Characterization of Immune Reconstitution Following High Dose Chemotherapy in Acute Myeloid Leukemia.

Recovery of immune function after initial treatment of acute myeloid leukemia (AML) is critical, not only for protection against infections but also for surveillance against recurrent disease. A better understanding of the nature of lymphocyte recovery following induction and consolidation therapy for AML could guide the design of immunotherapy strategies aimed at boosting the anti-leukemia activity of a reconstituted immune system. Prior studies examining thymic T cell production following bone marrow transplantation (BMT) have found varying levels of thymic output post-transplant, as measured by T cell Receptor Excision Circle (TREC) levels in the peripheral blood of adult patients. Of note, relapse of chronic myeloid leukemia (CML) following BMT is correlated with decreased levels of TREC positive T cells. In order to characterize immune reconstitution in AML, we studied 26 patients after induction or consolidation time sequential chemotherapy. Their median age was 52 (range 23–69). Thirteen patients received cytarabine, daunorubicin, and etoposide (AcDVP-16) induction therapy, 3 patients received cytarabine, daunorubicin, and cytarabine (AcDAc) consolidation therapy and 10 patients received flavopiridol, cytarabine, and mitoxantrone (FLAM) either as induction or consolidation therapy. Peripheral blood samples were collected approximately every other day for 3–5 time points after each patient’s white blood cell count exceeded 200 cells/cubic mm on three consecutive days. Among the four patients evaluated to date, flow cytometry results show that a majority of cells seen early in immune reconstitution are CD3+ lymphocytes (range 69–97%). Subset analyses on 3 of these 4 patients have shown CD4:CD8 ratios ranging from 3:1 to 4:1, while the fourth patient exhibited an inverse of this ratio at 1:5. In addition, CD25+FOXP3+ T cells represented a median of 5.1% (range 2.5–12.3%) of the CD3+ T cells. Since T cells represented the abundance of cells in the peripheral blood during early bone marrow recovery, we then assessed whether these cells represented recent thymic emigrants or naïve T cells by examining TRECs using real time PCR (RT-PCR). TRECs were present in 24 of the 26 patients with levels ranging between 100 and 100,000 copies per 100,000 cells. Furthermore, 4 control samples from normal volunteers (ages 37–43) revealed the absence of TREC positive cells. Further analyses of these time points and correlations between TREC levels and clinical responses are ongoing.