Addition of muramyl tripeptide to chemotherapy for patients with newly diagnosed metastatic osteosarcoma

A report from the Children's Oncology Group

Abstract

BACKGROUND:

The addition of liposomal muramyl tripeptide phosphatidylethanolamine (MTP-PE) to chemotherapy has been shown to improve overall survival in patients with nonmetastatic osteosarcoma (OS). The authors report the results of addition of liposomal MTP-PE to chemotherapy for patients with metastatic OS.

METHODS:

Intergroup-0133 was a prospective randomized phase 3 trial for the treatment of newly diagnosed patients with OS. The authors compared 3-drug chemotherapy with cisplatin, doxorubicin, and high-dose methotrexate (Regimen A) to the same 3 drugs with the addition of ifosfamide (Regimen B). The addition of liposomal MTP-PE to chemotherapy was evaluated.

RESULTS:

Five-year event-free survival (EFS) for patients who received liposomal MTP-PE (n = 46) was 42% versus 26% for those who did not (n = 45) (relative risk for liposomal MTP-PE, 0.72; P = .23; 95% confidence interval [CI], 0.42-1.2). The 5-year overall survival for patients who received MTP-PE versus no MTP-PE was 53% and 40%, respectively (relative risk for liposomal MTP-PE, 0.72; P = 0.27; 95% CI, 0.40-1.3). The comparison of Regimen A with Regimen B did not suggest a difference for EFS (35% vs 34%, respectively; relative risk for Regimen B, 1.07; P = .79; 95% CI, 0.62-1.8) or overall survival (52% vs 43%, respectively; relative risk for Regimen B, 1.1, P = .75; 95% CI, 0.61-2.0).

CONCLUSIONS:

When the metastatic cohort was considered in isolation, the addition of liposomal MTP-PE to chemotherapy did not achieve a statistically significant improvement in outcome. However, the pattern of outcome is similar to the pattern in nonmetastatic patients. Cancer 2009. © 2009 American Cancer Society.

The prognosis for patients with metastatic osteosarcoma (OS) remains poor despite aggressive multimodality therapy. In contrast to those with nonmetastatic disease, who have a 5-year survival of approximately 70% to 75%, survival for metastatic patients is significantly lower.1-8 Surgical resectability, the number of metastases at diagnosis, and site(s) of metastases remain important determinants of long-term survival.3, 5, 9 Several approaches using conventional cytotoxic therapies have been used in an effort to enhance survival for patients who present with metastatic disease, but they have done little to improve the outcome of patients with metastatic OS in the last 30 years. Most recently, an upfront window trial of topotecan in newly diagnosed patients with metastatic OS reported only 1 partial response and 1 clinical response (decreased radiotracer uptake on bone scan and decrease in pain) in a cohort of 27 evaluable patients.1 New approaches are needed.

Muramyl tripeptide phosphatidylethanolamine (MTP-PE) is a nonspecific immune modulator that is a synthetic analogue of a component of bacterial cell walls. Incorporation of MTP-PE into liposomes has allowed targeted delivery of MTP-PE to monocytes and macrophages in areas such as the lungs.10 Liposomal MTP-PE is able to activate these cells to become tumoricidal. Preclinical studies have already confirmed the antitumor effects of liposomal MTP-PE in rodent and canine OS models.10-21 In addition, it has been shown that concurrent administration of cytotoxic chemotherapy does not interfere with the antitumor effects of liposomal MTP-PE.17

From November 1993 through November 1997 the Children's Cancer Group and the Pediatric Oncology Group carried out Intergroup Study 0133. This was a prospective, randomized phase 3 trial of treatment of newly diagnosed OS in patients ≤30 years old. The study posed 2 questions in a 2-by-2 factorial design (Fig. 1). First, there was a randomization to a 3-drug chemotherapy regimen (doxorubicin, cisplatin, and high-dose methotrexate) or a 4-drug regimen using these agents with ifosfamide. The second was a randomization to receive or not to receive liposomal MTP-PE in addition to the randomly assigned chemotherapy. Outcome measures considered were event-free survival (EFS) or overall survival. The results of this trial for patients with nonmetastatic OS demonstrated a survival advantage (statistically significant improvement in overall survival, and a trend toward improved EFS) for patients receiving liposomal MTP-PE.22 We report the results for patients with metastatic disease enrolled in Intergroup Study 0133.

