours after the administration of AAV5-hFVIII-SQ
  - Symptoms resolved within 48 hours after the participant received Acetaminophen

Not reported 

8/9 pts. (59-128 U/L, 4-52 weeks)² and 11/15 pts. (peak and timing not reported)⁵

3/9 pts. (peak levels and timing not reported)² and 10/15 pts. (peak levels and timing not reported)⁴ 

Data regarding capsid directed T cell activation were reported by different publications as follows:

• 0/9 pts. (peak and timing not reported)² and Sporadic, transient positives in some pts.⁴ 
• All 15 pts. were tested negative for the AAV5-specific IFN-g responses at baseline⁷
• 4 /15 pts. were tested positive (R50 SFU/106 PBMCs) for AAV5-specific IFN-g responses at a single time point ^a post-administration and negative at all subsequent time points tested (Figure 4A) ⁷

^a PBMC samples were collected at baseline and at several time points after administration that varied by cohort but were generally every 2–4 weeks during the first 6 months, 4–8 weeks through the rest of year 1, and every 3 months thereafter

•  Yes, all pts. with elevated ALT levels were steroid responsive (mean duration of steroid treatment was 21 weeks in 7/9 pts.)²
•  Co. 3: Steroid treatment in 2/3 participants did not inhibit ALT elevations (duration not reported)⁴

• The increase in ALT levels led to decline in the FVIII activity in 1/9 (11 %) pts. in Co. 3²
  • The level in this pt. declined from 227 IU/dL to 52 IU/dL in conjunction with an increase in the ALT level from W28 to W35. 
  • The FVIII activity level subsequently increased, and no bleeding was reported
• The elevations in the ALT level appeared to have been temporally associated with decreases in FVIII expression in 4/9 pts. (P4, 5, 7, and 8) in the high dose cohort, Co. 3⁴ 

 Follow-up data regarding a diagnosed cancer in a pt. were taken from here⁸ and summarized as follows: 

• One SAE in a pt. was diagnosed as salivary gland cancer
• The cancerous mass was removed, and the pt. was monitored
• The study investigator considered the SAE as being unrelated to the treatment
   
• BioMarin announced the findings from genomic analysis from a tissue sample containing the mass, stating that: “The results  showed a comparable pattern of integration between healthy and tumor containing tissues, with no evidence emerging that vector integration contributed to the salivary gland mass.”

1. Gene Therapy Study in Severe Haemophilia A Patients (270-201). Available at: Study Details | Gene Therapy Study in Severe Haemophilia A Patients (270-201) | ClinicalTrials.gov
2. Rangarajan, S., et al., AAV5-Factor VIII Gene Transfer in Severe Haemophilia A. N Engl J Med, 2017. 377(26): p. 2519-2530. AAV5–Factor VIII Gene Transfer in Severe Hemophilia A | NEJM
3. McIntosh, J., et al., Therapeutic levels of FVIII following a single peripheral vein administration of rAAV vector encoding a novel human factor VIII variant. Blood, 2013. 121(17): p. 3335-44. Therapeutic levels of FVIII following a single peripheral vein administration of rAAV vector encoding a novel human factor VIII variant | Blood | American Society of Hematology (ashpublications.org)
4. Pasi, K.J., et al., Multiyear Follow-up of AAV5-hFVIII-SQ Gene Therapy for Haemophilia A. New England Journal of Medicine, 2020. 382(1): p. 29-40. Multiyear Follow-up of AAV5-hFVIII-SQ Gene Therapy for Hemophilia A | NEJM
5. Pasi, K., et al., Hemostatic Response is Maintained for up to 5 Years Following Treatment with Valoctocogene Roxaparvovec, an AAV5-hFVIII-SQ Gene Therapy for Severe Hemophilia A [presentation]. ISTH 2021 Congress, 2021. Hemostatic-Response-is-Maintained-for-up-to-5-Years-Following-Treatment-with-Valoctocogene-Roxaparvovec-an-AAV5-hFVIII-SQ-Gene-Therapy-for-Severe-Hemophilia-A.pdf (biomarin.com)
6. Pasi, K.J., et al., Persistence of haemostatic response following gene therapy with valoctocogene roxaparvovec in severe haemophilia A. Haemophilia, 2021. 27(6): p. 947-956. Persistence of haemostatic response following gene therapy with valoctocogene roxaparvovec in severe haemophilia A - PMC (nih.gov)
7. Long, B.R., et al., Early Phase Clinical Immunogenicity of Valoctocogene Roxaparvovec, an AAV5-Mediated Gene Therapy for Hemophilia A. Mol Ther, 2021. 29(2): p. 597-610. Early Phase Clinical Immunogenicity of Valoctocogene Roxaparvovec, an AAV5-Mediated Gene Therapy for Hemophilia A - PMC (nih.gov)
8. BioMarin follows up on previously-reported serious adverse event in its phase I/II gene therapy clinical trial for haemophilia A. 2022, WFH World Federation of Hemophilia. BioMarin follows up on previously-reported serious adverse event in its phase I/II gene therapy clinical trial for haemophilia A – WFH - World Federation of Hemophilia
9. Symington E, Rangarajan S, Lester W, Madan B, Pierce GF, Raheja P, Robinson TM, Osmond D, Russell CB, Vettermann C, Agarwal SK, Li M, Wong WY, Laffan M. Long-term safety and efficacy outcomes of valoctocogene roxaparvovec gene transfer up to 6 years post-treatment. Haemophilia. 2024 Mar;30(2):320-330. doi: 10.1111/hae.14936. Epub 2024 Feb 5. PMID: 38317480. Long-term safety and efficacy outcomes of valoctocogene roxaparvovec gene transfer up to 6 years post-treatment - PubMed (nih.gov)
10. Symington E, Rangarajan S, Lester W, Madan B, Pierce GF, Raheja P, Millar C, Osmond D, Li M, Robinson TM. Valoctocogene roxaparvovec gene therapy provides durable haemostatic control for up to 7 years for haemophilia A. Haemophilia. 2024 Jul 8. doi: 10.1111/hae.15071. Epub ahead of print. PMID: 38975624. Valoctocogene roxaparvovec gene therapy provides durable haemostatic control for up to 7 years for haemophilia A - PubMed (nih.gov)
11. Pasi, K., et al., Hemostatic Response is Maintained for up to 5 Years Following Treatment with Valoctocogene Roxaparvovec, an AAV5-hFVIII-SQ Gene Therapy for Severe Hemophilia A. ISTH 2021 Congress, 2021. Abstract Number: OC 67.1

AAV, Adeno-associated virus; ABR, Annualized bleeding rate; AEs: adverse events; AIR, Annualized FVIII/FIX infusion rate; ALT, alanine aminotransferase; AST, aspartate aminotransferase; CH, Chromogenic Assay; Co., cohort; DOACs, Direct oral anticoagulants; D, days; EDs, exposure days; FIX, factor IX; FIX-Padua, gain of function FIX variant; FVIII, factor VIII; gc, genome copies; HEK cells, human embryonic kidney cells; IQR, interquartile range; IRR, Infusion-related reaction; NAbs, neutralizing antibodies; OS, One-stage clotting assay; Pop., population; pt., patient/participant; pts., patients/participants; P1, Participant 1; PI, phase I; PBGD, porphobilinogen deaminase; PBMC, peripheral blood mononuclear cells; SAEs, serious adverse events; SFU, spot-forming units; TAC, tacrolimus; ULN, upper limit of normal; VCN, vector copy number; vg, vector genomes; W, weeks; WT, wild type; Y, year