High-dose chemoradiotherapy (HDC) in the Ewing family of tumors (EFT)

Critical Reviews in Oncology/Hematology 41 (2002) 169– 189 www.elsevier.com/locate/critrevonc

High-dose chemoradiotherapy (HDC) in the Ewing family of tumors (EFT) S. Burdach a,*, H. Ju¨rgens b a

Di6ision of Pediatric Hematology/Oncology and Children’s Cancer Research Center, Martin-Luther-Uni6ersity Halle Wittenberg, 06097 Halle, Germany b Department of Pediatric Hematology/Oncology, Westfa¨lische Wilhelms-Uni6ersity, 48129 Mu¨nster, Germany Accepted 21 December 2000

Contents 1. Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

170

2. Definition of the Ewing family of tumors (EFT) . . . . . . . . . . . . . . . . . . . . . . . . . .

170

3. The clinical biology of EFT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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4. Dose intensity concepts in EFT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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5. Clinical studies of HDC in EFT . . . . . . . . . . . . . . . . . . . . . 5.1. Stratification criteria . . . . . . . . . . . . . . . . . . . . . . . . . 5.1.1. Imaging issues . . . . . . . . . . . . . . . . . . . . . . . . . 5.2. Localized tumors . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.3. Metastatic tumors (lung versus bone/bone marrow) . . . . . . . 5.4. Relapsed tumors . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.5. Definition of risk strata . . . . . . . . . . . . . . . . . . . . . . . 5.6. Evaluation of HDC regimen . . . . . . . . . . . . . . . . . . . . 5.6.1. Single agent HDC. . . . . . . . . . . . . . . . . . . . . . . 5.6.1.1. Total body irradiation (TBI) . . . . . . . . . . . . 5.6.1.2. Single agent chemotherapy . . . . . . . . . . . . . 5.6.2. Combination regimens for HDC . . . . . . . . . . . . . . 5.6.2.1. TBI containing regimens . . . . . . . . . . . . . . 5.6.2.2. Chemotherapeutic regimens (containing no TBI) 5.6.3. Interpretation of clinical studies . . . . . . . . . . . . . . .

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172 172 172 173 174 175 175 175 175 175 175 176 176 178 179

6. Present HDC protocols . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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7. Future perspective . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7.1. Cytotoxic regimens for multifocal bone disease. . . . . . . . . . . . 7.2. Immunologic and genetic strategies exploiting the tumor specificity 7.2.1. Immunotherapy with cytokines and transgenic tumor cells . 7.2.2. Dendritic cells and sensitized T-cells . . . . . . . . . . . . . . 7.2.3. Allogeneic transplants . . . . . . . . . . . . . . . . . . . . . . 7.2.4. Gene repression . . . . . . . . . . . . . . . . . . . . . . . . . .

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180 180 180 180 181 181 182

8. Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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* Corresponding author. Tel.: + 49-345-5572387; fax: + 49-345-5572389. E-mail address: [email protected] (S. Burdach). 1040-8428/02/$ - see front matter © 2002 Elsevier Science Ireland Ltd. All rights reserved. PII: S 1 0 4 0 - 8 4 2 8 ( 0 1 ) 0 0 1 5 4 - 8

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S. Burdach, H. Ju¨ rgens / Critical Re6iews in Oncology/Hematology 41 (2002) 169–189 Reviewers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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Abstract EFT is defined by the expression of ews/ets fusion genes. The type of the fusion transcript impacts on the clinical biology. EFT requires risk adapted treatment. A risk-adapted treatment is determined by tumor localisation, tumor stage and volume. For metastatic and relapsed disease the pattern of spread and the time of relapse are the determinants of risk stratification. Staging of Ewing tumors has been considerably improved by magnetic resonance imaging and modern isotope scanning techniques. However, the determination of the extent of the metastatic spread in particular number of involved bones remains an unresolved issue. The prognosis for high-risk Ewing tumors has been improved by multimodal and high-dose radio/chemotherapy (HDC). The concepts for high-dose therapy in Ewing tumors are based on dose response and dose intensity relationships. In single agent HDC most experience exists with Melphalan. Several chemotherapeutic agents have been used in combination HDC with or without TBI such as Adriamycin, BCNU, Busulphan, Carboplatin, Cyclophosphamide, Etoposide, Melphalan, Thiotepa Procarbazin and Vincristine. Todate, superiority of any high-dose chemotherapy regimen has not been established. However, the clinical biology, the pattern of spread and the time of relapse determine the prognosis of patient who are eligible for HDC. In particular, patients with multifocal bone or bone marrow metastases have a poorer prognosis than patients with lung metastases. In addition, patients with a relapse within 24 months have a poorer prognosis than patients with a relapse later than 24 months after diagnosis. This review will analyze the results of single- and multi-agent chemotherapy with respect to agent combination, dose and risk stratum of patient population. Future therapeutic modalities for the treatment of EFT might encompass immunotherapeutic and genetic strategies including allogeneic stem cell transplantation. © 2002 Elsevier Science Ireland Ltd. All rights reserved. Keywords: Ewing family of tumors; High-dose chemotherapy; Risk adapted treatment; Stratification criteria; Stem cell transplantation; Immunotherapy; TBI

1. Introduction The topic of high-dose chemotherapy HDC in Ewing sarcoma has been excellently reviewed by a number of international experts [1– 6]. The special interest in treating this disease with HDC has produced a wealth of recent studies and publications in addition to novel immunogenetic therapeutic approaches, that justify a present review of the topic. In particular, the explosion of studies in the field of HDC in the Ewing family of tumors (EFT) has lead to the identification of specific risk stratification criteria to overcome the heterogeneity of patients within the conventionally defined clinical stages of localized, metastatic and relapsed disease.

2. Definition of the Ewing family of tumors (EFT) The EFT is defined by the expression of ews/ets fusion genes which confer transformation and transactivation [7,8]. The histogenetic origin of the Ewing tumor stem cell is a matter of debate since the original description of the tumor by James Ewing in the 1920s [9]. Ewing supposed it to be endothelial. More recent experimental evidence suggest that the Ewing tumor cell is capable of neuroectodermal differentiation. The de-

gree of neuroectodermal differentiation has been utilized for histologic subclassification of the EFT into: “ classical Ewing sarcoma; “ atypical Ewing sarcoma; and “ malignant peripheral neuroectodermal tumor (Table 1). In vitro, Ewing tumor cells have shown features of cholinergic neurons by synthesizing acetylcholine and expressing markers for synaptic vesicles [10–12]. The EFT is related to other embryonal and mesenchymal tumors of childhood and adolescence cliniTable 1 Classification of EFT Classification of EFT

Prognostic impact (?)

1. Morphology and differentiation markers Classical ES Atypical ES MPNT

Oa On nb

2. Molecular genetics ews/fli1 Type I (7-6 exon fusion) ews/ets non Type I

O n

a b

O, Improved. n, decreased prognosis.

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cally, morphologically as well as on the molecular basis. The clinical relationship consists of an excellent response to chemotherapy. The molecular relationship consists of the expression of tumor specific proteins due to chimeric oncofusion genes such as ews/ets in EFT and pax/fkhr in alveolar rhabdomyosarcoma. The molecular characterization of these tumors has helped tremendously in differentiating the morphologically related entities of small blue round cell tumors, including Ewing Tumors, soft tissue sarcomas and neuroblastomas and thus allowed the precise definition of the study of population of HDC in EFT. The change in precision of diagnosis by supplementation and possibly replacement of morphologic criteria by molecular definition is to be considered when comparing HDC studies from different historical periods and when analyzing registry data from centers with different diagnostic procedures.

