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Abstract 496 UKALL 2003 is an ongoing randomised controlled trial investigating treatment modification according to Minimal Residual Disease (MRD) status at the end of induction and week 11 in children and young adults (up to age 25 years) with ALL. Whilst the randomisation testing the benefit of intensification for an MRD defined high risk group remains open, a treatment reduction randomisation for MRD low risk patients closed in August 2009 after recruiting the target number of patients. Over 2600 patients have so far been enrolled into the trial and we have observed a significant improvement in 5 year event-free (EFS) compared with its predecessor, ALL 97/99, (EFS 87 vs 80%, p Patients in the trial are initially stratified by NCI risk, blast karyotype, and morphological marrow response at day 8/15 of induction for allocation to one of 3 treatment regimens, escalating in intensity (A, B, C). NCI standard risk (SR) patients receive a 3 drug induction (Regimen A) whilst high risk (HR) patients receive 4 drugs including daunorubicin (Regimen B). Patients with a slow early response at day 8 (NCI HR) or day 15 (NCI SR) of induction and patients with poor risk cytogenetics are transferred to Regimen C. Patients without poor risk cytogenetics and who have a rapid early response are eligible for the MRD randomisations. MRD was measured using Real Time Quantitative PCR with a sensitivity of 10−4. Five hundred and twenty-one patients who were either MRD negative at day 29 of induction or positive < 10−4 at day 29 but negative by week 11 entered a randomisation comparing two courses of delayed intensification (standard arm, n = 261) with one (treatment reduction arm, n = 260). Of these patients, 342 (65%) were NCI SR and 179 (35%) NCI HR. With follow-up to October 2009 (median 2 years 9 months), there is no difference in 5 year EFS for patients in the standard (94%) or reduced treatment arms (96%). As shown in table 1, the number of patients with relapse is similar in both arms but there was a non-significant excess of deaths in remission, serious adverse events and grade 3/4 toxicity in recipients of 2 DIs. 7/10 relapses have involved the CNS. The 3 CR deaths in the 2 DI arm were due to gram negative sepsis (DI1), pulmonary haemorrhage (DI2) and RSV pneumonitis (Maintenance, Down). There have been no relapses or CR deaths among 23 MRD low risk patients in the 16 – 25 age group. Table 1 1DI 2DI Statistics for 2DI Obs/N Obs/N Odds ratio(95% CI) 2p Any event 6/260 7/261 1.19 (0.40–3.53) 0.76 Relapse 6/260 4/261 0.69 (0.20–2.38) 0.57 Remission death 0/260 3/261 7.40 (0.77–71.14) 0.08 Grade 3/4 toxicity 177 (68%) 191 (71%) 0.2 SAEs 64 (25%) 77 (29.5%) 0.2 This randomised study provides evidence that treatment can be reduced for around 40% of children and young adults who show rapid clearance of MRD by the end of induction. These patients have an excellent outcome with < 5% risk of relapse on a treatment protocol that does not include cranial radiotherapy or high dose methotrexate and gives minimal exposure to anthracyclines and alkylating agents. Future trials should test the feasibility of further reductions in treatment for this group of patients. Disclosures: No relevant conflicts of interest to declare.

Cytotoxic T lymphocytes (CTL) are important for the recognition and lysis of virally infected cells, but their effectiveness can be limited by viral immune evasion mechanisms. We investigated the immunophenotype and function of human CD8+ T cells raised in response to herpes simplex virus (HSV). The expanded population contained cells of an activated and mature phenotype, as determined by the expressions of CD25, CD45RO, CD57, CD95 and HLA-DR. Cultured cells also expressed CD45RA. These cells lysed autologous and allogeneic HSV-infected lymphoblastoid cell line (LCL) targets via a non-major histocompatibility complex (MHC) restricted recognition pathway. Inhibition assays showed the mechanism of cytotoxicity to be calcium-dependent, granule exocytosis pathway, rather than the internal disintegration pathway. Cold target competition assays indicated that a common CTL population contributed to the recognition of autologous and allogeneic-infected targets. These effectors showed recognition of infected targets which was distinct from that of K562 cells. Non-MHC restricted lysis-associated molecule 2B4 (CD244) was upregulated on culturing and made a significant contribution to lysis of FcγR-bearing targets in a redirected killing assay. These findings suggest that CTL can recognize virally infected cells through a combination of non-MHC restricted mechanisms and may result in more efficient lysis than classical CD8+ T cells.

Hematopoietic chimerism was analyzed in serial bone marrow samples taken from 28 children following T-cell depleted unrelated donor bone marrow transplants (UD BMT) for acute lymphoblastic leukemia (ALL). Chimeric status was determined by polymerase chain reaction (PCR) of simple tandem repeat (STR) sequences (maximal sensitivity, 0.1%). At least two serial samples were examined in 23 patients. Of these, two had evidence of complete donor engraftment at all times and eight showed stable low level mixed chimerism (MC) (1% recipient hematopoiesis). All 10 of these patients remain in remission with a minimum follow-up of 24 months. By contrast, 13 patients demonstrated a progressive return of recipient hematopoiesis. Five of these relapsed (4 to 9 months post BMT), one died of cytomegalovirus pneumonitis and seven remain in remission with a minimum follow-up of 24 months. Five children were excluded from serial analysis as two serial samples were not collected before either relapse (3) or graft rejection (2). We conclude that as with sibling transplants, ex vivo T depleted UD BMT in children with ALL is associated with a high incidence of MC. Stable donor engraftment and low level MC always correlated with continued remission. However, detection of a progressive return of recipient cells did not universally correlate with relapse, but highlighted those patients at greatest risk. Serial chimerism analysis by PCR of STRs provides a rapid and simple screening technique for the detection of relapse and the identification of patients with progressive MC who might benefit from detailed molecular analysis for minimal residual disease following matched volunteer UD BMT for childhood ALL.