The way they did this study can be scrutinized at the NEJM website, but I am not going into it in any detail. Although the 'methods' part of the paper is vital for scientists who want to evaluate the work, a lay audience has to take them on trust.
The trial (ClinicalTrials.gov number, NCT01029366) was not looking at whether the treatment worked; that will have to come later. It was designed to assess the safety of the procedure and whether it was really feasible. It was approved by the institutional review board at the University of Pennsylvania. No commercial sponsor was involved in the study.
They designed a self-inactivating lentiviral vector (GeMCRIS 0607-793), which was subjected to preclinical safety testing. Lentiviruses are types of retroviruses that are now commonly used in Gene Therapy experiments and have lots of advantages. They can deliver a significant amount of genetic information into the DNA of the host cell and have the unique ability among retroviruses of being able to replicate in non-dividing cells, so they are one of the most efficient methods of a gene delivery vector. HIV is an example of a lentivirus.
T-cells from the patient were thawed and infected with lentivirus to express the CD19-specific chimeric antigen receptor. Four days before the cell infusion, the patient received chemotherapy designed for depletion of lymphocytes (pentostatin and cyclophosphamide without rituximab). Three days after chemotherapy but before cell infusion, the bone marrow was hypercellular with approximately 40% involvement by CLL. Cytogenetic analysis showed two separate clones, both resulting in loss of chromosome 17p and the TP53 locus. Four days after chemotherapy, the patient received a total of 3×10e8 T-cells, of which 5% were infected with the lentivirus, for a total of 1.42×10e7 infected cells (1.46×10e5 cells per kilogram) split into three consecutive daily intravenous infusions (10% on day 1, 30% on day 2, and 60% on day 3). No postinfusion cytokines such as IL-2 were given and there were no toxic effects of the infusions.
Clinical Response and Evaluations
Fourteen days after the first infusion, the patient began having chills and low-grade fevers associated with fatigue. Over the next 5 days, the chills got worse, and his temperature rose to 102.5°F, with more shivering attacks, feeling 'shocky', loss of apetite, nausea, and diarrhea. He had no lung or heart symptoms. Because of the fevers, CXR and blood, urine, and stool cultures were performed, but all were negative or normal. Tumor lysis syndrome was diagnosed on day 22 after the infusion with raised uric acid, phosphate and LDH levels. There was evidence of acute kidney damage, with a raised creatinine level. The patient was hospitalized and treated with fluid and rasburicase. The uric acid level returned to the normal range within 24 hours, and the creatinine level within 3 days and he was discharged on hospital day 4.
By day 28 after the CART19-cell infusion, the lymph nodes could no longer be felt, while on day 23, there was no evidence of CLL in the bone marrow. The karyotype was now normal, and FISH testing was negative for deletion TP53. Flow-cytometric analysis showed no residual CLL, and B cells were not detectable. CT scanning performed on day 31 after infusion showed no lymph node enlargement.
Three and 6 months after the CART19-cell infusion, physical examination remained unremarkable, with no lymph nodes to feel and CT scanning performed 3 months after the CART19-cell infusion showed sustained remission. Bone marrow studies at 3 and 6 months also showed no evidence of CLL, with a continued lack of normal B cells as well. Remission had been sustained for 10 months so far.