Double cord blood transplant

Whereas double UCB HCT may provide significantly more rapid neutrophil engraftment compared with use of single units, engraftment delays and failure rates remain higher when compared with bone marrow or mobilized blood sources. Furthermore, an area of continued investigation is to explain the observation of why only one unit will ultimately predominate.

In transplant recipients receiving 2 or more UCB units, one of the UCB units predominates, usually by 4 to 6 weeks after transplant. Infusion of the UCB that does not engraft may augment engraftment via immune activation, inhibition of recipient-mediated immune rejection, or the combination. On the other hand, relapse rates appear lower with 2 than with 1 unit. To date, there have been no convincing data that infusion of 2 rather than 1 UCB unit is superior; given the significantly higher cost and other issues, the standard of care remains uncertain.

Ex vivo UCB unit expansion: MSCs

De Lima and colleagues at the MD Anderson Cancer Center recently reported the early findings of a study whereby they ex vivo cocultured one of two UCB units infused in the course of a double UCB HCT. For expansion, they used either third-party haploidentical family member marrow-derived MSCs or commercially available mesenchymal progenitor cells (Angioblast Systems, Inc, New York, NY). Thirty-two patients received a MAC regimen consisting of fludarabine, melphalan, thiotepa, and antithymocyte globulin (ATG). The culture yielded a median 14-fold increase in total nucleated cells and 40-fold increase in CD34 ⁺ cells. Median time to neutrophil and platelet engraftment was 15 (range 9–42) days and 40 (range 13–62) days; there were no toxicities attributable to the expanded cells. The expanded unit contributed only to early blood cell recovery; long-term engraftment was provided by the unexpanded unit.

Ex vivo UCB unit expansion: TEPA

Peled and co-workers described a NOD/SCID mouse model in which the linear polyamine copper chelator TEPA augmented long-term ex vivo expansion of UCB-derived CD34 ⁺ cells and increased their engraftment potential. Subsequently, the MD Anderson Cancer Center group conducted a clinical trial in which 10 high-risk hematologic malignancy patients (n = 5 ALL; n = 2 AML) received high-dose cytotoxic therapy followed by infusion of a UCB unit that had been cryopreserved in 2 fractions. The GVHD prophylaxis regimen was combination tacrolimus with “mini-methotrexate” (3 days). One of the fractions was administered unexpanded, whereas the other component was CD133-selected and expanded in vitro in media containing stem cell factor (SCF), FLT-3 ligand, interleukin (IL)-6, thrombopoietin, and TEPA. The total nucleated cell dose in the manipulated component was expanded 219-fold and the CD34 ⁺ cell content increased sixfold. Despite the fact that the mean total nucleated cell content infused was low (1.8 × 10 ⁷ /kg), 9 of 10 patients engrafted at times comparable with those associated with infusion of unmanipulated single UCB units, for example, neutrophils at 30 days and platelets at 48 days. These data demonstrate that a graft with low cell numbers can be used successfully and justify exploring this approach in a larger patient series, in which methotrexate should be omitted from the GVHD prophylaxis regimen.

Ex vivo UCB unit expansion: Notch ligand

The Notch signaling pathway has been known to play an important role in hematopoiesis. Hence, the investigators at the FHCRC initiated a phase 1 MAC UCB HCT trial in 10 high-risk acute leukemia patients who were given one nonmanipulated and one ex vivo manipulated UCB graft. One UCB unit was thawed and the CD34 ^+– selected cells were expanded in serum-free media containing the Notch ligand Delta1 supplemented with SCF, thrombopoietin, FLT-3, IL-3, and IL-6. CD34 ⁺ cells were expanded (mean) 164-fold, and median (range) time to attain neutrophil recovery greater than 500/μL was 16 (range 7–34) days; these numbers compare favorably with values of 26 (range 16–48) days ( P = .002) from a concurrently treated cohort of 20 patients undergoing double UCB transplantation.

These novel UCB expansion projects are the subject of several investigations in various stages, undertaken by academic centers as well as corporate partners.

