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An evaluation of the effects of VP-16 on normal human marrow cells and representative lymphoma-leukemia cell lines was performed to assess this agent's applicability to ex vivo marrow purging. Tumoricidal dose curves were defined using malignant lymphoid (SK-DHL2 and Reh) and myeloid (HL-60) cells admixed with a 20-fold excess of irradiated marrow cells to simulate a borderline remission marrow. One-hour treatments yielded ID50 of less than 5 mumol/L of VP-16 for clonogenic units from each cell line; rare-to-zero clonogenic units survived exposure to 50 to 100 mumol/L. CFU-Mix, BFU-E, and CFU-GM were equal in their sensitivity to VP-16 (ID50s25 to 30 mumol/L). Marrows treated with 75 mumol/L were completely depleted of these colony-forming cells but produced CFU-GM in one-stage long-term marrow cultures (LTMCs). This dose had little adverse effect on the proliferative capacity of marrow stromal progenitors, as measured by CFU-F (ID50 271 mumol/L) and by the unperturbed development of adherent layers in LTMCs. Furthermore, these stromal layers were able to support hematopoiesis as well as controls in co-culture experiments with autologous marrow cells (two-stage LTMCs). In conclusion, doses of VP-16 that cleanse marrow of lymphoma-leukemia cells spare hematopoietic and stromal progenitors as demonstrated by LTMCs. These data favor the use of VP-16 in the clinical autotransplant setting.

3F8 is a murine monoclonal IgG3 antibody specific for the tumor- associated antigen ganglioside GD2. Previous in vitro studies suggest that tumor regressions observed in a phase I clinical trial of 3F8 may be attributable to complement activation by 3F8 and to 3F8-dependent cellular cytotoxicity (ADCC) with lymphocytes. We now describe 3F8- mediated ADCC of GD2-positive tumor targets (melanoma and neuroblastoma) with human granulocytes and report that recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) enhanced this phenomenon. Cytotoxicity required binding of 3F8 to the low-affinity Fc receptor type III (CD16) on the granulocytes and was poor with tumor-binding monoclonal antibodies of other immunoglobulin (ie, non-IgG3) subclasses. GM-CSF (2 to 20 ng/mL) increased ADCC by 93% to 267% at limiting dilutions of 3F8 (1 microgram/mL). With most GD2- positive cell lines tested, this effect translated into a tenfold or greater augmentation in 3F8 efficiency at mediating ADCC. Comparable enhancement occurred whether GM-CSF was present in the ADCC assay or granulocytes were incubated with GM-CSF and washed before the assay. Nonoxidative mechanisms may be important for ADCC since 3F8 mediated ADCC with granulocytes from two children with chronic granulomatous disease; this cytotoxicity was also enhanced by GM-CSF. Since GM-CSF induces a neutrophilia in patients, our data suggest that this cytokine may have the potential of amplifying 3F8 antitumor activity in patients by increasing effector cell numbers and by priming granulocytes for greater cytotoxicity.

A rationally devised induction regimen of vincristine, Adriamycin (Adria Laboratories, Columbus, OH), and sequentially-escalated cyclophosphamide (CPM), followed by S-phase-specific agents (5-fluorouracil [5-FU]/cytosine arabinoside [ara-C]/hydroxyurea), was used in 100 patients with neuroblastoma. Of 17 patients under 1 year of age at diagnosis, complete (CR)/good partial (GPR) responses with long-term disease-free survival were achieved in 11 (85%) of 13 new patients and in two of four previously treated patients; six of the GPRs also received myeloablative therapy with autologous bone marrow rescue to consolidate remission status. The 83 patients over 1 year of age at diagnosis included three groups: (1) 36 new patients whose N4SE included maximal-dose CPM (ie, up to 140 to 160 mg/kg/course); (2) an earlier group of 18 new patients whose N4SE included moderate-dose CPM (ie, up to 80 mg/kg/course); and (3) 29 previously treated patients who all received the maximal-dose N4SE regimen. For new patients, CR/GPR rates were 72% in the maximal-dose group v 39% in the earlier moderate-dose group (P = .029). A CR/GPR rate of 41% and a partial response rate of 17% were observed for the 29 previously treated patients, all but two of whom had large tumor burdens after therapy that included moderate doses of CPM. Despite consolidation with myeloablative therapy, many responders in the three groups ultimately relapsed. The N4SE was strongly myelosuppressive, but only two patients died from associated infection. Extramedullary toxicity was limited to hemorrhagic cystitis in four of 33 CPM previously treated patients; this problem did not occur in the 67 new patients. The data indicate that the maximal-dose N4SE is an effective induction regimen for neuroblastoma, can achieve marked regressions of disease resistant to less intensive therapy, and is sparing of major body organs. This high rate of remission induction must be coupled with improvements in consolidation therapy to assure long-term disease-free survival of poor-risk patients.

