Long-Term Safety Follow-Up of Subjects Previously Treated with Non-Replicating Retroviral Vector-Based Gene Therapies.

OBJECTIVE: Our objective was to evaluate the life-long safety profile of gene therapy using retroviral (non-replicating) vectors (nRCR), or cell products in 127 subjects with hemophilia, human immunodeficiency virus (HIV), or cancer, previously treated with such gene therapy. METHODS: We assessed the occurrence of serious adverse events (SAEs), deaths and presence of replication competent retrovirus (RCR). RESULTS: A total of 23 subjects remained until the data cut-off date of 31 July 2013 and provided safety information of up to 18 years. Of the 104 subjects who discontinued, the primary reason was loss to follow-up (47.2 %; n = 60). The follow-up period for the 60 subjects lost to follow-up was 7-10 years. A total of 41 subjects experienced at least one SAE, and 15 subjects died. We reviewed SAEs and cause of death (none related to the active therapy), but no evidence was found for safety signals related to new malignancy or neurologic, rheumatological, autoimmune, or hematologic disorder. RCR results were negative, indicating no evidence for in vivo vector persistence. CONCLUSION: Despite the loss of follow-up, which is the limiting factor in this long-term safety trial, the findings from this long-term follow-up study are encouraging.

Assays for the quantification of melphalan and its hydrolysis products in human plasma by liquid chromatography-tandem mass spectrometry.

Two high-performance liquid chromatography tandem mass spectrometry (LC-MS/MS) assays are described for the quantification of melphalan in human plasma. N-phenyldiethanolamine was tested as internal standard. The first assay consisted of a protein precipitation by cold methanol and a reversed-phase HPLC whereas the second one was based on a solid phase extraction and a hydrophilic interaction chromatography. Both provided a very satisfactory mean extraction yield with a small volume of sample. The first method was simple, rapid and used as a routine assay. The second one was developed in order to determine melphalan hydrolysis products and to avoid scarce cases when interferences from biological matrix alter the quantification of melphalan using the first method. The two assays were linear and sensitive in the range of 1-500ng/mL for the first one and in a range of 25-2000ng/mL for the second one. Concentrations out of the range fixed with the first method were also validated. The procedure was reliable with precision and accuracy below 10%. All compounds were detected after positive mode electrospray ionization in selected reaction monitoring mode. These new analytical procedures were developed for melphalan pharmacokinetic studies or therapeutic drug monitoring.