Clofarabine-based

Clofarabine, a new potent purine nucleoside antimetabolite that inhibits DNA polymerase and ribonucleotide reductase, is approved by the Food and Drug Administration for the treatment of relapsed or refractory childhood ALL. Biochemical and clinical studies suggest synergy with cytarabine and significant activity in acute leukemia; hence, this agent has been combined in preparative regimens. Ricci and colleagues reported a combination of clofarabine, cytarabine, and TBI followed by allogeneic HCT in 12 poor-risk children, adolescents, and young adults with acute leukemia. Clofarabine was dose-escalated to the maximum tolerated dose of 52 mg/m ² /d for 5 days. TRM at 100 days was 0%; 2 patients had progressive disease while 8 patients remain alive in continuous complete remission at a median (range) of 209 (40–770) days. At the 2011 BMT tandem meeting, Agura and colleagues described preliminary results of a single-institution phase 2 clofarabine and busulfan RIC regimen in 20 hematologic malignancy (n = 16 AML) patients. Eleven subjects with active malignancy attained complete remission by day 30; 7 experienced relapse and overall, 12 died but 6 remain in remission at a median (range) follow-up of 31 (13–41) months. Kirschbaum and colleagues performed a phase 1 dose-escalation study of clofarabine and melphalan in 16 AML patients, median age 63 (range 30–66) years. The investigators did not identify dose-limiting toxicities; 2 patients expired early from multiorgan toxicities. Only 2 subjects relapsed and 11 remain in remission at a median (range) of 23 (4–35) months after HCT. The regimen of 5 days of clofarabine 40 mg/m ² /d plus 1 day of melphalan 100 mg/m ² appears to be an effective combination worthy of additional study. These clofarabine-containing regimens appear to be relatively well tolerated and show promising antileukemic activity.

Treosulfan-based

Treosulfan, pharmacologically similar to busulfan but more potent in vitro, is a bifunctional, alkylating agent with myeloablative as well as immunosuppressive properties. Unlike busulfan, treosulfan does not appear to require dose adjustment and has an excellent toxicity profile. Furthermore, in contrast to busulfan, treosulfan is active against both primitive and committed hematopoietic progenitor cells. Danylesko and colleagues recently reviewed this agent in HCT. Several interesting studies reported included a prospective, multicenter RIC trial from France that combined treosulfan (12 mg/m ² /d for 3 days) with fludarabine (30 mg/m ² /d for 5 days) and ATG (2.5 mg/kg/d for 2 days) in 56 hematologic malignancy patients. Therapy was well tolerated and median overall survival was not reached at a median follow-up of 13 months, with a 52% 3-year probability of survival.

Nemecek and colleagues undertook a prospective, multicenter trial in 60 patients (n = 44 AML) at high risk for relapse or TRM. Median (range) age was 46 (5–60) years. Therapy consisted of treosulfan 12 to 14 mg/m ² /d for 3 days and fludarabine 30 mg/m ² /d for 5 days, followed by HCT from HLA-identical siblings (n = 30) or unrelated donors (n = 30). All patients engrafted, and the 2-year nonrelapse mortality was 8%. With a median follow-up of 22 months, 2-year relapse-free survival for all patients was 58% and 88% for patients without high-risk cytogenetics. The cumulative incidence of relapse at 2 years was 33%. Newell and colleagues reported the results of a single-institution double UCB HCT protocol in 15 hematologic malignancy patients aged 4 to 63 (median 48) years including 7 AML subjects. Conditioning consisted of a regimen of treosulfan 14 mg/m ² /d for 3 days, fludarabine 30 mg/m ² /d for 5 days, and low-dose TBI 200 cGy. Median time to neutrophil recovery was 22.5 days and only one patient failed to engraft. Treosulfan appears to be an effective and well tolerated, interesting new addition to the HCT armamentarium, although there are no clinical HCT studies directly comparing this agent with busulfan alone or in combination.

Plerixafor and G-CSF

Plerixafor recently has been incorporated into conditioning regimens. The marrow microenvironment contributes to leukemia cell chemoresistance for those cells residing in niches. Preclinical models indicate that inhibition of the chemokine receptor CXCR4 will mobilize leukemia cells into the circulation and facilitate sensitization to cytotoxic therapy. Konopleva and colleagues at the MD Anderson Cancer Center reported in preliminary fashion a phase 1/2 study in which G-CSF and plerixafor therapy were initiated in escalating dose before the start of a standard fludarabine and busulfan regimen. The investigators showed preferential mobilization of clonal leukemia cells over normal cells in this 27-patient study, without toxicities ascribed to the G-CSF/plerixafor treatment. This approach will require validation in a larger series and longer follow-up to determine efficacy.