Figure 1. A treatment schema is shown. HDMTX indicates high-dose methotrexate; MTP, muramyl tripeptide phosphatidylethanolamine. *Resection of metastatic lesions, when possible, occurred after definitive surgery.

MATERIALS AND METHODS

Patients

Patients enrolled in the study had histologically confirmed high-grade intramedullary OS. Metastatic patients were defined to be those patients in whom at least 1 site of metastasis had been reported at the time of enrollment and before any therapeutic measures. Histologic confirmation of metastasis was not required. Estimate of disease resectability was determined at study enrollment and was not subsequently revised after induction chemotherapy. Patients with clinically detectable metastatic disease were eligible at Children's Cancer Group institutions, but ineligible at Pediatric Oncology Group institutions. Detailed eligibility requirements have previously been described.23

Treatment

Treatment details have also been previously described (Fig. 1).23 Briefly, patients were assigned randomly to 1 of 4 regimens. There were 2 chemotherapy arms, Regimens A (cisplatin, doxorubicin, and high-dose methotrexate) and B (ifosfamide, doxorubicin, high-dose methotrexate, and cisplatin), and within those, patients were assigned randomly to receive liposomal MTP-PE or not. Although liposomal MTP-PE treatment did not begin until Week 12 of protocol therapy, randomization of treatment assignment was done at entry. This resulted in 4 treatment arms: A or B for chemotherapy, both with and without liposomal MTP-PE.

Both regimens called for an initial period of chemotherapy, designated as induction, that lasted 10 weeks, followed by definitive resection of the primary tumor. Maintenance was scheduled to begin at Week 12, but did not begin until the surgeon determined that wound healing was adequate. Maintenance continued until Week 31 in Regimen A and Week 38 in Regimen B.

Half the patients were assigned randomly at entry to receive liposomal MTP-PE beginning at Week 12 of protocol therapy. It was administered at a dose of 2 mg/m². Liposomal MTP-PE was administered twice weekly for 12 weeks beginning at Week 12, and weekly for an additional 24 weeks beginning at week 24. MTP-PE was not interrupted for delays in chemotherapy.

Approval from the institutional review board (IRB) was required at every institution before enrollment. Informed consent was obtained from all patients or their guardians, and the appropriate IRB-approved written informed consent was signed.

Statistical Methods

Patient status current to August 31, 2005 was considered in the intent-to-treat analysis. EFS is defined to be the time from study entry to progression of disease, diagnosis of second malignancy, death, or last follow-up, whichever occurred first. Local progression at the end of induction chemotherapy is specifically excluded as an event, because we intended all patients in the study to undergo surgery whenever possible. In prior Children's Oncology Group studies, patients with apparent radiographic tumor progression after induction therapy often had significant necrosis observed in the primary tumor after resection. For this reason, we did not consider apparent radiographic progression after induction to be an event; such patients remained in the study. Patients who experienced disease progression, diagnosis of second malignancy, or death were considered to have suffered an analytic event. Otherwise, the patient was censored at last follow-up.

Overall survival is defined to be the time from study entry to death or last follow-up, whichever occurred first. Patients who died were considered to have experienced an event. Otherwise, the patient was censored at last follow-up.

The EFS and overall survival functions were estimated by the method of Kaplan and Meier.24 The statistical significance of the comparisons of risk for adverse event was assessed by means of the log-rank test,24 stratified by factors on which the randomization was balanced, namely, lactate dehydrogenase (LDH) at study enrollment. Relative risks and associated confidence intervals were estimated using a relative hazards model with the characteristic of interest as the only variable in the model.24

Interaction between assigned chemotherapy and assigned biological agent was assessed using the relative hazards regression. The hazard of an event (either analytic-event or death) was modeled as:

• 

The following terms were included in the regression model: 1) c, chemotherapy, coded as 1 if the patient was assigned Regimen B and 0 otherwise; 2) m, biological agent, coded as 1 if the patient was assigned to receive liposomal MTP-PE and 0 otherwise; and 3) interaction, coded as the product of c and m, that is, 1 if the patient received both Regimen B and liposomal MTP-PE and 0 otherwise. A P value associated with the test of hypothesis β₁₂ = 0 of .10 or less was considered evidence of a significant interaction. If there was no evidence of interaction, the effect associated with liposomal MTP-PE was estimated by performing a stratified log-rank test, with chemotherapy randomization as the stratification factor.24

