Assessment of Doxorubicin and Pembrolizumab in Patients With Advanced Anthracycline-Naive Sarcoma: A Phase 1/2 Nonrandomized Clinical Trial.

IMPORTANCE: Anthracycline-based therapy is standard first-line treatment for most patients with advanced and metastatic sarcomas. Although multiple trials have attempted to show improved outcomes in patients with soft-tissue sarcoma over doxorubicin monotherapy, each has fallen short of demonstrating improved outcomes.
OBJECTIVE: To evaluate the safety and efficacy of doxorubicin in combination with pembrolizumab in patients with advanced, anthracycline-naive sarcomas. DESIGN, SETTING, AND PARTICIPANTS: This nonrandomized clinical trial used a 2-stage phase 2 design and was performed at a single, academic sarcoma specialty center. Patients were adults with good performance status and end-organ function. Patients with all sarcoma subtypes were allowed to enroll with the exception of osteosarcoma, Ewing sarcoma, and alveolar and embryonal rhabdomyosarcoma.
INTERVENTIONS: Two dose levels of doxorubicin (45 and 75 mg/m2) were tested for safety in combination with pembrolizumab. MAIN OUTCOMES AND MEASURES: Objective response rate (ORR) was the primary end point. Overall survival (OS) and progression-free survival (PFS) were secondary end points. Correlative studies included immunohistochemistry, gene expression, and serum cytokines.
RESULTS: A total of 37 patients (22 men; 15 women) were treated in the combined phase 1/2 trial. The median (range) patient age was 58.4 (25-80) years. The most common histologic subtype was leiomyosarcoma (11 patients). Doxorubicin plus pembrolizumab was well tolerated without significant unexpected toxic effects. The ORR was 13% for phase 2 patients and 19% overall. Median PFS was 8.1 (95% CI, 7.6-10.8) months. Median OS was 27.6 (95% CI, 18.7-not reached) months at the time of this analysis. Two of 3 patients with undifferentiated pleomorphic sarcoma and 2 of 4 patients with dedifferentiated liposarcoma had durable partial responses. Tumor-infiltrating lymphocytes were present in 21% of evaluable tumors and associated with inferior PFS (log-rank P = .03). No dose-limiting toxic effects were observed. CONCLUSIONS AND RELEVANCE: In this nonrandomized clinical trial, doxorubicin plus pembrolizumab was well tolerated. Although the primary end point for ORR was not reached, the PFS and OS observed compared favorably with prior published studies. Further studies are warranted, especially those focusing on undifferentiated pleomorphic sarcoma and dedifferentiated liposarcoma. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02888665.

Introduction

Patients with advanced soft-tissue sarcoma (STS) generally receive first-line therapy with doxorubicin alone or with ifosfamide.[1] Median overall survival (OS) for doxorubicin alone ranges from 12.8 to 20.4 months, with median progression-free survival (PFS) of 4.1 to 6.8 months and objective response rates (ORRs) of 12% to 20%.[1,2,3,4,5] Multiple trials combining doxorubicin with investigational agents have failed to show improved outcomes.[1,2,3,4,5,6]

Programmed cell death ligand 1 (PD-L1) is variably expressed by STS, especially undifferentiated pleomorphic sarcomas (UPSs).[7,8] Responses to pembrolizumab monotherapy have been seen in STS, particularly UPS,[9] and some, though not all,[10] combination therapies may be associated with improved programmed cell death 1 (PD-1) blockade.[11,12,13] Chemotherapy may synergize with immunotherapy by depleting suppressive immune cells, releasing damage-associated molecular patterns and increasing tumor antigen presentation through tumor toxicity.[14] We conducted a phase 1/2 trial to assess the tolerability and outcomes of treatment with pembrolizumab in combination with doxorubicin.

Methods

Patients, Treatment Schedules, and Supportive Care

All research herein was reviewed and approved by the Fred Hutchinson Cancer Research Center Institutional Review Board. All patients provided written informed consent in accordance with the Helsinki Declaration. The trial protocol can be found in Supplement 1. Race and ethnicity data were self-reported and collected for institutional reporting requirements.

Pathology was confirmed at the University of Washington. Adult patients with sarcoma with Eastern Cooperative Oncology Group performance status of 0 or 1 and adequate organ function were eligible. Patients with osteosarcoma, Ewing sarcoma, and alveolar and embryonal rhabdomyosarcoma were ineligible because well-established alternative regimens exist (eFigure 1 in Supplement 2).

