Abstract

Introduction: Gene therapy may provide long-term benefit in hemophilia B with a single administration. BAX 335 (AAV8.sc-TTR-FIXR338Lopt) delivers a self-complementary codon-optimized hyperactive F9 transgene (FIXR338Lopt), driven by the liver-specific transthyretin (TTR) promoter in an adeno-associated virus 8 (AAV8) capsid. Shedding is an area of concern with vectors of viral origin and must be evaluated as part of the regulatory application process.

Objective: A secondary objective of this ongoing, phase 1/2, open-label study is to determine the duration of BAX 335 genome shedding into blood, saliva, semen, urine, and stool after single-dose administration in subjects with hemophilia B. Results of an unplanned preliminary analysis of data on shedding from this study are presented.

Methods: Up to 16 adult men with severe hemophilia B are to be given a single intravenous dose of BAX 335 in up to 4 sequentially ascending dose cohorts: Cohort 1 (2.0x1011 vector genome [vg]/kg), Cohort 2 (1.0x1012 vg/kg), Cohort 3 (3.0x1012 vg/kg), and Cohort 4 (4.0x1012 vg/kg). A quantitative real time polymerase chain reaction (qPCR) is used to detect vector genomic DNA sequences (BAX 335 genomes) in body fluids collected from each subject. Whole blood, saliva, semen, urine, and stool samples are obtained before study drug administration; on day 1; at weeks 1, 2, and 3; and at weekly intervals thereafter until negative (2 consecutive samples below limit of detection).

Results: The study is ongoing and preliminary results are available for 7 subjects (S1-S7; Table). All subjects were negative for BAX 335 genomes in all samples at screening. In blood, BAX 335 genomes were detected at the first weekly visit in all subjects, with peak levels achieved at that time. BAX 335 genomes were considered negative in blood between 5 and 9 months after treatment in the 4 subjects (S1-4) for whom follow-up is complete. The remaining 3 subjects (S5-S7) have not yet achieved a negative result in blood (S5) or have not yet reached the month 9 visit (S6 and S7) at the time of this abstract and are still being followed (Table). BAX 335 genomes were also identified at lower levels in saliva (n=7), semen (n=5), stool (n=7), and urine (n=4), with peak values achieved between 1 day and 2 weeks and persisting for 1-5 weeks. The safety profile for all subjects enrolled to date was acceptable, including no inhibitors to factor IX (FIX).

Table 1.

BAX 335 Genome in Whole Blood and Semen After Single Intravenous Dosing

Subject No. First Time Detected* Peak Time Detected* Last Time Detected Time Negative 
Cohort 1 (2.0x1011 vg/kg) 
S1
Blood
Semen 
D 1 (2908)
ND 
D 1 (2908)
ND 
M 4 (99)
ND 
M 5
ND 
S2
Blood
Semen 
D 1 (828513)
D 1 (154) 
D 1 (828513)
D 1 (154) 
M 4 (445)
Wk 1 (67) 
M 5
Wk 2 
Cohort 2 (1.0x1012 vg/kg) 
S3
Blood
Semen 
D 1 (>1x106)
D 1 (273) 
D 1 (>1x106)
D 1 (273) 
M 5 (105)
Wk 3 (137) 
M 6
Wk 4 
S4
Blood
Semen 
D 1 (>1x106)
D 1 (4771) 
D 1 (>1x106)
Wk 2 (6421) 
M 6 (93)
Wk 3 (753) 
M 9
Wk 5 
S5
Blood
Semen 
D 1 (>1x106)
ND 
D 1 (>1x106)
ND 
M 12 (61)
ND 
NR
ND 
Cohort 3 (3.0x1012 vg/kg) 
S6
Blood
Semen 
D 1 (>1x106)
D 1 (2098) 
D 1 (>1x106)
D 1 (2098) 
M 6 (185)
Wk 2 (751) 
NR
Wk 3 
S7
Blood
Semen 
D 1 (>1x106)
D 1 (2236) 
D 1 (>1x106)
Wk 1 (10680) 
Wk 12 (66815)
Wk 3 (196) 
NR
Wk 4 
Subject No. First Time Detected* Peak Time Detected* Last Time Detected Time Negative 
Cohort 1 (2.0x1011 vg/kg) 
S1
Blood
Semen 
D 1 (2908)
ND 
D 1 (2908)
ND 
M 4 (99)
ND 
M 5
ND 
S2
Blood
Semen 
D 1 (828513)
D 1 (154) 
D 1 (828513)
D 1 (154) 
M 4 (445)
Wk 1 (67) 
M 5
Wk 2 
Cohort 2 (1.0x1012 vg/kg) 
S3
Blood
Semen 
D 1 (>1x106)
D 1 (273) 
D 1 (>1x106)
D 1 (273) 
M 5 (105)
Wk 3 (137) 
M 6
Wk 4 
S4
Blood
Semen 
D 1 (>1x106)
D 1 (4771) 
D 1 (>1x106)
Wk 2 (6421) 
M 6 (93)
Wk 3 (753) 
M 9
Wk 5 
S5
Blood
Semen 
D 1 (>1x106)
ND 
D 1 (>1x106)
ND 
M 12 (61)
ND 
NR
ND 
Cohort 3 (3.0x1012 vg/kg) 
S6
Blood
Semen 
D 1 (>1x106)
D 1 (2098) 
D 1 (>1x106)
D 1 (2098) 
M 6 (185)
Wk 2 (751) 
NR
Wk 3 
S7
Blood
Semen 
D 1 (>1x106)
D 1 (2236) 
D 1 (>1x106)
Wk 1 (10680) 
Wk 12 (66815)
Wk 3 (196) 
NR
Wk 4 

D=day; M=month; ND=not detected; NR=not reached; vg=vector genome; Wk=week.

*Values in parentheses are the number of copies/10 mL blood or semen.

Subject still being followed.

Conclusions: From these preliminary data for the first 7 subjects dosed in the study, a trend towards dose response in terms of both magnitude and duration of AAV shedding into whole blood, saliva, stools, urine, and semen was observed. Shedding has also been reported in other studies of AAV-mediated gene transfer in subjects with severe hemophilia B (Manno CS et al. Nat Med. 2006;12:342-7; Nathwani AC et al. N Engl J Med. 2014;371:1994-2004). The peak values for shedding in our study were observed between day 1 and week 2 in all subjects. Four subjects have completed the follow-up with 6 months post-infusion being the last time a positive sample was detected. Three subjects (S5-S7) are still being followed. S5 has for the past year had sustainable expression levels of FIX at 20% or above and 12 months post-infusion is persistently positive for vector genome in blood (all other fluids negative since week 3).

Disclosures

Litchev:Baxalta: Employment. Monahan:Pfizer: Honoraria; Baxter/Baxalta: Consultancy, Honoraria, Research Funding; Bayer: Consultancy; Asklepios BioPharmaceutical: Consultancy, Patents & Royalties: Author I.P. licensed by UNC to AskBio, Research Funding; Novo Nordisk: Consultancy, Honoraria, Research Funding; Prolor: Research Funding; Chatham LLC: Consultancy; CSL Behring: Consultancy, Honoraria. Prener:Novo Nordisk: Other: previous employee; Uniqure: Other: previous consultant; Baxalta: Employment.

Author notes

*

Asterisk with author names denotes non-ASH members.