The prognostic significance of selected patient characteristics determined at study enrollment was assessed using a relative hazards model with the characteristic of interest as the only variable in the model.24 To explore the joint relationships between therapy assignment, patient characteristics, and outcome, factors considered significantly related to outcome as single characteristics were incorporated into a relative risk regression model along with the randomized therapeutic assignment. Backward stepwise regression was used to evaluate whether therapeutic assignment was significantly related to outcome after adjustment for those previously identified important risk factors.

RESULTS

Patient Characteristics

Of the 777 eligible patients enrolled on Intergroup Study 0133, 91 were found to have metastatic disease at initial diagnosis and are the subjects for this report. Characteristics determined at enrollment for these patients are described in Table 1. Median time of follow-up for patients who had not experienced an adverse analytic event was 89 months (range, 1-141 months).

Table 1. Patient Characteristics

	No. of Patients	%
ULN indicates upper limit of normal. * The ULN for this analysis was the ULN in the institution where the patient was treated.
Sex
Male	56	62
Female	35	38
Age at enrollment, y
0-11	25	28
12-14	24	26
15-30	42	46
Race
White	54	59
Hispanic	18	20
Black	13	14
Asian	1	1
Other	5	6
Primary tumor site
Femur	59	65
Below knee	16	18
Humerus	7	8
Pelvis	5	5
Other axial	3	3
Not determined	1	1
Alkaline phosphatase*
<ULN	43	47
≥ULN	48	53
Lactate dehydrogenase
<ULN	38	42
≥ULN	53	58
Lung involvement
Right lung only	7	8
Left lung only	6	7
Both lungs	30	36
Involved, but laterality not specified	13	15
No lung involvement	28	33
Not reported	7
Metastatic bone involvement
Yes	20	24
No	62	76
Not reported	9
Soft tissue involvement
Yes	21	24
No	68	76
Not reported	2

Patient Outcomes: EFS

As of August 2005, the 5-year EFS for the entire cohort of 91 patients was 34% (95% confidence interval [CI], 24%-45%). When analyzed according to chemotherapy regimen, the 5-year EFS for each of the regimens was as follows: 1) Regimen A without MTP-PE = 29% (95% CI, 11%-51%); 2) Regimen A with MTP-PE = 41% (95% CI, 21%-60%); 3) Regimen B without MTP-PE = 23% (95% CI, 8%-43%); and 4) Regimen B with MTP-PE = 44% (95% CI, 23%-64%). There was no statistical difference among the regimens (log-rank = 0.55; Fig. 2A). By using the statistical test described above for risk for adverse analytic event, there was no evidence of an interaction between chemotherapy and liposomal MTP-PE assignment (P = .20).

Figure 2. (A) Event-free survival (EFS) by is shown by treatment regimen assignment. (B) EFS is shown by liposomal muramyl tripeptide (MTP) assignment. (C) EFS is shown by chemotherapy regimen assignment.

The relative risk for adverse analytic event associated with randomization to receive liposomal MTP-PE was 0.72 (P = .23; 95% CI, 0.42-1.2; Table 2). The EFS at 5 years was 42% for those randomized to receive MTP-PE versus 26% for those who were not (Fig. 2B). The relative risk for adverse analytic event associated with randomization to receive chemotherapy Regimen B was 1.07 (P = .79; 95% CI, 0.62-1.8; Table 2). The EFS at 5 years was 34% for those randomized to 4-drug chemotherapy versus 35% for those randomized to 3-drug chemotherapy (Fig. 2C).

Table 2. Relative Risks Based on Treatment Regimens

	Relative Risk of Death	Relative Risk of Adverse Event
L-MTP-PE indicates liposomal muramyl tripeptide phosphatidyl ethanolamine.
L-MTP-PE assigned (n=46)	0.72	0.72
No L-MTP-PE assigned (n=45)	1.0	1.0
P	.27	.23
Regimen A (n=43)	1.0	1
Regimen B (n=48)	1.1	1.07
P	.75	.79

Local progression was specifically excluded as an event for this analysis, as we intended all patients enrolled on study to undergo surgery. Only 2 patients in this current cohort had radiographic evidence of local progression at the end of induction chemotherapy; 1 patient had a concurrent new site of metastatic disease and was deemed a treatment failure, another patient was lost to follow-up and removed from the protocol.