3. The clinical biology of EFT In 95% of EFT reciprocal translocation of the ews gene on 22q12 with e.g. fli1 or erg located on 11q24 or 21q22 is detectable [13]. ews is an ubiquitously expressed gene. fli1 and erg belong to the ets oncogene family and they have DNA binding domains with 98% identity [7], [14]. ets genes are transcription factors [15]. They are involved in regulation of normal development, including embryological and blood cell development [16 –20]. All ets genes are transcriptional activators, repressors have not been found [21,22]. By mapping of chromosomal translocation breakpoints, different ews/fli1 and ews/erg fusion transcripts have been identified. Presently, more than 10 different ews/fli1 and four ews/erg fusion types are known [23–26]. The most frequent formations are ews/fli1 type 1 with ews exon 7 fused to fli1 exon 6 (50%) and type 2 characterized by exons 7– 5 fusion ( 25%) [27]. The product of ews/ets fusion gene is an aberrant transcription factor, that is capable to transform NIH3T3 fibroblasts in vitro [7,8]. Fli1 or erg are deregulated by translocation to the ews promoter. The fusion transcript has a higher transactivating potential than the wild type ets components [21,28]. Recently, association between adenovirus E1A and ews/fli1 was described. After transfection or retroviral infection of human cells esw/fli1 fusion transcript was detected [29]. However, these result were challenged by [30]. The clinical impact of morphologic subtyping [31] has been demonstrated by some studies but the reproducibility of morphologic subtyping in a larger variety of institutions has been challenged by others [32]. Schmidt et al., showed a significant difference in EFS in

171

Ewing’s sarcoma patients compared with MPNT patients when diagnosis of MPNT was based on the presence of Homer-Whright rosettes and/or the expression of at least two neural markers, respectively Ewing’s sarcoma was diagnosed in cases lacking Homer-Wrihgt rosettes and expressing no neural marker or only one in immunohistochemistry [33]. Moreover, using this classification patients with MPNT or Ewing’s sarcoma were also different in tumor site and mean age. Hartman et al., found a poorer prognosis for patients with EFT of an atypical appearance [34]. However, this has not be confirmed by various other authors [35]. More recently, a clinical impact of molecular subclassification based on the fusion type of the chimeric gene has been validated. Amann et al. demonstrated an association of increased neuroglial markers expression with a non-type 1 ews/fli1 gene fusion [36]. Analysis of different ews chimeric transcripts suggested a possible advantage in EFS for patients with localized disease and fusion type 1 transcripts [27]. This assumption was confirmed by De Alava et al., showing the significance of the type 1 fusion transcript to overall survival to all patients regardless of stage (1998). A basis for a molecular explanation of the clinical association of the type 1 fusion with better patient survival may be the finding that the type 1 ews/fli1 fusion encodes a less active chimeric transcription factor than the type 2 fusion [37]. The impact of the detection of the fusion gene product on assessment of minimal residal disease and purging of autologous graft is discussed in chapter 7.3. (allogeneic transplants). Nevertheless, the main risk factor may still be timely diagnosis and appropriate treatment design including quality control, planning of radiation and surgical therapy of the compartment disease as well as appropriate treatment systemic disease component.

4. Dose intensity concepts in EFT In pediatric sarcomas two extremes of contrasting concepts of chemotherapy can be identified. The first therapeutic concept is based on a dose response relation. The second concept is based on duration exposure. The dose response relation concept has been introduced by Vincent de Vita into clinical oncology as dose intensity paradigm [38]. Despite his convincing clinical work, the dose response relationship for tumor cells mainly has been demonstrated for experimental tumors and less convincingly for solid tumors in general and the EFTs in particular. However, in comparison to the epithelial tumors of adulthood the there is more evidence for a dose response relationship in pediatric tumors in general and in pediatric sarcomas in particular. Those response relationship has been demonstrated

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for radiotherapy [39] and for some such as anthracyclins but not all chemotherapeutic agents that are used to treat Ewing Tumors [40]. The duration exposure concept has been based on cell kinetic models [41]. Prolonged therapy (i.e. total duration of therapy in months) has reduced relapses in earlier studies [42,43]. Based on the work of [44] at the Memorial Sloan Kettering Cancer Center, Ju¨ rgens et al., have introduced a prolonged dose intensive combination chemotherapy utilizing intensive polychemotherapy with EVAIA (Etoposide (E) 900 mg/m2, Vincristine (V) 3 mg/m2, Doxorubicin (A) 60 mg/m2, Ifosfamide (I) 1200 mg/m2, Actinomycin D (A) 1.5 mg/m2) in localized tumors in combination with radiotherapy and surgery over 14 months [45,46]. More recently, short term dose very intensive treatment with maximum tolerated dose short of needing PBSC rescue chemotherapy has been advocated. [47] treated 36 patients with a median age of 17 years (range: 1.5–36) with seven courses of chemotherapy with HD-CIV containing 4.2 g/m2 Cyclophosphamide given over 2 days and IFO/VP-16. Total duration of therapy was 6 months; 24 patients had regional tumors. The EFS for these patients is 77% at 2 years (20/24 progression free, two patients relapsed, one died of leukemia without relapse). 5/6 patients with lung metastases survive progression free 7– 36 months from diagnosis. None of the six patients with multifocal bone marrow disease is progression free. [48] confirmed poor results in patients with multifocal bone disease with short term dose intensive treatment with inframyeloablative chemotherapy: 1/4 patients with metastatic lung disease and 0/2 patients with lung plus bone/bone marrow metastases survived after eight cycles VACIME chemotherapy with surgery after cycle 6 and radiotherapy after cycle 8. Total duration of therapy was 6 months. However, there were problems with hematopoietic reconstitution raising the necessity of repeated stem cell infusion support as previously advocated [49,50]. Todate, it has not been clearly clarified beyond doubt whether long and low dose chemotherapy or short and high-dose chemotherapy is superior in which stage of EFT (Table 2). However, in recent studies the dose intensification concept has been preferred in Ewing tumors, in particular for advanced disease. Despite of its toxicity [51] HDC appears more attractive and feasible to many clinicians as compared to extended intermediate dose chemotherapy. Of interest in [52] protocol the cumulative dose of chemotherapy is lower for patients with a higher risk strata than for patients with a lower risk strata. This surprising fact is due to the attractiveness of the concept of dose intensification by high-dose chemotherapy. Lower cumulative dose of chemotherapy may help to reduce second malignancies after Ewing sarcoma [53].

Table 2 Dose intensity concept Intensity

Duration

Authors

Low High (infra-myeloablative)

Longb Long

High (infra-myeloablative)c High (supra-myeloablative)d High (supra-myeloablative) repeated

Shorta Short Short

[42] [45] [44] [47,48] [128,121,131,102] [166,50,49]

Short, 6 months. Long, \12 month. c Inframyeloablative intensity, chemotherapy short of necessitating stem cell rescue. d Supramyeloablative intensity, chemotherapy necessitating stem cell rescue. a

b

5. Clinical studies of HDC in EFT

5.1. Stratification criteria 5.1.1. Imaging issues Diagnostic imaging is not an unresolved issue in patients with Ewing tumors in particular with regard to detection of the number of bone metastases and with regard to response to chemotherapy. Clearly, today magnetic resonance imaging (MRI) is the method of choice for staging the Ewing tumor [54,55] except in the lung where Computed tomography (CT) is superior. Total body MRI reveals additional lesions as compared to conventional imaging in patients with bone metastases [56–58]. STIR sequences are most sensitive for detecting involved compartments [59] but are not very specific [60,61]. In particular, cytokine stimulated marrow recovery has to be differentiated from tumor lesions [62,63]. Nevertheless, if using STIR sequences the pattern symetry can be used to differentiate Ewing tumor lesions from regenerating marrow. However, in systemic Ewing tumor with wide spread marrow disease this might be a problem (Bloem personal communication, [64]. STIR sequences are particularly useful in pretreated patients, who have little or no viable tumor left but do need assessment of the initial extent of disease for planning involved compartment irradiation. However, low specifity of STIR sequences require specific techniques such as fat saturated contrast enhanced T1 weighted [65– 67] or T2 weighted images [68], although some controversy arises from these publications whether contrast enhanced T1 or T2 weighted images are preferable. Nevertheless these techniques can not reliably differentiate viable tumor from reactive tissue [69–71]. Picci et al. have observed within the Italian–Scandinavian-Sarcoma-Study-Group, that the variation of soft tissue component at consecutive CT or MRI correlates best with the histological response [72]. Earlier

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Table 3 EFT staging and stratification criteria

Reports could not demonstrate prediction of chemotherapy response of soft tissue lesions by MRI signal changes [73]. Nevertheless, soft tissue components are not specific for malignancy [74]. In general MRI imaging techniques are very sensitive but specificity is unsatisfactory. Very recently, the use of F-18-flourodeoxy-glucose (FDG)-PET to detect pulmonary metastases has been analyzed. Although spiral CT was superior to FDGPET in the sensitivity of detection of pulmonary metastases, the FDG-PET was useful in follow up of positive lesions [75,76]. In addition FDG-PET is a sensitive and functional test to detect bone metastases because of their increased glucose metabolism [77,76].

