Transplantation of Cord Blood Expanded Ex Vivo with Copper Chelator.

Cord blood (CB) is used to restore hematopoiesis in transplant patients lacking marrow donors. CB is associated with higher rates of delayed/failed engraftment. Peled et al developed an expansion technology using the copper chelator tetraethylenepentamine (TEPA), which enhanced the expansion of primitive CB populations when combined with early acting cytokines. A phase I clinical trial employing this technology was initiated. 10 patients with high-risk, heavily pre-treated hematologic malignancies (AML-2, ALL-5, HD-2, and NHL-1) have been enrolled with CB units that were cryopreserved in 2 fractions [20:80% (n=2), 40:60% (n=5) or 50:50% (n=3)]. 21days prior to infusion, AC133+ cells were isolated from the smaller (if unequal) or 50% CB fractions using the CliniMACS device and cultured for 21 days in media containing 10% FBS and SCF, FLT-3, IL6, TPO plus the copper chelator TEPA (Gamida). Patients then received myeloablative therapy with ATG and either fludara and busulfan (AML), or fludara, melphalan, thiotepa (ALL, HD, NHL) with infusion of the unmanipulated CB fraction on day 0, and the expanded fraction on day +1. GVHD prophylaxis was methotrex 5 mg/m² days 2, 4,7, and tacrolimus for 6 months. The median age was 21 (range 7–51) and weight 69 (range 31–156) kg. The CB units were matched at 4/6 (n=8) or 5/6 (n=2) HLA antigens. The pre-thaw total nucleated cell (TNC) dose of the CB units was a median of 2.5x10⁷/kg with post-thaw TNC of 2.4 x10⁷/kg. Following AC133-selection, the manipulated CB fractions were a median of 73 (range 38–95)% AC133+ with a median of 0.650 (range 0.16–2.7) x10⁶ TNCs, which were placed in culture. After 21 days of culture the expanded fraction had 69 (range 2–1638) x10⁶ TNCs representing a 207 (range 2–616) fold TNC expansion. Patients received a total (expanded plus unmanipulated) median of 1.8 (range 1.1–6.1) x10⁷ TNC/kg and 1.6x10⁵ (range 0.4–49.9) CD34+ cells/kg. Two patients have CB cultures in progress. Of the 8 patients transplanted, 1 had autologous recovery with relapse of AML on day 30 and death. Of the remaining patients, 7 were evaluable for neutrophil engraftment and 4 of them for platelet engraftment (2 too early for platelet evaluation and 1 early death). The median time to engraftment was 27 days for neutrophils (range 16–46) and 48 days for platelets (range 27–96). Preliminary analysis suggest a correlation between a shorter time to neutrophil engraftment and total TNC/kg infused (p=0.02), and a trend for CD34+ cells/kg infused (p=0.09). Three patients have developed grade ≤2 acute skin GVHD and one had chronic extensive GVHD of the skin and GI tract; all resolved with steroids. One patient (without GVHD) died of a systemic viral infection on day 56, despite adequate neutrophil recovery (not platelets). All of the remaining patients are all alive and free of malignancy at a median follow-up of 4 (range 1–16) months.

Conclusion: There was no toxicity associated with infusion of the TEPA-expanded CB cells. Additional data is necessary to determine the efficacy of this approach. Future directions include the expansion of the entire CB unit and removal of methotrex from the GVHD regimen to improve time to neutrophil engraftment, as well as comprehensive assessment of immune reconstitution.