Patient Outcomes: Overall Survival

As of August 2005, the 5-year overall survival for the entire cohort of 91 patients was 47% (95% CI, 35%-58%). When analyzed according to chemotherapy regimen, the 5-year overall survival for each of the chemotherapy groups were as follows: 1) Regimen A without MTP-PE = 53% (95% CI, 28%-73%); 2) Regimen A with MTP-PE = 50% (95% CI, 26%-69%); 3) Regimen B without MTP-PE = 30% (95% CI, 13%-50%); and 4) Regimen B with MTP-PE = 57% (95% CI, 33%-75%). There was no statistical difference among the regimens (log-rank = 0.60; Fig. 3A). By using the statistical test described above for risk of adverse analytic event, there was no evidence of an interaction between chemotherapy and liposomal MTP-PE assignment (P = .39).

Figure 3. (A) Overall survival is shown by treatment regimen assignment. (B) Overall survival is shown by liposomal muramyl tripeptide (MTP) assignment. (C) Overall survival is shown by chemotherapy regimen assignment.

The relative risk for death associated with randomization to receive liposomal MTP-PE was 0.72 (P = .27; 95% CI, 0.40-1.3; Table 2). The survival at 5 years was 53% for those randomized to receive MTP-PE versus 40% for those who were not (Fig. 3B). The relative risk for death associated with randomization to receive chemotherapy Regimen B was 1.1 (P = .75; 95% CI, 0.61-2.0; Table 2). The survival at 5-years was 43% for those randomized to ifosfamide versus 52% for those who were not (Fig. 3C).

Impact of Clinical Factors on Survival

To explore joint relationships between therapy assignment and these prognostic factors, we performed an analysis of a relative risk regression model including these prognostic factors and therapy assignment. The relative risk for EFS and survival in this backward stepwise regression analysis was not different from the univariate analysis of the relative risks of therapy assignment alone (Table 3).

Table 3. Final Relative Risk Regression Incorporating Prognostic Factors*

	Relative Risk	P	95% CI
CI indicates confidence interval; L-MTP-PE, liposomal muramyl tripeptide phosphatidylethanolamine. * Adjusting for sex (male vs female), race (white vs nonwhite), and serum alkaline phosphatase at enrollment (at or below upper limit of normal vs above upper limit of normal). † Risk relative to patients randomized to receive chemotherapy A. ‡ Risk relative to patients randomized not to receive L-MTP-PE.
Event-free survival
Regimen B†	1.3	.42	0.71-2.3
L-MTP-PE‡	0.67	.13	0.37-1.13
Overall survival
Regimen B†	0.98	.94	0.53-1.8
L-MTP-PE‡	0.77	.41	0.43-1.41

Several clinical factors correlated with worse EFS and overall survival in this cohort of patients. Male patients, patients with high LDH, patients with high alkaline phosphatase (AP), patients with metastatic bone involvement (either alone or in combination with other sites of metastatic disease), and non-Caucasian patients had consistently worse outcome. We identified sex, race, and baseline AP as the strongest predictors of outcome for the patients who presented with metastatic disease (Tables 4-6).

