Biodistribution (within the body) and Vector Shedding (excretion/secretion)

Valoctocogene roxaparvovec-rvox transgene DNA levels (total amount of vector DNA) in various tissues (evaluated in nonclinical studies), blood, and shedding matrices were determined using a quantitative polymerase chain reaction (qPCR) assay. This assay is sensitive to transgene DNA, including fragments of degraded DNA. It does not indicate whether DNA is present in the vector capsid, in cells or in the fluid phase of the matrix (e.g., blood plasma, seminal fluid), or whether intact vector is present. Plasma and semen matrices were further evaluated by measuring encapsidated (potentially infectious) vector DNA using an immunoprecipitation quantitative PCR assay in Studies 270-201 and 270-301.

Nonclinical Data

Biodistribution of ROCTAVIAN was assessed in adult male mice. Following intravenous administration of 2.1 × 1014 vg/kg, the highest vector DNA concentration was detected in the liver, followed by lower levels in the lung, heart, lymph nodes, kidney, spleen, bone marrow, testis, and brain through six months post-administration. The expression of the hFVIII mRNA transcripts were primarily detected in the liver, with no or minimal expression in extrahepatic tissues.

Clinical Data

ROCTAVIAN biodistribution and vector shedding were investigated on samples from blood, saliva, semen, stool, and urine. Administration of ROCTAVIAN at the dose of 6 × 1013 vg/kg resulted in detectable vector DNA in blood and all shedding matrices evaluated at the dose of 6 × 1013 vg/kg, with peak concentrations observed between 1 and 9 days post-administration. The peak vector DNA concentrations were observed in blood, followed by saliva, semen, stool, and urine. The peak concentration observed to date in blood across two clinical studies was 2 × 1011 vg/mL. The maximum concentration observed in any shedding matrix was 1 × 1010 vg/mL. After reaching the maximum in a matrix, the transgene DNA concentration declines steadily.

In patients treated in two clinical studies, encapsidated (potentially transmissible) vector DNA was detectable in plasma up to 10 weeks after ROCTAVIAN administration.

All patients treated in clinical studies achieved the first of 3 consecutive measurements below the lower limit of quantification (LLOQ) for vector DNA in semen by 36 weeks, and all except one patient achieved 3 consecutive measurements below limit of detection (BLOD) or negative by the time of the data cut. The maximum time to the first of 3 consecutive measurements BLOD for encapsidated (potentially transmissible) vector DNA in semen was 12 weeks.

In clinical studies, all patients achieved 3 consecutive measurements below the LLOQ for vector DNA in urine and saliva, and 126 (89%) patients achieved 3 consecutive measurements below the LLOQ for vector DNA in stool by the time of the data cut. The maximum time to the first of 3 consecutive LLOQ measurements was 8 weeks for urine, 52 weeks for saliva, and 131 weeks for stool. All patients in the first study achieved 3 consecutive measurements BLOD or negative in urine, saliva, and stool by five-year post-dosing. All patients in the second study achieved 3 consecutive measurements BLOD or negative in urine, and saliva, and 85 (63%) patients achieved 3 consecutive measurements BLOD or negative in stool by three-year data cut.

Magnitude and duration of shedding appear to be independent of the patient's attained factor VIII activity.