We report an unusual case of progressive disseminated neuroblastoma occurring in a child with a family history and stigmata of von Recklinghausen's disease. A review of the literature confirms the extreme rarity of finding these two neurocristopathies in a single individual and thus undermines the widely held notion of an association--genetic or otherwise--between these two entities. We propose that synchronous neuroblastoma and von Recklinghausen's disease is accounted for by chance alone and therefore represents a randomly occurring phenomenon.

A review of the Memorial Sloan-Kettering Cancer Center experience with second malignancies (SM) after childhood Hodgkin's Disease (HD) identified 17 SM in 320 patients who survived more than 1 year from, and were 15 years old or younger at the time of, HD diagnosis (1949 to 1983). Of 254 previously untreated patients, 12 SM were noticed as compared with 0.606 expected on the basis of rates in the general pediatric population (relative risk, 19.8; 95% confidence interval, 10.2 to 34.6). For patients who received multi-agent chemotherapy, the cumulative probability of developing acute nonlymphocytic leukemia (ANLL) or bone sarcoma was 6.2% and 5.5%, respectively, at 10 years from the initiation of therapy; the cumulative risk of all SM in this group reached 18.7% at 15 years. For patients who received radiation alone or with single-agent chemotherapy, the cumulative risk of SM rose from 0% at 10 years and 2% at 15 years, to 10.7% at 25 years from the initiation of treatment. The risk of ANLL after childhood HD was highest in the first 5 to 10 years after combined modality treatment, and aggressive forms of NHL were associated with excessive immunosuppression. Bone sarcomas predominated in solid SM in the first decade after HD treatment, whereas "adult-type" cancers, for example, breast and colon carcinomas, were more delayed. Our findings, supported by a literature review, point to a therapy-related enhanced risk of approximately age-appropriate solid SM. This possibility mandates careful surveillance of long-term survivors of childhood HD.

Neuroblastoma metastatic to the testes is thought to be an extremely rare event and is therefore not routinely considered in the evaluation of neuroblastoma patients. A review of metastatic sites in a series of neuroblastoma patients, however, revealed 11 cases with testicular involvement. Analysis of these cases suggests that the phenomenon may not be uncommon in a subset of neuroblastoma patients, namely, male patients with infra-diaphragmatic primaries and advanced-stage disease. Because the testes constitute a sanctuary site for tumor cells, occult testicular involvement may play a role in the relapse of patients judged to be in complete remission. Accordingly, it is proposed that in the above group of patients special diagnostic attention should be directed at the scrotal contents and, if testicular involvement is detected, orchiectomy or local irradiation should be considered. Cancer 56: 1730-1732, 1985.

The identification of prognostic factors has greatly facilitated the rational choice of therapies in individual patients. Intensive chemotherapy, supplemented with radiation and surgery, has increased the remission rate of patients with widespread disease. The persistence of microscopic foci of malignant cells, however, remains a difficult hurdle for long-term disease-free survival. Highly toxic myeloablative therapies have had at most a modest impact on the overall cure rate of poor-risk patients. The use of novel biological therapies has provided new information on the mechanisms and potentials of immune-mediated tumor cytotoxicity. Timely clinical trials are needed to test their role in adjuvant treatment of occult microscopic disease.

The phenomenon of familial neuroblastoma is discussed in the context of case reports describing disseminated neuroblastoma in two of three half-brothers who share a common unaffected mother and who each have a different father. This family's cytogenetics proved to be unremarkable; also, the mother's peripheral blood DNA did not show tumorigenic capacities in transfection-nude mice experiments. An analysis of reported cases permits an updated examination of the clinical features of this entity and defines the limits of genetic counseling of families of all neuroblastoma patients. Multiple primaries are a hallmark of familial neuroblastoma. Most diagnoses are made in the first 18 months of life and at ages that fall within 12 months of the age of diagnosis of the other affected family member. Difficulties in determining the incidence and penetrance of an inherited susceptibility to neuroblastoma derive from undiagnosed tumors that have undergone regression or spontaneous maturation to benign ganglioneuroma, as well as from early deaths or long-term treatment complications that preclude reproduction and multigenerational pedigrees. Nevertheless, the risk of neuroblastoma in siblings or offspring of the large majority of persons with neuroblastoma appears to be less than 6%. Recent observations concerning chromosomal aberrations and oncogenes in embryonal malignancies are presented in an integrated model of tumorigenesis that corresponds to clinical experience.