Table 4. Impact of Clinical Factors on Event-Free Survival

Clinical Characteristic	No. of Patients (Events)	Relative Risk of Event	95% CI	P
CI indicates confidence interval; ULN, upper limit of normal. * Maximal tumor size determined by computed tomography or magnetic resonance imaging. † Log-rank trend test.
Sex
Male	56 (42)	1.00	—	.001
Female	35 (14)	0.38	0.21-0.70
Age at diagnosis, y
0-11	25 (12)	1.00	—	.512
12-14	24 (14)	1.39	0.64-3.01
15+	42 (30)	1.48	0.76-2.88
Race
White	54 (33)	1.00	—	.0025
Hispanic	18 (9)	3.05	1.55-6.00
Black	13 (12)	1.06	0.51-2.22
Other	6 (2)	0.47	0.11-1.96
Tumor site
Distal extremity	16 (8)	1.00	—	.423
Proximal extremity	66 (41)	1.41	0.66-3.00
Axial skeleton	8 (6)	2.01	0.70-5.81
Not determined	1 (1)	—	—
Maximum tumor size, cm*
<7	10 (4)	1.00	—	.074†
≥7 but <9	12 (7)	1.03	0.41-2.57
≥9 but <11	20 (13)	1.17	0.54-2.51
≥11	37 (23)	1.33	0.67-2.64
Not determined	12 (9)	—	—
Lactate dehydrogenase
<ULN	38 (18)	1.00	—	.0011
≥ULN	53 (38)	2.50	1.42-4.40
Alkaline phosphatase
<ULN	43 (17)	1.00	—	<.0001
≥ULN	48 (39)	4.39	2.44-7.91
Lung involvement
Unilateral	31 (19)	1.00	—	.702
Bilateral	35 (20)	0.88	0.47-1.66
No. of lung nodules
1	18 (11)	1.00	—	.460
2	13 (9)	1.22	0.50-2.95
3+	35 (19)	0.79	0.38-1.67
Site of metastasis
Lung only	44 (24)	1.00	—	.0147
Bone only	6 (6)	3.49	1.40-8.67
Lung + bone only	8 (6)	2.62	1.07-6.46
Soft tissue + other	21 (14)	1.47	0.76-2.86
Unknown	12 (6)	—	—

Table 5. Impact of Clinical Factors on Overall Survival

Clinical Characteristic	No. of Patients (Events)	Relative Risk of Death	95% CI	P
CI indicates confidence interval; ULN, upper limit of normal. * Maximal tumor size determined by computed tomography or magnetic resonance imaging. † Log-rank trend test.
Sex
Male	56 (35)	1.00	—	.0072
Female	35 (11)	0.41	0.21-0.80
Age at diagnosis, y
0-11	25 (9)	1.00	—	.359
12-14	24 (13)	1.82	0.78-4.26
15+	42 (24)	1.58	0.73-3.41
Race
White	54 (27)	1.00	—	.0166
Hispanic	18 (8)	3.22	1.46-7.10
Black	13 (9)	1.39	0.63-3.09
Other	6 (2)	0.69	0.16-2.90
Tumor site
Distal extremity	16 (8)	1.00	—	.678
Proximal extremity	66 (32)	1.08	0.50-2.35
Axial skeleton	8 (5)	1.60	0.52-4.90
Not determined	1 (1)	—	—
Maximal tumor size, cm*
<7	10 (3)	1.00	—	.160†
≥7 but <9	12 (6)	1.02	0.38-2.77
≥9 but <11	20 (9)	0.86	0.36-2.08
≥11	37 (20)	1.16	0.55-2.41
Not determined	12 (8)	—	—
Lactate dehydrogenase
Below ULN	38 (14)	1.00	—	.0008
At or above ULN	53 (32)	2.84	1.51-5.35
Alkaline phosphatase
Below ULN	43 (11)	1.00	—	<.0001
At or above ULN	48 (35)	4.91	2.47-9.77
Lung involvement
Unilateral	31 (15)	1.00	—	.742
Bilateral	35 (15)	0.89	0.43-1.82
No. of lung nodules
1	18 (9)	1.00	—	.287
2	13 (8)	1.48	0.57-3.85
3+	35 (13)	0.68	0.29-1.59
Site of Metastasis
Lung only	44 (16)	1.00	—	.0059
Bone only	6 (6)	3.49	1.40-8.67
Lung + bone only	8 (5)	2.62	1.07-6.46
Soft tissue + other	21 (14)	1.47	0.76-2.86
Unknown	12 (5)	—	—