5.2. Localized tumors For stratification of risk-adapted treatment it is essential to identify prognostic risk factors. As in many other solid tumors prognosis depends on stage of disease defined as localized, metastatic or relapsed stages of tumor. A number of workshops of the International Society of Pediatric Oncology SIOP (Vienna, 1996) as well of the Euro-E.W.I.N.G. Group have defined several criteria that influence survival rates as there are tumor volume, tumor site, histologic response to treatment, local therapy, age, tumor load, markers such as LDH, site of primary metastasis and time of relapse (Table 3).

In localized tumors, it is generally excepted, that tumor volume is a major prognostic factor. Ju¨ rgens et al. showed the prognostic significance of tumor volume for localized Ewing’ sarcoma [78]. Within the Cooperative Ewing’s Sarcoma Study (CESS of the German Society of Pediatric Oncology, one of the predators of Euro-E.W.I.N.G.) 51 patients were treated from 1981 to 1984. The 3-year disease-free survival rate was 75% for patients with a tumor volume B 100 ml compared to 10% for patients with a tumor ] 100 ml volume. These results were confirmed in consecutive analysis’ of the study [79,80]. The significance of tumor volume as prognostic factor has been shown in different analysis by several authors [3,81]. As a consequence, the CESS 86 regimen was stratified according to tumor volume: Patients with a tumor volume less than 100 ml belonged to the standard risk group, patients with a tumor volume greater than 100 ml to the high risk group. Recent analysis of the results of the CESS 86 demonstrated again the importance of the tumor volume as prognostic factor [82]. However, in addition to the prognostic discrimination obtained by a cut off of 100 ml tumor volume an additional discrimination of prognosis was obtained by using a cut off of 200 ml tumor volume. This may reflect improvement of chemotherapy. Thus, the stratification criteria was shifted toward larger volumes, from 100 to 200 ml tumor volume. The

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8-year EFS rate of patients with tumors \200 ml was 42% compared to 70 and 63% for patients with tumors of 100–200 and \100 ml volume. On the basis of these results, in EE99 the cut point for stratification according to tumor volume is 200 ml. Tumor site as prognostic risk factor has been discussed controversially. McLean showed, that primary site in the pelvis was associated with a poor outcome for patients with no metastatic disease [83]. In the UK Children’s Cancer Study Group (UKCCSG), patients with localized Ewing tumor in the pelvis had the worst prognosis of all patients [84]. Fizazi found by analyzing 182 patients with Ewing tumor that pelvic primary lesion and tumor size were independent prognostic factors for survival [85]. In contrast, the comparison of overall survival in 43 children presenting with metastatic Ewing sarcoma showed no significant differences between pelvic and non-pelvic primaries [86]. Ahrens analyzing the CESS data questioned the influence of pelvic tumor site on prognosis [82]. The inferior outcome for patients with pelvic tumors might be explained by large tumor size with extensive soft tissue invasion and the higher percentage of patients presenting with metastases at diagnoses. However, there is also in primary pelvic tumors a strong influence of tumor volume on prognosis [87]. The mode of local therapy may also impact on outcome since local failures are associated with poor prognosis. Therefore adequate local therapy is essential for long term survival. Local treatment modalities in EFT consist of surgery and/or radiation therapy. Irradiation had been preferred for a long time, although functional result were sometimes poor due to impairment of bone growth by irradiation. The improvement in surgical procedures including endoprothesis made surgery an attractive alternative. In the UKCCSG, local relapse rate was 27% for radiotherapy alone and 11% for surgical resection [84]. In the CESS trials, the local or combined relapse rate after surgery with or without irradiation was 7% compared with 31% after definitive irradiation [88]. In contrast, the frequency of local recurrences was similar after surgery (7%), radiation therapy (7%) and combined surgery and radiotherapy (6%) in the Italian Cooperative Study [89]. Dunst et al., showed that radiotherapy is as effective as surgery if selection of patients with regard to tumor volume is comparable [90]. Very recent data of Dunst et al. show that the presence of perfused necrotic areas at time of diagnosis as assessed by contrast enhanced magnetic resonance imaging is associated with an increased risk of metastases especially in unfavourable pattern of metastatic spread at diagnosis [91]. This finding confirms prior observations using traditional morphologic techniques [92]. Histologic response to preoperative chemotherapy is classified according to the criteria by [93]. As well in

localized [81,89] as in metastatic Ewing tumor [94,87], it proved to be an important clinical predictor for outcome. Based on the results of CESS 81, therapy effect was classified as good (B 10% viable tumor cells in the surgical specimen) or poor response (\ 10% viable tumor cells). However, due to risk-adapted treatment, responsiveness to chemotherapy was no longer a significant prognostic factor in CESS 86 [82]. Other risk factors might be age at diagnosis and serum lactic dehydrogenase (sLDH). sLDH levels before treatment was of prognostic significance in an analysis of 618 patients with Ewing’s tumors of the extremities. The time to relapse was significantly shorter for patients with elevated sLDH than in patients with normal values [95]. In the UKCCSG study, patients with extremely elevated levels of sLDH had a poorer prognosis [84]. High sLDH levels may reflect high tumor volume and tumor lysis in turn may be in consequence of tumor necrosis. The negative impact of tumor necrosis on prognosis of Ewing tumor has very recently been demonstrated [91]. In the Italian Cooperative Study age less or equal 14 years affected the outcome of patients with localized tumors positively [89]. In the UKCCSG/MRC studies better results were obtained for patients younger than 15 years [84]. Delepine showed a significantly positive prognostic role in univariate analysis for patients younger than 18 years compared with patients older than 18 years [96]. [97] identified age as a major determinant of local tumor extension (1999). Age was also a significant risk factor for patients with primary multifocal bone disease [49].

5.3. Metastatic tumors (lung 6ersus bone/bone marrow) The prognosis for patients with metastases at the time of diagnosis is clearly worse than for patients with localized Ewing tumors. Moreover, and at least of similar importance, prognosis depends on site of metastases. In the German/European Cooperative Intergroup Ewing’s Sarcoma Studies the 5 year EFS for patients with primary lung metastases was 35%, for patients with primary extrapulmonary metastases less than 20% [98] This analysis includes patients who have received HDC. In addition, earlier reports about patients who have not received HDC also report event free survival between 30 and 37% for patients with lung metastases alone [99,100]. Earlier dated studies from 1984, an era before HDC in EFT show an even more dismal prognosis. Patients with primary lung metastasis had a prognosis of 50% event free survival and 0% event free survival for patients with primary bone metastases [101]. This difference may possibly be due to the introduction of high-dose chemotherapy and stem cell transplantation for patients with primary extrapulmonary metastases.