Selective radiation approaches

While TBI is used to prevent immunologic graft rejection and can eradicate residual malignant cells, toxic effects may be significant, and include cardiac toxicity, pulmonary fibrosis, renal failure and, especially in the pediatric population, an excess number of secondary malignancies. Some investigators have attempted to improve on TBI using helical tomotherapy, a modality that integrates computed tomography image-guided radiotherapy and intensity-modulated radiation therapy into a single device. This technique can deliver conformal targeted radiation to the marrow and lymphatic systems. Rosenthal and colleagues recently described the results of a phase 1/2 investigation using total marrow and lymph node irradiation (TMLI) to augment RIC transplantation for 33 advanced, poor-risk hematologic malignancy patients with a median age of 55 years. The addition of TMLI 1200 cGy (administered as 150 cGy/fraction in 8 fractions over 4 days) to fludarabine 25 mg/m ² /d for 5 days and melphalan 140 mg/m ² /d for 1 day, did not appear to increase toxicity of this chemotherapy preparative regimen. At 1 year after HCT, overall survival, event-free survival, and nonrelapse-related mortality rates were 75%, 65%, and 19%, respectively. The addition of TMLI to RIC is feasible and safe, and the centers equipped with this device can offer it to patients with advanced hematologic malignancies who might not otherwise be candidates for RIC.

Targeted myeloablative radioimmunotherapy

An alternative strategy for acute leukemia patients undergoing HCT is myeloablative radioimmunotherapy (RIT), a technique that targets radiotherapy to the bone marrow using radiolabeled monoclonal antibodies. Candidate isotopes usually are the β-emitter radionuclides ( ¹³¹ I, ⁹⁰ Y, ¹⁸⁸ Re), which are conjugated to monoclonal antibodies including anti-CD33, anti-CD45, and anti-CD66. The cross-firing β particles create a field radiation effect, potentially killing antigen-negative resident marrow tumor cells. Most clinical studies have used ¹³¹ I, a long-lived β-particle emitter, whose γ emissions allow dosimetry studies to be performed easily, but require patient isolation. ⁹⁰ Y is a pure β emitter and the absence of γ emission allows outpatient administration of high doses, but dosimetry calculations are extremely complex. ¹⁸⁸ Re is another radioisotope suitable for biodistribution and dosimetry as it also has γ emission. On the other hand, α particles are helium nuclei emitted from the decay of radioisotopes. RIT using α emitters, such as ²¹³ Bi, ²²⁵ Ac, and ²¹¹ At, should result in less nonspecific toxicity to normal bystander cells and provide a more efficient single-cell killing than β-emitting constructs. Clinical experiences have been very limited.

Although the efficacy of RIT depends on a variety of factors, the most important appears to be biodistribution. Kletting and colleagues described physiologically based pharmacokinetic models to estimate biodistribution and maximize anti-CD45 antibody and anti-CD66 RIT efficacy. These approaches have not yet penetrated clinical practice. On the other hand, Schulz and colleagues recently reported an open-label, single-center pilot study of 30 pediatric and adolescent patients undergoing HCT for malignant (n = 16) and nonmalignant (n = 14) disorders using treatment with a ⁹⁰ Y-labeled anti-CD66 monoclonal antibody. This patient population was considered at high risk for nonrelapse mortality due to advanced disease, ongoing severe infections, second HCT, and significant visceral organ damage. The therapeutic ⁹⁰ Y activity was calculated to deliver a marrow dose of 3000 to 4000 cGy to patients in the malignant group and 1600 to 2000 cGy in the nonmalignant group. Patients also received additional conditioning with either TBI-based or intravenous busulfan, melphalan, and fludarabine-containing therapy. No excess acute organ toxicity was observed in any of the patient groups. Twenty-three patients are alive at a median (range) of 35 (19–71) months after HCT. The overall nonrelapse mortality in this extremely poor-risk group was only 13%, and supports continued exploration of this approach.

Pretargeted myeloablative radioimmunotherapy

While the use of radiolabeled antibodies has shown promise in both imaging and therapeutic applications, the clinical adoption has not fulfilled the original expectations, due either to poor image resolution and contrast in scanning, or to insufficient radiation doses delivered selectively to tumors for therapy. Pretargeting involves the separation of the localization of tumor with an anticancer antibody from the subsequent delivery of the imaging or therapeutic radionuclide.

Pretargeted RIT (PRIT) circumvents the issue of suboptimal therapeutic index (target-to-nontarget ratio) by separating the prolonged-circulating antibody construct (“targeting vehicle”) from the therapeutic radioisotope (“effector”). Conceptually, this sequential administration allows for maximal antibody targeting to take place prior to delivery of the therapeutic radionuclide while maintaining targeting specificity. PRIT substantially reduces whole-body radiation because of radionuclide delivery via small molecules that yield rapid tumor uptake and fast renal excretion of nontumor-bound radioactivity. Synthetic chasers (“clearing agents”) have been introduced as an additional refinement to PRIT. At present, this technique has not yet been implemented in clinical practice but holds considerable promise as a means of enhancing the therapeutic index of RIT.