The promising antineoplastic agent diaziquone is associated with prolonged aplasia and rare instances of bone marrow necrosis, but only mild extramedullary toxicity. To explore the drug's potential as a myeloablative agent prior to bone marrow transplantation, we compared its effects on hematopoietic versus marrow stromal cells. After short- term (one to six hours) or prolonged (three to seven days) exposure to the drug, marrow was assayed for hematopoietic (CFU-Mix, BFU-E, CFU-GM) and stromal (CFU-F) colony-forming cells and studied in long-term marrow culture (LTMC). One- and three-hour treatments produced little cytotoxicity, even at 5000 ng/mL. After six-hour treatments with this dose, marrow was depleted of CFU-Mix, BFU-E, and CFU-GM, but produced CFU-GM in LTMCs, indicating an ongoing input of CFU-GM from a surviving pre-CFU-Mix population. In contrast, elimination of the latter may be inferred from the absence of CFU-GM in LTMCs exposed for three to seven days to diaziquone at only 150 ng/mL. Under these conditions, CFU-F recovery was 40% and adherent stromal layers in LTMCs were similar to untreated controls regarding rate of development and cellular composition. Our in vitro pre-CFU-Mix-ablative regimen correlates with clinical data that show prolonged but reversible myelosuppression at steady-state diaziquone plasma levels of 101 +/- 10 ng/mL (mean +/- standard error of mean) during 7-day constant infusions. In conclusion: hematopoietic cells are more sensitive than marrow stromal cells to the dose- and highly time-dependent cytotoxicity of diaziquone, a direct drug-induced noxious effect on the marrow microenvironment is an unlikely cause of the isolated episodes of marrow necrosis after the use of diaziquone in vivo, and prolonged infusion of diaziquone represents an attractive means for achieving myeloablation in selected clinical situations.

We studied the ex vivo sensitivity of continuously cultured neuroblastoma cells from 3 different patients towards interleukin-2-induced cell-mediated cytotoxicity. A mean ( ± SD) target cell lysis (4 h 51Cr release) of 49 ± 11, 46 ± 8, and 32 ± 11 % in SMS-SAN, LA-N-1, and SK-N-BE2 cell lines, respectively, was achieved when neuroblastoma cells were co-cultured at an effector-to-target (E:T) ratio of 50:1 with peripheral blood mononuclear cells (PBMC) that had been preincubated for 4 days in the presence of recombinant interleukin-2 (rIL-2; 100 U/ml). Under identical conditions, 93 ± 9 % of Daudi cells (a standard target for rIL-2-activated killer cells) were lysed. Preincubation of rIL-2-induced PBMC cultures in the presence of irradiated neuroblastoma targets (LA-N-1, SK-N-BE2) resulted in a significant cytolytic augmentation. At E:T ratios of 50:1 and 10:1, day-4 rIL-2/LA-N-1-stimulated PBMC produced 69 ± 7 and 41 ± 11% lysis of LA-N-1 cells, as compared to 46 ± 8 and 22 ± (mean ± SD) 7 % lysis by untargeted PBMC that were preincubated with rIL-2 (100 U/ml) in the absence of LA-N-1 target cells (p < 0.05). Co-incubation of rIL-2-induced PBMC preparations with irradiated LA-N-1 and SK-N-BE2 cells, respectively, did not significantly enhance the cytolytic activity against other neuroblastoma targets and the standard Daudi cell line (p ≧ 0.3). In summary, our results suggest that cell-mediated cytotoxicity using rIL-2-activated effector cells may be potentially useful in the immunotherapy of human neuroblastoma.

Although rarely reported, neuroblastoma in monozygotic siblings merits attention because study of its features may aid in elucidating mechanisms of tumorigenesis. We describe disseminated neuroblastoma in one of monozygotic triplets, including both documentation of monozygosity and long-term follow-up of the unaffected co-twins. Information from reports of monozygotic twins concordant and discordant for neuroblastoma reinforces the hypothesis that hereditary factors may be predominant in neuroblastoma diagnosed in infants, whereas nonheritable random mutational genetic events may be more important in neuroblastoma diagnosed after infancy.