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Number of involved bones and age were additional risk factors for patients with primary multifocal bone disease or early relapse [49]. Patients with multifocal bone or bone marrow disease have the worst prognosis [102,47].

5.4. Relapsed tumors The results of treatment after relapse are mostly dismal [103], although some controversy exists. Early data from Memphis show a 60% overall survival [41]. The European data show uniformly poor survival [101,104]. These conflicting data may be explained by distinct prognosis for patients with early and late relapse. In clinical terms, it has been useful to divide a relapsed tumor into early relapse within 2 years of diagnosis and late relapse after 2 years of diagnosis. Data from the European Intergroup (EICESS, a more recent predator of Euro-E.W.I.N.G.) show, that patients with relapsed Ewing tumors benefit from highdose therapy by improving their prognosis from 7% without high-dose therapy to 19% with high-dose therapy. This difference is almost entirely due to the benefit in the group of patients with early relapse (B 2 years after diagnosis) who increase their prognosis from 2% without chemotherapy to 17% whereas in patients with late relapse (\2 years after diagnosis) there is no significant difference between high-dose therapy and conventional therapy [104]. In fact, data again from the European Intergroup (EICESS) show that patients with early relapse had a poorer prognosis before stem cell transplantation was in use.

5.5. Definition of risk strata The discussion of the stratification criteria demonstrates that the current staging into localized versus metastatic disease does not preclude considerable clinical heterogeneity of tumors in these two categories. Even within the group of localized tumors there is a considerable heterogeneity depending on tumor localization [86,84,85,83,82] and tumor volume [78,80,81]. Axial tumors and most importantly pelvis tumors were found to have a poorer prognosis than limb tumors. However, localization may be associated with tumor volume. This heterogeneity applies also for metastatic disease which sometimes has also been termed as stage IV disease (ISG/SSG IV protocol 1999), although stage II and stage III in the Ewing family of tumors have not been well defined. Nevertheless, metastatic disease is heterogeneous depending on localization and number of metastases. In the majority of studies specifically addressing this issue, lung disease has a better prognosis than bone and/or bone marrow disease. In addition, the number of bone metastases is also of prognostic relevance [49] and

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has recently also been used as a stratification criteria [105]. In addition, histogenetic differentiation has been used as a stratification criterion although problems exist with methodology and reproducibility of differentiation assessment [36]. Recently, a molecular classification has been found to be more useful for risk stratification. The fusion transcript type has been utilized to predict prognoses [27,106,36]. In addition, prognostic impact has been attributed to the p53 mutation status [107,108]. The chemotherapy response has an impact on a prognosis [93] and has been utilized for clinical stratification. In addition to chemotherapy response, many necroses at time of diagnoses have been found to have an impact on pattern of metastatic spread [92,91]. Tumors with necrosis at time of diagnosis may be particular genetic entity with a distinct clinical biology because they grow faster than their vascular supply. Based on the described inadequacies of the current staging system with its confined differentiation between localized and metastatic disease the European Ewing Tumor Working Initiative of National Groups (EuroE.W.I.N.G.) has developed a new risk stratification system which is implemented in the EE99 protocol (Figs. 1 and 2).

5.6. E6aluation of HDC regimen In the ensuing analysis we have first assessed phase I/II studies with single agents including TBI as single agent. Next we have assessed the results of combination chemotherapy regimens with and without TBI. In this analysis we have specifically analyzed the stages and risk factors defined above. HDC studies in patients with measurable disease have been very valuable since they have assessed the efficacy of various HDC regimens in this disease. Of note, patients not responding to conventional chemotherapy can not be rescued by HDC. The place of HDC is in the treatment of patients responding to conventional chemotherapy being at high risk for relapse because of minimal residual disease

5.6.1. Single agent HDC 5.6.1.1. Total body irradiation (TBI). The employment of TBI to control systemic Ewing tumors was originally reported in the early 1980s [109]. They reported an objective response rate of 50% with a 33% survival at 4–27 months. TBI has been pursued by various groups in vitro and in the clinic [110–112] demonstrating that it is relatively ineffective when the tumor burden is high. This finding was expected given that 48–54 Gy are required to eliminate overt EFT. 5.6.1.2. Single agent chemotherapy. The efficacy of highdose single chemotherapy with single agents has been

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Table 4 Agents with proven activity in Phase II studies Agent

Dosage

Melphalan

180–215 120–210 120–125 155–160 375–625

VP 16

mg/m2 mg/m2 mg/m2 mg/m2 mg/m2

1800 mg/m2 Busulfan Thiotepa Cyclophosphamide

900 mg/m2 7 g/m2

Patient-no

Response rate

Authors

3 7 2 4 3 7 15 3 3 4

0.66 0.71 0.50 0.50 0.33 0.28 0.20 0.66 0.66 0.50

[113] [114] [115] [116] [117] [42] [118] Burdach, unpublished [120] [122]

reviewed by Craft in 1987. In spite of repeated recommendations for further work in phase I and phase II studies [102] and randomized cooperative studies [58] little additional information has been gained since then (Table 4). The most extensive experience with high-dose single agents exists with Melphalan. Although high-dose Melphalan did not provide cure in the early studies, it demonstrated activity against advanced disease. Additional studies assessed the efficacy of high-dose Melphalan with a valuable disease [113– 116] with a total response rate of 10 out of 16 patients with three complete remissions and six partial remissions. In addition, two patients were treated with minimal residual disease as defined at that time, one of two stayed in continuos remission at least for 4 years. With Etoposide, three different studies show six out of 25 responses in patients with a evaluable disease [42,117,118]. Busulfan as a single drug has not been widely assessed in EFT tumor in spite of its wide spread use in high-dose conditioning regimens. Research in www.ncbi.nlm.nih.go6 utilizing the terms B Busulfan AND Ewing \ as of November 20, 2000 reveals thirteen publications, none of them assessing busulfan as a single drug. Chemotherapy combinations including Busulfan will be discussed below. Limited data show that Busulfan can be active as a single drug in patients with multiple relapsed Ewing tumors, partial remissions were observed of two in three patients refractory to chemotherapy after multiple relapse (Burdach, unpublished observation). However, preclinical data did not demonstrate activity of Busulfan in peripheral neuroectodermal tumors [119]. High-dose Thiotepa activity was demonstrated in producing partial remission in 2/3 patients with Ewing sarcoma refractory to conventional chemotherapy doses in a phase II study [120]. An earlier report from this group demonstrated one partial and one minor response in two patients. [121]. Two out of four patients with refractory EFT sar-

coma responded to high-dose Cyclophosphamide (7g/ m2) with one complete response [122]. Conflicting results have been reported regarding the activity of single agent Cis-Platinum in Ewing tumors [123,124]. However, Platinum containing regimens such as PAI have been utilized by the Scandinavian Sarcoma Group in Ewing tumors [125]. Agents that are not active as single drugs include Carboplatinum (Pediatric Oncology Group, unpublished data).

5.6.2. Combination regimens for HDC 5.6.2.1. TBI containing regimens. Most of the high-dose combination chemotherapeutic regimens used for treatment of the advanced Ewing tumors did contain total body irradiation or derivatives. The first encouraging reports from the 1980s [126,127] show responses that have fostered future clinical studies. Miser et al., used 8 Gy total body irradiation and combination chemotherapy of Cyclophosphamide, 1200 mg/m2 per day times two, Adriamycin 35 mg/m2 per day times two and Vincristine 2 mg/m2. His study population has included patients with localized Ewing tumors of the central axis and patients with metastasized Ewing sarcomas. The location of metastases was not specified. Event free survival rate was 50% at 30 months after autologous bone marrow transplantation (BMT) [128,129]. Horowitz et al., have reported the results of studies pursuing Miser’s concept of total body irradiation with a total dose of 8 Gy in combination with Adriamycin 70 mg/m2, Cyclophosphamide 2400 g/m2 and Vincristine 2 mg/m2. One subject of the study were patients with metastatic Ewing sarcoma or rhabdomyosarcoma. Five of 31 patients (15%) survived 6 years after diagnosis with no difference in outcome between Ewing sarcoma and rhabdomyosarcoma. Only patients who received a complete remission were eligible for this study. Metastatic site was not specified [130].