Table 6. Characteristics of Patients Among the Treatment Arms

Regimen	A	A + MTP	B	B + MTP
MTP indicates liposomal muramyl tripeptide phosphatidylethanolamine; ULN, upper limit of normal.
Patients, No.	21	22	24	24
Age, y
0-11	8	7	5	5
12-14	2	5	7	10
15-30	11	10	12	9
Sex
Male	12	11	17	19
Female	9	11	7	8
Race
White	9	11	18	16
Hispanic	4	4	5	5
Black	4	6	0	3
Other	4	1	1	0
Lactate dehydrogenase
≤ULN	8	10	10	10
>ULN	13	12	14	14
Alkaline phosphatase
≤ULN	9	10	11	13
>ULN	12	12	13	11
Tumor site
Distal extremity	1	5	5	5
Proximal extremity	16	15	17	18
Axial skeleton	3	2	2	1
Not determined	1	0	0	0
Lung disease
Unilateral	11	8	8	4
Bilateral	5	9	8	13
Missing	5	5	8	7
Sites of disease
Lung only	12	10	12	10
Bone only	2	0	3	1
Lung + bone only	1	1	3	3
Soft tissue + other	4	8	4	5
Not reported	2	3	2	5

Toxicity

Toxicity data were collected for patients enrolled in the protocol. Toxicity grading was based on the Children's Cancer Group Toxicity and Complications Criteria, a scale similar to but not identical to the National Cancer Institute's Common Toxicity Criteria (available upon request from the Children's Oncology Group Operations Center). No toxic deaths were reported. No significant differences were noted between the study arms. Select grade 3 and 4 toxicity data are presented in Table 7.

Table 7. Select Grade 3 and Grade 4 Toxicity Among Treatment Arms

Toxicity Code	Regimen
	A No MTP (n=21), Grade				A MTP (n=22), Grade				B No MTP (n=24), Grade				B MTP (n=24), Grade				All (N=91), No.
	3		4		3		4		3		4		3		4
	No.	%	No.	%	No.	%	No.	%	No.	%	No.	%	No.	%	No.	%
WBC indicates white blood cells; ANC, absolute neutrophil count; HGB, hemoglobin; AST, aspartate aminotransferase; ALT, alanine aminotransferase; AP, alkaline phosphatase; CrCl, creatinine clearance; GI, gastrointestinal.
Hematologic, WBC	3	14.3	3	14.3	2	9.1	1	4.5			6	25.0	2	8.3	6	25.0	23
Hematologic, ANC	1	4.8	9	42.9	3	13.6	5	22.7			6	25.0	3	12.5	8	33.3	35
Hematologic, platelet	2	9.5	4	19.0	1	4.5	3	13.6	1	4.2	2	8.3	2	8.3	6	25.0	21
Hematologic, HGB	2	9.5	1	4.8			1	4.5	1	4.2			1	4.2	3	12.5	9
Hepatic, AST	7	33.3	2	9.5	6	27.3	3	13.6	7	29.2	3	12.5	10	41.7	3	12.5	41
Hepatic, ALT	6	28.6	5	23.8	6	27.3	4	18.2	12	50.0	4	16.7	11	45.8	6	25.0	54
Hepatic, AP	1	4.8													1	4.2	2
Hepatic, total bili	2	9.5	1	4.8									2	8.3	2	8.3	7
Renal, creatinine													1	4.2			1
Renal, CrCl							1	4.5	1	4.2					2	8.3	4
GI, stomatitis	2	9.5	5	23.8	4	18.2	3	13.6	5	20.8	2	8.3	2	8.3	7	29.2	30
GI, nausea & vomiting	1	4.8	3	14.3	4	18.2	1	4.5	2	8.3			4	16.7	1	4.2	16
Cardiac, rhythm									1	4.2					1	4.2	2
Nervous, peripheral sensory	1	4.8															1
Nervous, centrocerebellar	1	4.8											1	4.2	1	4.2	3
Skin									1	4.2			1	4.2			2
Hearing, objective					1	4.5			2	8.3			1	4.2			4
Infection	4	19.0			3	13.6	1	4.5	5	20.8			4	16.7	1	4.2	18
Fever					1	4.5							2	8.3	1	4.2	4
Performance status			1	4.8													1

DISCUSSION

The poor prognosis of patients who present with metastatic OS highlights the need for new active agents to be identified. We have learned that dose intensification of conventional cytotoxic agents, although leading to greater necrosis after preoperative chemotherapy, does not alter the 20% long-term survival of patients with metastatic OS.25, 26 Addition of conventional cytotoxic agents that appear to have activity in the preclinical setting, such as topotecan, has also not led to improvement in patient survival.1

