Significantly increased CD70 up regulation on TEL-AML positive B cell precursor acute lymphoblastic leukemia cells following CD40 stimulation.

BACKGROUND: TEL-AML the most common genetic alteration in childhood precursor B acute lymphoblastic leukemia (BCP-ALL) is associated with a favorable prognosis. PATIENTS AND METHOD: We studied the expression of nerve growth factor/tumor necrosis factor receptor (NGFR/TNFR)/ligand family members on 108 primary BCP-ALL samples by flow cytometry and compared both their baseline expression and CD40-induced modulation on TEL-AML positive and negative leukemia samples. RESULTS: Our findings demonstrate that TEL-AML positive patients exhibit a significantly higher percentage of CD40, CD27 and p75NTR positive blasts at diagnosis. This might well contribute to the improved relapse-free survival of these patients assessed in Kaplan Meier analysis as CD27 and p75NTR directly mediate apoptotic signals. Furthermore CD40 ligation enhances antigen presenting and T cell stimulatory capacity via significant up regulation of CD70 while adequate response to physiological maturation signals as indicated by concomitant down regulation of CD27 is retained in TEL-AML positive leukemia. CONCLUSION: These data provide novel insights in immunological control mechanisms preserved in this leukemia subtype and suggest that not only treatment with chemicals such as HDAC inhibitors but also retained in vivo response to CD40 ligation contributes to improved immune surveillance in these patients which may add to a superior relapse-free survival observed particularly in the presence of other risk factors.

Reduced expression and defective modulation of TNF receptor/ligand family molecules on proB-ALL blasts.

BACKGROUND: There is a subgroup of pediatric patients with an immature immunophenotype of proB-ALL that still poses a therapeutic challenge, even if the overall prognosis in B cell precursor acute lymphoblasic leukemia (BCP-ALL) is very good. Due to impaired treatment response these patients are prone to suffer relapse and are thus by definition stratified into the clinically defined high risk group receiving intensified chemotherapy. Besides response to chemotherapy long term prognosis is also influenced by immunological control mechanisms. Thus, high expression of the TNF receptor CD40 has been shown to prevent particularly late relapse in BCP-ALL suggesting a pivotal role of this regulatory molecule for maintenance of the remission status. PATIENTS AND METHODS: We therefore determined the baseline expression and CD40-mediated modulation of TNF receptor and costimulatory molecules in 5 patients with proB-ALL, 8 with preB-ALL and 22 with c-ALL performing FACS analysis. We particularly compared the TNF receptor status on proB-ALL blasts to the expression on more mature preB- and c-ALL blasts. RESULTS: Here, we demonstrate for the first time a significantly lower baseline expression and CD40-induced modulation capacity of TNF receptor and costimulatory molecules in pediatric proB-ALL compared to more mature precursor B-ALL blasts. CONCLUSION: The lower expression and defective capacity of proB-ALL blasts to respond to CD40 ligand stimulation might resemble the immature feature of these blasts and besides increased chemoresistance contribute to the impaired prognosis of these patients due to escape from apoptosis and immunological control mechanisms.

Herpes simplex virus type-1 amplicon vectors for vaccine generation in acute lymphoblastic leukemia.

For leukemia vaccine generation, high-efficiency gene transfer is required to express immunomodulatory molecules that stimulate potent antileukemic immune responses. In this context, herpes simplex virus type-1 (HSV-1)-derived vectors have proven to be a promising tool for genetic modification of lymphoblastic leukemia cells. Yet, vector-associated viral protein expression might inadvertently modulate vaccine efficacy facilitating both immune evasion and immune stimulation. To explore the issue of immune-stimulation versus immune-suppression in immature lymphoblastic leukemia cells, two types of HSV-1 amplicon vectors, helper virus-dependent and helper virus-free that express the immunomodulatory molecules CD70 and IL-2, were compared with regard to their vector-associated immunomodulatory potential. We first established that lymphoblastic cell lines and primary acute lymphoblastic leukemia (ALL) cells express HSV receptor genes. Lymphoblastic cell lines were transduced with high efficiency, and in primary ALL cells high gene transfer rates of 47+/-15 and 42+/-14% were obtained with helper virus-dependent and -free HSV-1 amplicon vectors, respectively. The efficacy of the two amplicon vectors to induce antineoplastic responses was assessed in a vaccine setting in mice with pre-existing highly malignant lymphoblastic disease. Treatment of mice with vaccine cells transgenically expressing CD70+IL2 significantly suppressed lymphoblastic cell proliferation and improved survival. Of note, when helper virus-dependent HSV-1 amplicon vectors were used for vaccine preparation, the high immunogenic potential of the vector itself, in the absence of transgenic CD70+IL2 expression, seemed to be sufficient to mediate protection comparable to the antineoplastic response achieved by expression of immunomodulatory molecules. Thus for vaccine generation in B lymphoblastic leukemia, the immunogenic potential of HSV-1 helper virus-dependent amplicon vectors does provide additional benefit to the high transduction efficiency of HSV-1-derived vectors.

Characterization of CD34+ progenitor-derived dendritic cells pulsed with tumor cell lysate for a vaccination strategy in children with malignant solid tumors and a poor prognosis.

BACKGROUND: Children and adolescents with primary multifocal, refractory or relapsed malignant extracranial solid tumors still have a poor prognosis inspite of intensive standard radio-/chemotherapy. Here complementary immunomodulatory treatment modalities may prove beneficial as consolidation therapy following cytoreduction. Neuroblastoma, Ewing tumor and soft tissue sarcoma cells have principally been shown to be susceptible towards both cytotoxic and humoral effector mechanisms. Yet in vivo they are not capable of inducing an effective antitumor response which has been attributed to low level MHC expression and lack of costimulatory surface molecules. Professional antigen - presenting cells such as dendritic cells (DCs) in contrast are capable of activating unprimed T cells and are therefore ideal tools for vaccine generation. RESULTS: Here we demonstrate that DCs may be generated from CD34+ progenitor cells to clinical scale in a three to four week cell culture process including an initial expansion and subsequent differentiation and maturation steps. DCs derived from CD34+ progenitors express the expected marker profile and are highly effective in stimulating allogeneic T cell effectors. We also demonstrate that they effectively take up fluorescence-labelled tumor cell lysate. DISCUSSION: Having established a cell culture process for clinical scale DC production utilizing CD34+ progenitors as the cellular source we discuss the role of CD34+ derived DCs in clinical vaccination protocols. The rationale for a phase I/II DC dose escalation study for high risk pediatric patients with extracranial solid tumors assessing safety, immunological and clinical efficacy of serial combined intranodal and subcutaneous injections of tumor cell lysate-pulsed autologous DCs is delineated.