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Emminger et al. have treated nine patients with the EFT with 12 Gy fractionated total body irradiation and Melphalan 120–140 mg/m2 plus Etoposide 60 mg/kg 9 Carboplatin 1.5 g/m2. Five of the nine patients had bone metastases or multifocal bone disease at diagnosis and two of the nine patients had a pulmonary relapse. They had three of nine patients in complete remission with observation times from nine to 46 month after BMT. Two of six patients without Carboplatin (29 and 46 month) and one of three patients treated with Carboplatin (9 month after BMT) stayed in remission [131]. These results have been further pursued in a cooperative trial utilizing the same program in a larger group of 17 patients with extremely poor prognosis, i.e. primary multifocal bone disease or early relapse [102]. Nine of 17 patients had multifocal bone disease or bone marrow involvement. Eight patients had early relapse. Seven of 17 patients relapsed and two of 17 patients died of complications. Event free survival at 5 year after initial diagnosis was 45%. In a comparison to a matched cohort comprising 41 patients with the same extent of disease event free survival was 2% (40 of 41 patients relapsed). The recent follow up shows the results of this protocol with 36 patients. 17 of these 36 patients had multifocal primary Ewing tumors, 18 of these 36 had early, multiple or multifocal relapse and one patient had a unifocal late relapse. Nine of these 36 patients died of treatment related toxicity, 18 of 36 patients suffered relapse. Three patients developed secondary malignancy. Event free survival for the whole group is 21%. Patients with allogeneic transplantation had no better outcome (20%) than patients with autologous transplants (21%). The median observation time was 89 months, range 56– 139 from diagnosis and 81 months, range 48– 132 after transplantation. Thus, late relapse and development of secondary malignancy decreased event free survival. Particular to this condition and regimen was the use of involved compartment irradiation for such bones, that were involved in the disease as assessed by Tc scan, MRI and histology. In addition, 24 out of 36 patients were transplanted in complete remission (CR), whereas 12 out of 36 were transplanted in partial remission (PR). Seven of 19 patients had early relapse, three of 19 patients had late but multifocal relapse and two of 19 patients had two relapses before stem cell transplantation. 17 of 36 patients had multifocal primary disease. Of the total group, 27 of 36 patients (75%) presented with bone metastases either at primary diagnosis or at relapse. Nine out of 27 patients had one bone with metastases, ten out of 27 had 2–5 bones with metastases and eight of 27 patients had more than five metastatic bones. Parts of this concept, i.e. to use Melphalan plus TBI for consolidation of patients with primary metastatic bone disease have been pursued by other groups.

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Meyers et al. and the Children’s Cancer Group have reported 16% event free survival at 2 years in 24 patients with bone or bone marrow metastases using a regimen of induction chemotherapy followed by Melphalan, Etoposide and TBI with peripheral blood stem cell support. Two patients died of toxicity [132]. Although the authors conclude, that this strategy did not improve prognosis for this group of patients, it is to be considered that the results for this group of patients reported without HDC consolidation is generally B5% [104,101]. Martinez-Aguillo et al. have treated six patients with either metastatic or relapse or refractory disease with high-dose Melphalan with TBI as a consolidation treatment. One of six patients is in CR at 28 months after transplant [133]. Most recently the concept of total body irradiation and involved compartment irradiation combined with high-dose chemotherapy was modified by using total marrow irradiation instead of whole body irradiation. A recent study from Seattle compared combination chemotherapy Busulfan 12 mg/kg, Melphalan 100 mg/ m2 and Thiotepa 500 mg/m2 9 total marrow irradiation (TMI) following recovery from combination chemotherapy [134]. The median total marrow irradiation dose was 12 Gy, TMI was derived by a modification of TMI employing lung and liver shielding. Only those patients were eligible for TMI that had not a prior dose limiting of radiotherapy that would cause excession of normal organ tolerance. A total of 16 patients were treated in this study, seven patients were excluded from TMI because of extensive prior radiation therapy, inadequate peripheral blood stem cell harvest, early disease progression or patient refusal. The disease status prior to myeloablative therapy was CR1 in three, CR2 in nine and second PR in one, CR3 in one and progressive disease in two patients. Three of 16 patients died of toxicity, eight of 16 died of relapse. Six of 16 patients are alive without evidence of disease with a median observation time between 27 and 66 months. All surviving patients received total marrow irradiation. The event free survival at 3 years is 36%. One of the six survivors had multifocal disease at relapse. One of the surviving patients had lung metastases at diagnosis and a local relapse that lead to transplantation and one patient had initial regional lymph node metastases. All other surviving patients had localized relapsed. Taeb et al have reviewed the Florida College of Medicine experience in 30 patients with metastatic Ewing tumors with 2 TBI containing regimens: Vincristine, Adriamycine and Cyclophosphamid or Etoposide and Cyclophosphamid. No indication of metastatic organ site was given for this population. A total of 19 patients died, two from transplant related complications, 17 patients relapsed, 11 patients remained alive — 1–19 years (overall survival: 37%). Of the surviving patients, six had lung metastases, all survivors were under 18

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years of age. No patient with bone marrow metastases survived more than 14 months [135]. [136] compared the European bone marrow transplant registry data for patients with Ewing tumors who received first or second complete remission high-dose chemoradiotherapy reported by 21 European transplant centers in eight European countries after a median follow up time of 4-years (range 1 month to 10 years) since megatherapy. 32 patients had metastatic disease at diagnosis with 21 patients having metastases to the bone or the bone marrow. 4/4 patients with bone metastases had an event free survival of 5 years after transplantation. In this analysis, patients with isolated lung metastases did not have a better outcome than patients with primary metastases to the bone or the bone marrow. However, a follow up of the registry data in 1999 did show a significantly poorer outcome in metastatic patients with non lung metastases as compared to lung metastases [137]. The number of involved bones was not assessed in the registry. 31 of 62 patients grafted in second remission, which had event free survival, had localized disease at diagnosis. Of interest in this study, the time to relapse with a cut off point of 2 years did not influence prognosis. The high-dose chemoradiotherapy regimens were grouped into six categories, four without TBI, two with TBI. There was no advantage of a particular regimen. However, patients treated with Busulfan containing regimens fared better than patients with TBI containing regimens. Of particular interest, four out of four patients treated for metastatic bone disease with Busulfan containing regimens survived with an observation time from 15 to 36 month, median 22 month after megatherapy. In this registry data, only one out of ten patients treated with TBI containing regimens survived 5 years after megatherapy. Thus, future long term follow up of patients with bone metastases treated with high-dose Busulfan containing regimen is of great interest. Interpretation of registry data appears difficult since risk strata such as metastatic site and time to relapse maybe modulated by different chemotherapeutic regimens. Thus, it needs to be kept in mind when comparing different high-dose chemoradiotherapy regimens, that the clinical biology of the tumor before transplant may already been affected by induction chemotherapy.