As an immune modulator, liposomal MTP-PE has been studied extensively and presents a novel avenue of therapy for these patients. The initial analysis of the nonmetastatic cohort of Intergroup Study 0133 detected interaction between the 2 factors under study.23 This led to a preliminary conclusion that the addition of liposomal MTP-PE to traditional chemotherapy did not confer a survival advantage to these patients.23 However, with longer follow-up of the same cohort, the test of the hypothesis of interaction no longer met a conventional level of significance, allowing us to perform the factorial analyses originally designed for this study. We identified a trend toward improved EFS and a statistically significant improvement in overall survival for those who received liposomal MTP-PE, independent of chemotherapy regimen.22 Specifically, in the nonmetastatic cohort of patients (n = 662), the relative risk of death for patients randomized to receive liposomal MTP-PE was 0.71 (95% CI, 0.52-0.96; P = .03), and the hazard ratio for EFS for patients who received liposomal MTP-PE was 0.80 (95% CI, 0.62-1.0; P = .08). Interestingly, the same pattern of survival enhancement seen in the nonmetastatic, resectable group of patients was also seen in the metastatic cohort reported here: a trend toward improved EFS and overall survival with addition of liposomal MTP-PE to chemotherapy. In addition, there was no statistically significant difference between the 2 chemotherapy regimens with respect to EFS or overall survival in the metastatic cohort presented here. The study was not powered to detect differences between the 4 study arms. We cannot make firm conclusions about the apparent differences of survival between the study arms. A larger, adequately powered study would help to resolve potential differences between chemotherapy regimens.

Previous studies described clinical characteristics in the population of metastatic patients that correlate with outcome, including age, sites of metastases, number of pulmonary nodules, and levels of biochemical markers (AP, LDH).3, 5, 7, 9 Many of these studies were performed at single institutions or described a population of patients who were treated with different protocols over a span of decades. Although this study was not powered to detect outcome differences in a subpopulation of patients with metastatic disease, it does provide valuable clues as to potentially prognostic clinical characteristics. Several clinical factors in this particular cohort of patients were related to worse outcome, including sex (male), race (non-Caucasians), extent of disease involvement (bone metastases), and nonspecific biochemical markers of tumor burden (high AP and LDH). When incorporated into a relative risk regression model, both sex and baseline alkaline phosphatase levels were related to overall survival and EFS. In this study, female patients had better EFS and overall survival. Although this recapitulates the findings of published studies by some, but not all, investigators (most notably, the Scandinavian Sarcoma Group recently published data supporting the better prognosis of female OS patients),5, 7, 27 the reason for this consistent observation remains unclear. In addition, these results reinforce the notion that increased tumor burden (as suggested by higher AP) leads to worse prognosis. Of interest is the lack of impact the number and location of lung nodules had on both EFS and overall survival. This is likely a reflection of the small sample size of the study population. A larger population would have provided an opportunity to detect the impact of lung tumor burden in these patients. Also, age at diagnosis did not correlate with outcome in this group of patients. In addition to these 2 characteristics, race was significantly related to risk of adverse analytic event, with non-Caucasian patients having worse EFS and overall survival.

Although the small number of patients in this cohort preclude precise estimation of treatment effects, it is still important to note that both EFS and overall survival appear to be improved for the patients who received liposomal MTP-PE. In addition, the reductions in risk of adverse analytic event and death are very similar to the pattern in the much larger cohort of patients who presented without clinically detectable metastatic disease. These data suggest that liposomal MTP-PE might provide benefit when added to chemotherapy for the treatment of patients with osteosarcoma who present with metastatic disease. With the recent approval of liposomal MTP-PE by the European Medicines Agency, the agent should become widely available for inclusion in additional, larger studies defining its efficacy in osteosarcoma.

Acknowledgements

We thank Dr. Judith Sato and Dr. Helen Nadel for valuable contributions to this study.

Conflict of Interest Disclosures

Supported by the Children's Oncology Group (COG) Chair's Grant U10 CA98543 and the Statistics & Data Center Grant U10 CA98413 from the National Institutes of Health. A complete listing of grant support for research conducted by CCG and POG before initiation of the COG grant in 2003 is available online at: http://www.childrensoncologygroup.org/admin/grantinfo.htm

Research funding: Paul A. Meyers, IDM Pharma. Expert testimony: Eugenie S. Kleinerman, IDM Pharma (uncompensated); Paul A. Meyers, IDM Pharma (uncompensated).