Absence of Replication-Competent Retrovirus in Vectors, T Cell Products, and Patient Follow-Up Samples

Main Text

Two papers published in this issue of Molecular Therapy report on the results of testing for replication-competent lentivirus (RCL) or retrovirus (RCR) in the vector product as well as the infused vector-transduced T cell lines and patient follow-up samples in recipients of genetically modified T cells.1, 2 In the first study, the National Gene Vector Biorepository (NGVB) reports results from 26 trials using third-generation lentiviral vectors produced by transient transfection and that all assays for RCL in vector lots, 460 infused products, and 296 follow-up samples from patients were negative.¹ In the second report, investigators at the University of Pennsylvania reported data for 17 vector lots, 375 manufactured T cell products, and 308 patient follow-up samples, showing no evidence of RCR/L.² Both groups concluded that, with this body of safety testing, it may be appropriate to re-evaluate the current guidelines for T cell gene therapy product testing and long-term patient monitoring.

In 2012, the experience of our group with RCR testing of vector products, transduced cell lines, and patient samples was included in a multicenter report that detailed negative testing on 297 infused T cell lines and 629 patient follow-up samples.³ Here we present updated data that includes testing performed in the last 6 years and not reported in our previous publication. As summarized in Table 1 we have manufactured vectors for 17 clinical trials, using 9 different retroviral constructs where we have tested 9 lots. A total of 266 cell lines have been manufactured, and all have tested negative for RCR using a co-culture assay. 220 patients have been infused with these products and have had follow-up RCR assays run by the NGVB in the first year post infusion, with subsequent follow-up assays archived. A total of 549 patient samples have been assayed, and all have been negative.

Table 1.

Testing Results for RCR

Clinical Trial No	Retroviral Vector	Vector Supernatant Results	Cell Line Results	Patient Follow-Up Results
NCT00368082 NCT02379520 NCT02065362	TGF-β RIIdcyt (DNRII)	negative	17 negative 5 negative 7 negative	54 negative 4 negative 15 negative
NCT005863914	CAR-CD19. CD28. ζ and CAR-CD19. ζ	negative	27 negative	37 negative
NCT01853631	CAR-CD19. CD28. ζ and CAR-CD19.41BB .CD28. Ζ	negative	34 negative	33 negative
NCT007108925 NCT014941036	iCaspase9. ACD19	negative	11 negative 14 negative	33 negative 26 negative
NCT02050347 NCT008408537	CAR-CD19.CD28. ζ	negative	6 negative 21 negative	10 negative 50 negative
NCT00889954	CAR-HER2. CD28. ζ TGFβ RIIdcyt (DNRII)	negative	22 negative	42 negative
NCT009020448 NCT011090959	CAR-HER2. CD28. ζ	negative	26 negative 18 negative	68 negative 35 negative
NCT01192464 NCT0131614610	CAR-CD30. CD28. ζ	negative	4 negative 10 negative	10 negative 37 negative
NCT0088192011	CAR-Kappa.CD28. ζ	negative	17 negative	49 negative
NCT0182265212 NCT01953900	CAR-GD2.CD28. OX40. ζ	negative	18 negative 9 negative	31 negative 15 negative
Total	–	9 negative	266 negative	549 negative

These data presented on RCR testing supplement our previous report and support the data on RCL testing presented in the two papers in this issue, which suggest that RCR/RCL testing of the transduced lines and patient follow-up samples are no longer needed if the viral vector supernatant used to genetically modify the T cells initially tested negative for replication-competent virus.

Author Contributions

D.L. supervised the RCR assays and collated the data. N.L. and A.P.G. provided QA review. All authors reviewed the letter.