5.6.2.2. Chemotherapeutic regimens (containing no TBI). [138,121] treated 32 children (1–16 years, median 10 years) for metastatic Ewing sarcoma with various highdose combination therapeutic regimens. 22 children had metastases at diagnosis, 12 patients had relapse. Metastatic location was lung, bone or bone marrow, the proportion of patients with lung metastases versus bone/bone marrow metastases was not specified as well as time of relapse. 14 patients were treated with measurable disease with 18 courses of high-dose chemother-

apy. Eight courses with Melphalan9 BCNU and Procarbazin resulted in one CR, two PR, three minor responses and two non-responders. Eight courses with Busulfan and Cyclophosphamide and/or Melphalan resulted in three CR, four PR and one minor response. Two courses with Thiotepa resulted in one partial response and one minor response. 18 patients were treated in complete remission, 14 of these 18 patients received a combination of BCNU Procarbazine and Melphalan. 14 have received two high-dose courses and five of 14 have received one high-dose course with BCNU, Procarbacin and Melphalan. Four of 18 patients in complete remission have received two different high-dose chemotherapy courses, the first consisting of BCNU and Melphalan and the second consisting of Busulfan and Cyclophosphamide. One of 14 patients with measurable disease was alive 6 years after grafting and four of 18 patients were treated in complete remission are alive. The probability of event free survival for patients grafted in remission is 18% for the median observation time of 3 years. [139] have treated 18 patients with Ewing sarcoma with high-dose Busulfan and Melphalan. The median age at diagnosis was 14.2 years (range 2.75–30 years). Eleven patients had metastatic disease, ten of the 11 had pulmonary metastases and one had bone marrow involvement. Six of the seven non-metastatic patients had \ 100 ml primary tumors and the seventh patient was transplanted in second complete remission. Time between diagnosis and relapse in this patient was not specified. Twelve patients were in CR1, four in CR2, one in PR and one in progressive disease at time of high-dose chemotherapy. With a median follow up of only 2 years after high-dose therapy (range 2 month to 7 years), 13 of 18 patients are surviving in complete remission. Six of the 11 patients with metastatic disease are surviving. Among the survivors are five with lung disease and one with bone marrow disease at time of diagnosis. Five of 18 patients have relapsed and one of 18 patients succumbed to early death due to CMV pneumonitis. One additional patient is surviving after pulmonary relapse in rescue treatment with lobectomy and unilateral lung radiation 18 months post high-dose therapy. [140] have treated 15 patients with EFT, 11 of 15 patients were in CR 1, two of 15 patients were in CR 2 and two were in PR. Stage of disease (localized versus metastasis) was not specified. Actuarial EFS is 64% at 4 years after transplantation with a median observation time of 12 months (range 1–46 months). Pession et al. have performed a phase one study with a combination of Busulfan, Etoposide and Thiotepa in three patients with Ewing tumors, two patients in CR2 and one patient in partial remission. Patient eligibility was age younger than 17 years and metastatic or relapsed tumors. Site of metastatic disease or time of

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relapse was not specified. One of three patients is surviving with no evidence of disease 5 years after transplant. This patient was in partial remission at time of transplant, two other patients relapsed [141]. A re-analysis in 1999 of the EBMT registry data of 289 patients grafted for Ewing tumors revealed some new findings. Some patients reported in the mono-institutional European studies may also have been reported to the registry. 28 patients were grafted with local disease because of tumor volume and/or poor response. 138 patients had primary metastatic disease and 123 patients had relapse. The 5 year overall survival was 30% for the 289 patients with a median follow up of 6.5 years after BMT. The overall survival was better in patients with localized disease (n= 28, 60%) than in patients with metastatic primary disease (n = 128, 30%). Of interest in this analysis is, that patients who received Busulfan containing regimens did better (n =46, 60%) versus patients who received Busulfan not containing regimens (n = 120, 26%). This difference was mainly due to an improvement of outcome in localized disease where 16 Busulfan receiving patients had a 5 year overall survival of 75% whereas twelve patient receiving other regimens had a 38% overall survival. Patients with lung metastases did not fare significantly better with Busulfan (n= 12, 66%) as compared to non Busulfan containing regimens including irradiation (n= 15, 39%). In addition, in patients with metastases other than lung the difference between Busulfan receiving patients (n=18, 44%) and those receiving other regimens (n= 93, 23%) did not quite reach statistical significance. This study did not specify the location of metastases other than lung. In addition, it did not specify number of involved bones in patients with bone metastasis. Again no superiority of Busulfan was found in patients with relapse. In contrast to a previous registry report [136], metastatic site other than lung was an adverse prognostic factor [137]. [142] have reviewed the Children%s Hospital of Los Angeles experience in 17 patients with pediatric metastatic sarcomas utilizing high-dose chemotherapy with Melphalan (210 mg/m2), Carboplatinum (1200– 1700 mg/m2) and Etoposide (640– 1600 mg/m2). Amongst these patients were nine with metastatic Ewing tumors. 10 of the 17 patients had bone, bone marrow or combined bone/bone marrow metastases. 17 patients had lung metastases. The progression free survival at 3 years was 44912% for transplanted patients as compared to 189 8% for a control group (n =13). In this analysis longer survival was associated with transplantation and absence of bone metastases [142].

5.6.3. Interpretation of clinical studies A comparison of different high-dose chemotherapy regimens containing combinations of high-dose chemotherapeutic agents with or without total body

179

irradiation is difficult, given the clinical heterogeneity of patient groups and regimens. In these studies patients with localized and metastatic tumors have been treated as well as patients with relapsed tumors. The clinical stratification criteria such as tumor volume, metastatic location, number of bone metastases respectively number of involved bones and time of relapse after diagnosis have not been addressed uniformly in these studies. In contrast the aforementioned criterion of remission status has been universally applied. From a comparison of these studies the following can be concluded: In patients with disease refractory to conventional chemotherapy, cure can not be achieved by HDC be it with or without total body irradiation. HDC is a consolidation treatment. The irradiation of all involved compartment in multifocal bone disease and multi-compartment disease in conjunction with combination HDC seems to produce the only long term survivals in the setting of multifocal bone disease or early relapse. This is the patient group with the poorest prognosis. Busulfan appears to be a promising chemotherapeutic agent for the treatment of localized and lung and possibly bone disease although the efficacy of Busulfan as a single agent in Ewing tumors has not been demonstrated well so far. It has been argued that drugs are not effective as single agents may display an effect in combination chemotherapy, although the experimental evidence for this concept is lacking so far. This is mainly due to the lack of experimental systems. Systems in use to test the control of the human tumor (Ewing family of tumor) in mice so far had involved xenografting in pharmacological immunosupressed mice [143,144]. The engraftment of Ewing tumor cells in NOD-SCID mice has only recently been achieved [145,146]. This experimental progress may open new avenues for rational design of HDC regimens.

6. Present HDC protocols Present studies have specified the indications for high-dose chemoradiotherapy in patients with advanced Ewing tumors. Here we will discuss some of the current protocols in high-dose chemotherapy protocols in use for the Ewing family of tumors (Table 5). The European Intergroup study EURO-E.W.I.N.G. 99 defined three risk groups (Fig. 1). Risk group one (R1) comprised patients with tumors B 200 ml and patients with a good histological response at time of surgery in a neoadjuvant setting. This European protocol sees no role for high-dose radiochemotherapy in risk group one. Risk group two (R2) comprises patients with a histological poor response, patients with tumors \ 200 ml and patients with lung metastases. In R2, high-dose Busulfan/Melphalan is compared in a randomized fashion to seven courses of chemotherapy with