Patients’ initial cycle was pembrolizumab (200 mg administered intravenously) alone. Cycles were 21 days. Starting with cycle 2, doxorubicin was given prior to pembrolizumab, same day, every 3 weeks, for up to 6 cycles. After cycle 7, pembrolizumab treatment continued for up to 2 years (see eFigure 2A in Supplement 2). Growth factors were not permitted until cycle 3 during phase 1. Imaging, performed every 12 weeks, was assessed by Response Evaluation Criteria in Solid Tumors (RECIST), version 1.1 with the option of confirming progressive disease (PD) in clinically well patients.

Trial Design, Statistical Basis, and End Points

Two doses of doxorubicin (45 and 75 mg/m2) were evaluated with pembrolizumab using a 3 + 3 design. A dose-limiting toxic effect was defined as any possibly related, unexpected, grade 3 or greater serious adverse events, or any possibly related adverse event requiring discontinuation of either drug during the first 6 weeks of combination treatment (eFigure 2B in Supplement 2).

The primary end point of the phase 2 portion of the study was the ORR as assessed by RECIST 1.1.[15] The 2-stage study was designed to rule out ORR of 15% or less with 85% power if the true ORR was 35%, using a 1-sided 5% level test. If 2 or more responses were observed in the 20-patient first stage, an additional 15 patients would accrue. If 10 or more responses of 35 patients (29%) were observed, this would rule out a 15% ORR. Although the primary end points were evaluated for the phase 1 and 2 cohorts separately, data were combined in other analyses. Data were analyzed as of September 2019.

The OS was calculated as the duration from the start of treatment to death due to any cause, and PFS as the duration from start to progression or death. Outcomes were censored at the date of last contact for living patients (OS) or living and progression free (PFS). For these secondary end points, 2-sided P values less than .05 were considered significant. All analyses were performed using SAS, version 9.4 (SAS Institute); Excel, version 16.33 (Microsoft Corp); and Prism, version 8.4 (GraphPad Software) (see eFigure 3 in Supplement 2 for detailed methods regarding correlative analyses).

Results

Patient Demographic Characteristics

In the combined phase 1/2 trial, 37 patients (22 men; 15 women) were treated, including 6 phase 1 patients—3 patients at each dose. Both phase 2 stages enrolled, but accrual was closed at 31 of 35 planned patients because of an insufficient number of second-stage partial responses (PRs), indicating that the study would not achieve the primary end point. The most common histology was leiomyosarcoma, present in 11 patients (30%), 3 being uterine leiomyosarcomas. Demographic characteristics, including histology, are summarized in Table 1.

Table 1.

Patient Demographic Characteristics

Characteristic	No. (%) (N = 37)
Age at day 1, median (range), y	58.4 (25.8-80.4)
Sex
Female	15 (41)
Male	22 (59)
Race
American Indian or Alaska Native	2 (6)
Asian	2 (6)
Black or African American	1 (3)
White/other	32 (85)
No. of prior lines of systemic treatment
0	28 (76)
1	7 (19)
2	2 (5)
Doxorubicin dose, mg/m2
45	3 (8)
75	34 (92)
Best response
Not evaluable	1 (3)
Partial response	7 (19)
Progressive disease	7 (19)
Stable disease	22 (59)
Disease
Alveolar soft-part sarcoma	1 (3)
Angiosarcoma	1 (3)
Clear cell chondrosarcoma	1 (3)
Conventional chondrosarcoma	3 (8)
Dedifferentiated liposarcoma	4 (11)
Endometrial stromal sarcoma	2 (5)
Epithelioid hemangioendothelioma	2 (5)
Epithelioid sarcoma	1 (3)
Extraskeletal myxoid chondrosarcoma	1 (3)
Hemangiopericytoma	1 (3)
Leiomyosarcoma	11 (30)
Myxofibrosarcoma	1 (3)
Pleomorphic liposarcoma	1 (3)
Pleomorphic rhabdomyosarcoma	1 (3)
Solitary fibrous tumor	2 (5)
Spindle cell sarcoma	1 (3)
Undifferentiated pleomorphic sarcoma	3 (8)

Safety

No dose-limiting toxic effects were observed. The phase 2 dose was 75 mg/m2. In both phase 1 and 2 cohorts, the most common toxic effects were nausea (n = 32) and fatigue (n = 21) (see Table 2; eTable 1 in Supplement 2). No grade 5 toxic effects were seen; the only attributable grade 4 toxic effects were neutropenia (n = 6), leukopenia (n = 1), and febrile neutropenia (1), all of which resolved. Two patients had grade 3 reductions in ejection fraction attributable to doxorubicin. Notable pembrolizumab-related toxic effects included grade 3 adrenal insufficiency (n = 1) and hypothyroidism (n = 7).

Table 2.