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Vincristine, Actinomycin and Ifosfamide plus lung irradiation. Risk group three (R3) comprises patients with metastases to bone and/or bone marrow. In R3 one course of Busulfan/Melphalan high-dose chemotherapy is offered alternatively to the Meta-EICESS regimen which comprises pretransplant involved compartment irradiation and tandem high-dose chemotherapy with Melphalan and Etoposide (Fig. 2). The latter protocol has evolved from the studies utilizing total body or total marrow irradiation [128,102] whereas the Busulfan/Melphalan protocol has mainly been derived from work of the Villejuif group utilizing Busulfan for advanced Ewing tumors [121] as well as from the analysis of the EBMT registry data. The Italian–Scandinavian ESG/SSG IV protocol addresses metastatic patients only if they have lung and/or pleural metastases and only one bone metastasis. It utilizes high-dose Busulfan/Melphalan after eight cycles of induction chemotherapy with Vincristine, Adriamycin, Ifosfamide, Cyclophosphamide, Etoposid and uses total lung irradiation ten weeks after high-dose chemotherapy. The Fred Hutchinson Cancer Research Center assesses the role of a dual transplant approach comprising Busulfan and Melphalan for the first graft and total marrow irradiation before the second graft. This study is open for patients with relapsed EFT and has two objectives: (1) estimate the maximum tolerated dose of total bone marrow irradiation (TMI) that can be administered as planned consolidation utilizing autologous peripheral blood stem cell graft following local radiotherapy and prior high-dose therapy with Busulfan/Melphalan and Thiotepa; and (2) examine the efficacy of this dual transplant approach for high risk patients with EFT with recurrent or metastatic disease. The Memorial Sloan Kettering Cancer Center pursues high-dose short term chemotherapy short of necessitating stem cell rescue utilizing Cyclophosphamide at a total dose of 4.2 g/m2 given on day one and day two, Doxorubicin total dose of 75 mg/m2 given from day one to day three, Vincristine total dose of 2 mg/m2 given from day one to day three, alternating with a total dose of Ifosfamide of 9 g/m2 and VP16 at a total dose of 0.5 g/m2 each given from day one to day five.

7. Future perspective See Table 6

7.1. Cytotoxic regimens for multifocal bone disease Future studies need to address the role of total body irradiation and total marrow irradiation in comparison to Busulfan in preventing relapse in multifocal bone disease. In addition, the role of pregraft involved compartment irradiation needs to be assessed. The term pre-involved compartment irradiation describes irradiation of all involved bones in multifocal bone disease. This may end up to irradiating more than 50% of the bone marrow volume necessitating stem cell support [56,58,50]. Future cytotoxic strategies for advanced Ewing tumors could utilize regimens for advanced Ewing tumors consisting of: (1) induction; (2) local intensification; and (3) systemic intensification. The induction chemotherapy may consist of two cycles of chemotherapy for in vivo purging and two cycles of chemotherapy followed by stem cell apheresis. The local intensification will consist of two cycles of chemotherapy simultaneously targeting the involved compartments with radiotherapy. The systemic intensification will utilize repeated two high-dose chemotherapy or chemo, followed by a TBI or TMI-regimen. A prospective study design is necessary to assess the role of the involved compartment irradiation as compared to solely irradiating the primary tumor. These essentials have been implemented in the Meta-EICESS protocol. In addition, pharmacological gene therapy [147–149] and pharmaco immunogene therapy [150,151] may help to control multimetastatic or refractory disease.

7.2. Immunologic and genetic strategies exploiting the tumor specificity of fusion genes 7.2.1. Immunotherapy with cytokines and transgenic tumor cells Several cytokines can enhance immune response against tumor cells and their systemic administration has been shown to produce clinical tumor responses. However, application is often limited by toxic side

Table 5 Phase III studies for patients with bone/bone marrow metastases or early relapse Patient number

Age

Observation time

Results

Authors

5 14 21 9 32 36

n.i. 6–31 years 12 years (0–21) 10–19 years n.i.a 16 years (4–30)

24 days–29 months 49 months (4 months–4 years) 4 years (3–106 months) 12–62 months 2 years 5 year (minimum) (5–15)

2/5 CR 6/14 CR 7/21 CR 2/9 CR 0.16 EFS 0.21 EFS

[131] [102] [136] [134] [132] [49]

a

n.i., not indicated.

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Fig. 1. The European Intergroup study EURO-E.W.I.N.G. 99 defines three risk groups.

effects. Gene modified tumor cells can produce locally high amounts of cytokines and induce immunity without significant systemic toxicity. As has been demonstrated in animal models, this anti-tumor response is directed not only against the gene modified cells itself but also against pre-existing tumor cells. A multitude of cytokines including IL1, IL2, IL4, IL6, INFg, INFa, TNFa and G-CSF have been transfected into tumor cells and gene modified tumor cells have been administered as tumor vaccines. There is some evidence that the Ewing tumor family is a suitable target for immunotherapy. TNF alpha and IF gamma, which can be stimulated by IL2, induce a synergistic antiproliferative response in human Ewing tumor cells in vitro [152]. IL2-induced killer cells showed lytic activity against Ewing sarcoma cells in culture [153]. In children with Ewing tumors, systemic IL2-application after stem cell transplantation led to increased transplantation-related activation of the immune system on cellular and humoral levels [154] and increased EFS [49]. Intratumoral injection of transgenic IL2-secreting fibroblasts in a child with a peripheral neuroectodermal malignancy induced a CD3+ / CD56 + lymphocyte population, cytokine-induced killer cells [155]. Immunotherapy with transgenic tumor cells has been

assessed in neuroblastoma. Immunotherapy with transgenic tumor cells has been undertaken with allogeneic [156] and autologous tumors [157]. At the University of Halle Medical Center presently one protocol is under IRB for patients with HLA 2 and A24, utilizing allogeneic tumor cell lines expressing IL2 as a transgene.

7.2.2. Dendritic cells and sensitized T-cells The utilization of tumor specific translocations in pediatric sarcomas for immunotherapy has been pursued in the United States [158] and in Europe [150,151]. In mice it has been demonstrated that the PAX-3FKHR fusion product of alveolar rhabdomyosarcoma is capable of generating immune responses [158]. EFT specific peptide has been designed that bind to specific HLA haplotypes [151]. Presently NCI protocols 97-C0050 and 97-C-0052 are pursuing this approach ([email protected]). A European protocol is presently under IRB review ([email protected]). 7.2.3. Allogeneic transplants Allogeneic transplants in advanced EFT tumors have been pursued for a number of reasons among which the following two are most prominent: Number one is the

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variable contamination rate (10– 90%) of grafts with tumor cells [159–163]. The second reason is a possible graft versus tumor effect of allogeneic transplant. Results of allogeneic transplant as compared to autologous transplant are difficult to assess because of the problems with patient heterogeneity outlined previously in this review. In our analysis, we did not find a superiority of outcome with allogeneic transplant which may be due to high transplant related mortality of 40% in the allogeneic transplant group [49]. However, an earlier EBMT registry analysis reports an improved outcome with 50% survival with allogeneic bone marrow transplantation [164]. A decrease of allogeneic transplant related mortality utilized less toxic conditioning regimens may improve the outcome of allogeneic transplants.

7.2.4. Gene repression Ewing tumor specific fusion gene products have also been targeted by genetic repression therapy utilizing ribozymes [165]. In a distinct approach p53 deficiency is utilized by an E1B deleted replication competent adanovirus (ONYX 015) in the MCI-T99-0021 study chaired by Galanis. Although p53 mutation are rare in a primary patient population with primary Ewing tumor they are higher in cell lines derived from patients with Ewing tumors at the Mayo Clinic [107,108] which might indicate that p53 serve as a marker of poor prognosis. In addition, this finding points out the very selective and unusual nature of existing Ewing tumor

cell lines, which are mostly derived from metastatic patients and may not necessarily represent the full spektrum of the clinical biology of Ewing tumors.

8. Summary Ewing tumors can be defined on a molecular basis of ews/ets fusion genes. Todate, the role of dose intensity versus cumulative chemotherapy dose with prolonged duration of therapy has not been clarified beyond doubt. However, most investigators feel that a higher dose intensity might be beneficial for patients with advanced disease. That preference effects, that higher dose intensity often leads to lower cumulative doses of chemotherapy in patients with a higher risk. The controversy about high-dose chemotherapy in patients with Ewing tumors has previously not focused on this issue, but on issues such as which chemotherapeutic agents to use or whether to use total body irradiation or Busulfan. Stratification of the disease is extremely important to judge about the efficacy of therapy in EFT specifically the efficacy of high-dose therapy. Stratification criteria in HDC studies in EFT must include the following localized versus metastatic versus relapse. In localized primary tumors the volume and response to chemotherapy is to be assessed, in metastatic patients the site and number of metastases is to be differentiated, patients with lung metastases have to be differentiated from patients with bone/bone marrow

Fig. 2. In R3 one course of Busulfan/Melphalan high-dose chemotherapy is offered alternatively to the Meta-EICESS regimen which comprises pretransplant involved compartment irradiation and tandem high-dose chemotherapy with Melphalan and Etoposide.