Adverse Events

Adverse event	No.				Total No. of events
	Grade
	1	2	3	4
Adverse events with at least 2 grade 3 or 4 events
Ejection fraction decreased	0	0	2	0	2
Lymphocyte count decreased	0	0	2	0	2
Febrile neutropenia	0	1	1	1	3
White blood cell count decreased	0	0	2	1	3
Anemia	2	1	2	0	5
Neutrophil count decreased	0	1	2	6	9
Mucositis, oral	3	7	3	0	13
Anorexia	9	7	2	0	18
Other adverse events with at least 4 events
Rash, maculopapular	2	2	0	0	4
Upper respiratory tract infection	2	2	0	0	4
Dyspnea	3	1	0	0	4
Headache	3	1	0	0	4
Constipation	4	1	0	0	5
Rash	4	2	0	0	6
Hypomagnesemia	6	0	0	0	6
Weight loss	4	1	1	0	6
Fever	6	1	0	0	7
Hypothyroidism	2	4	1	0	7
Dry eye	7	1	0	0	8
Diarrhea	5	2	1	0	8
Pruritus	8	1	0	0	9
Dysgeusia	7	3	0	0	10
Vomiting	5	6	0	0	11
Dry mouth	10	1	0	0	11
Alopecia	4	10	0	0	14
Fatigue	11	10	0	0	21
Nausea	15	16	1	0	32

Tumor Response

In the combined phase 1/2 trial, confirmed PRs were seen in 7 of 37 patients (19%), 4 in the phase 2 cohort (13%) and 3 in the 75 mg/m2 phase 1 cohort (Figure, A). Two patients had unconfirmed PRs, and 11 patients had stable disease with tumor regression as their best response (Figure, B). One patient came off study for increasing symptoms prior to follow-up imaging. Two of 3 patients with UPS, and 2 of 4 patients with dedifferentiated liposarcoma had durable PRs (eFigure 4 in Supplement 2). Three patients with chondrosarcoma had tumor regression, including 1 conventional chondrosarcoma with a 26% decrease in size.

Figure.

Patient Responses

A, Waterfall plot demonstrating the percentage change in tumor size from baseline constituting the best response for each patient. B, Spider plots showing responses for all patients.

Survival Outcomes

Median PFS was 8.1 (95% CI, 7.6-10.8) months, with 29 of 37 patients (78.4%) having had an event and 4 patients with continuing stable disease or PR at the time of this analysis. The PFS rates at 12 and 24 weeks were 81% (95% CI, 64%-90%) and 73% (95% CI, 56%-84%), respectively. At 12 months, the PFS was 27% (95% CI, 14%-42%). Median OS was 27.6 (95% CI, 18.7-not reached) months at the time of this analysis.

Correlative Studies

Immunohistochemistry was evaluable for 29 patients; 66% had PD-L1 expression scores of 0, reflecting a low level of PD-L1 expression (eTable 2 in Supplement 2). Expression of PD-L1 was not associated with PFS or OS. Tumor-infiltrating lymphocytes were present in 21% of evaluable tumors and associated with inferior PFS (log-rank P = .03) (eFigure 5 in Supplement 2). This was confirmed in a multivariate Cox regression analysis that adjusted for age, sex, and number of prior therapies (P = .04; eTable 3 in Supplement 2). Nanostring data were available for 24 patients (eTable 4 in Supplement 2). No gene was significantly associated with PFS after correction for multiple comparisons.

Serum cytokine levels were assessed before treatment and during cycles 1 and 2. Granulocyte macrophage–colony-stimulating factor levels increased each cycle, and IL-15 levels dropped following doxorubicin treatment. Circulating IL-2R, IP10, and CD30 levels rose sharply after cycle 1, while levels of IL-8 dropped.

Discussion

This nonrandomized phase 1/2 trial demonstrated that doxorubicin plus pembrolizumab can be given safely and may be associated with clinical benefit for patients with advanced sarcoma. While the combined ORR observed here is similar to prior published series, the PFS and OS seen here are encouraging.[2,3,4,6] In correlative studies, we found that tumor-infiltrating lymphocytes were associated with inferior PFS. This may reflect more aggressive tumor biology rather than as association with the PD-1 inhibitor.

Limitations

Similar to other sarcoma trials, histologic subtypes likely influenced these results. Our trial, like others, demonstrated a higher response rate and clinical benefit of pembrolizumab in UPS compared with other sarcoma subtypes,[11] despite their generally worse prognosis.[5] Durable PRs were also seen in 2 of 4 patients with dedifferentiated liposarcoma. Chondrosarcomas are generally resistant to doxorubicin; inclusion of these patients in the study likely lowered the ORR. Still, 2 patients with chondrosarcoma had durable disease regression, suggesting that doxorubicin/pembrolizumab may benefit these patients. For patients with these selected subtypes, follow-up studies are warranted.