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Table 6 Future directions Therapeutic mode

Protocol address

Mitomycin, D and Cisplatin plus ONYX-015 for metastatic sarcoma V,D, C and Dexrazoxan with and without ImmTher a meromyl dipaptide Tumor-specific peptide vaccination and IL-2 with and without autologous T-Cell Transplantation in Sarcomas (97-C-0050) Autologous T-Cell transplantation with vaccine driven expansion of anti-tumor effectors (97-C-0052) C, D, V, E and I [email protected]

Mayo Clinic Cancer Center, Rochester, Minnesota 55905, Evanthea Galanis USA: [email protected] Memorial Sloan Kettering Cancer Center, New York 10021, Paul A. Meyers USA: [email protected] National Cancer Institute, 10 Cloister Court, Bethesda, Chrystal Mackall Maryland 20892, USA: [email protected]

V, D, C plus E for metastatic EFT Busulfan, Thiotepa and Melphalan followed by ASCT+TMI for high risk EFT EVAIA for metastatic EFT Double ME with ASCT for metastatic EFT (Meta-EICESS) Vaccination with allogenic IL2 transgenic Ewing tumor cells for advanced high-risk EFT

National Cancer Institute, 9000 Rockmill Pike, Bethesda, Maryland 20892-4754, USA: [email protected] Memorial Sloan Kettering Cancer Center, New York 10021, USA: [email protected] Memorial Sloan Kettering Cancer Center, New York 10021, USA: [email protected] Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA: [email protected] Children’s Hospital, University of Mu¨ nster, 48129 Mu¨ nster, Germany: [email protected] Children’s Hospital Medical Center, University of Halle, 09097 Halle, Germany: [email protected] Children’s Hospital Medical Center, University of Halle, 06097 Halle, Germany: [email protected]

Principal investigator

Chrystal Mackall Brian H. Kushner Mark Lawrence Bernstein Jean E. Sanders Heribert Ju¨ rgens Stefan Burdach Stefan Burdach

V, Vincristine; D, Doxonubicin; C, Cyclophosphamide; I, Ifosfamide; E, Etoposide.

metastases. In patients with bone metastasis the number of involved bones is to be enumerated. In patients with relapsed disease the time of relapse is to be considered as well as the response to chemotherapy. A risk stratification of Ewing tumors based on these stratification criteria defines the clinical heterogeneity of Ewing tumors that is not described by the current staging system differentiating between localized metastatic and relapsed disease. Taking these risk stratification into consideration, there is no role for high-dose chemotherapy in patients with small localized tumors (B 100 ml) who have a good chemotherapeutic response (R1 stratum in the EE99 protocol). High-dose chemotherapy needs to be assessed in patients with large volume localized disease (\ 200 ml) and in patients with poor response to induction chemotherapy as well as in patients with primary lung metastasis (R2 stratum in EE 99). In patients with primary multifocal bone disease and/or bone marrow involvement, high-dose chemoradiotherapy studies need to address the following issues: 1. What is the role of involved compartment irradiation in multifocal bone disease? 2. What is the role of total body irradiation or total marrow irradiation in multifocal bone disease? 3. What is the role of Busulfan in controlling multifocal bone disease? In addition, new therapeutic agents targeting the bone including pharmaco-gene therapy approaches are needed for this group of patients. Future developments of therapy for patients with advanced Ewing tumors

will clarify also the role of immunotherapeutic and genetic approaches including transgenic tumor cells, fusion gene product presenting dendritic cell, immunization of T-cells, genetic repression strategy and role of allogeneic transplantation in this disease. In general, the assessment of the efficacy of high-dose chemoradiotherapy in EFT has to take into consideration that the prognostic factors depend on the pretreatment before grafting and that not only the substances but the scheduling and the cumulative doses of induction chemotherapeutic regimen may modulate the clinical biology and the prognostic risk factors in EFT.

Reviewers Prof. Dr. Ju¨ rgen Dunst, Dept. of Radiation Oncology, Martin-Luther University Halle-Wittenberg, D06097 Halle, Germany. Prof. Alan W. Craft, Head, Dept. of Child Health, The Royal Victoria Infirmary, Queen Victoria Road, Newcastle upon Tyne, NE1 4LP, U.K. Dr. Patrice Viens, Department of Medical Oncology, Institute Paoli Calmettes 232, Blvd Sainte Marguerite, B.P. 156, F-13273 Marseille Cedex 9, France.

Acknowledgements ‘Deutsche Krebshilfe’ and ‘Elterninitiative Kinderkrebsklinik e.V.’. The expert help of Almut

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Meyer-Bahlburg, in editing of the manuscript in preparation of the figures and tables as well as the helpful comments of Professor Ulrich Go¨ bel and Dr Wolfgang Hirsch are greatly appreciated. In addition the word processing and the bibliographic work assistance of Peggy Patzner is acknowledged.

[19]

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Biographies Stefan Burdach, Professor Dr med, born April 1952, Berlin, Germany, one son. Education and Training: Friedrich-Wuheim College, Cologne, 1971; AlbertusMagnus-University, Cologne, 1980, MD; HeinrichHeine-University, Du¨ sseldorf, 1989, Ph.D. Resident and Fellow Children’s Hospital Medical Center, Cologne, 1978–1983; Research Fellow, Stanford University School of Medicine, 1984–1987; Primary Care Physician, Fred Hutchinson Cancer Research Center, Seattle, 1986; Attending Physician Children’s Hospital Medical Center, Du¨ sseldorf 1988–1998. Appointments: Associate Professor 1995, Full Professor 1998 Current position: Chairman, Department of Pediatrics and Pediatric Hematology/Oncology; Director, Cell and Gene Therapy Program; Children’s Hospital, Martin-LutherUniversity Halle-Wittenberg, Halle, Germany. Author and co-author of over 120 original papers and 19 book chapters. Memberships: American Society of Hematology, American Association for the Advancement of

S. Burdach, H. Ju¨ rgens / Critical Re6iews in Oncology/Hematology 41 (2002) 169–189

Science, International Society of Pediatric Oncology, International Society of Experimental Hematology, European Society of Paediatric Haematology and Immunology, German Society of Paediatric Haematology and Oncology. Herbert Ju¨ rgens, Professor Dr med, born July 1949, Bonn, Germany, married to Christine, two sons, two daughters. Education: Medical School Du¨ sseldorf, Glasgow, 1968– 1974; board examination 1974;doctorate 1976; habilitation 1983. Medical residency, 1975–1976; Paediatric training Du¨ sseldorf, 1976–1981; Special Fellow Paediatric Oncology, MSKCC, New York, 1978–1979. Appointments: Associate Professor 1989, Full Professor 1991. Current posi-

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tion: Director Department of Paediatric Haematology/Oncology, University Children’s Hospital, Mu¨ nster, Germany. Author and co-author of over 250 original papers and over 40 book chapters. Memberships: German Society of Paediatrics; German Society of Paediatric Oncology and Haematology, President, chairman Co-operative Ewing’s Sarcoma Study; Cancer Society North Rhine Westphalia, board member; German Cancer Society, Vice President 1996– 1998; International Society of Paediatric Oncology, member 1990–1996 and chairman 1994– 1996 Scientific Committee, board member 1994– 1996; American Society of Clinical Oncology; European Musculo-Skeletal Society, board member 1995–1999.