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Journal of Clinical Oncology, Vol 23, No 4 (February 1), 2005: pp. 866-873 © 2005 American Society of Clinical Oncology. DOI: 10.1200/JCO.2005.07.088 Molecular and Clinical Analysis of Locally Advanced Dermatofibrosarcoma Protuberans Treated With Imatinib: Imatinib Target Exploration Consortium Study B2225From the Peter MacCallum Cancer Centre, East Melbourne, Australia; Dana-Farber Cancer Institute and Harvard Medical School, Sarcoma Center; Department of Pathology, Brigham & Women's Hospital, Boston, MA; Universiteit Ziekenhuis Gasthuisberg dienst oncology; Cytogenetics and Molecular Genetics of Human Malignancies, Department of Human Genetics, Catholic University of Leuven, Leuven, Belgium; Departments of Medicine and Pathology, Oregon Health Sciences University Cancer Institute and Portland Veterans Affairs Medical Center, Portland, OR; and Clinical Research Oncology, Novartis Pharma AG, Basel, Switzerland Address reprint requests to information: Grant McArthur, MB, BS, PhD, Peter MacCallum Cancer Centre, St Andrews Place, East Melbourne, Australia 3002; e-mail: grant.mcarthur{at}petermac.org
PURPOSE: The cutaneous malignant tumor dermatofibrosarcoma protuberans (DFSP) is typically associated with a translocation between chromosomes 17 and 22 that places the platelet-derived growth factor–B (PDGFB) under the control of the collagen 1A1 promoter. The purpose of this study was to evaluate molecular, cytogenetic, and kinase activation profiles in a series of DFSPs and to determine whether these biologic parameters are correlated with the clinical responses of DFSP to imatinib. PATIENTS AND METHODS: We analyzed the objective radiologic and clinical response to imatinib at 400 mg twice daily in eight patients with locally advanced DFSP and two patients with metastatic disease. RESULTS: Each of eight patients with locally advanced DFSP had evidence of t(17;22) and showed a clinical response to imatinib. Four of these patients had complete clinical responses. The two patients with metastatic disease had fibrosarcomatous histology and karyotypes that were substantially more complex than those typically associated with localized DFSP. One patient with metastatic DFSP and an associated t(17;22) had a partial response to imatinib but experienced disease progression after 7 months of therapy. In contrast, the other patient with metastatic disease had a tumor lacking t(17;22), and there was no clinical response to imatinib. Unexpectedly, there was minimal platelet-derived growth factor receptor–beta phosphorylation in the untreated DFSP, despite the documented presence of a PDGFB autocrine mechanism. CONCLUSION: Imatinib has clinical activity against both localized and metastatic DFSP with t(17;22). However, fibrosarcomatous variants of DFSP lacking t(17;22) may not respond to imatinib.
Dermatofibrosarcoma protuberans (DFSP) is a rare malignant tumor of the subcutaneous tissues characterized by slow but infiltrative growth. Standard management of localized disease is local surgical resection with wide margins.1 However, local recurrence rates are high (13% to 60%),1-3 especially in DFSP with fibrosarcomatous histopathology.4 In retrospective series, the use of Mohs micrographic surgery, where histologic examination is used intraoperatively to ensure clear margins, has been associated with lower rates of recurrence.5 However, Mohs surgery is not practiced in many centers because of practical difficulties implementing intraoperative pathology. In addition to local control issues, 1% to 4% of DFSP may develop distant metastases.4 Effective systemic therapy for locally advanced, inoperable, or metastatic disease would clearly be useful in the management of this tumor. More than 90% of DFSP feature a translocation involving distinct regions of chromosomes 17 and 22.6 Most commonly, the t(17;22) breakpoint region is tandemly repeated within a supernumerary ring chromosome.7 The translocation breakpoint generally involves the second exon of the platelet-derived growth factor-B (PDGFB) gene on chromosome 22, which is fused with the strongly expressed collagen 1 alpha 1 (COL1A1) gene on chromosome 17. This distinctive translocation mechanism results in transcriptional upregulation of the PDGFB gene, in the form of a COL1A1-PDGFB fusion oncogene. The associated COL1A1-PDGF-B fusion protein8-10 is post-translationally processed to yield mature and fully functional PDGFB.11 Therefore, the t(17;22) results in PDGFB-mediated activation of platelet-derived growth factor receptor–beta (PDGFRB) by autocrine and paracrine production of a functional ligand for PDGFRB. Because PDGFRB is the major PDGFR isoform expressed on DFSP,12,13 the t(17;22) most likely activates PDGFRB rather than PDGFR-alpha (PDGFRA).11 The identification of deregulated expression of PDGFB as a result of the t(17;22) led to the hypothesis that inhibitors of PDGFRB, such as imatinib, might have activity in DFSP. After supportive preclinical studies,11,12 there have been several reports of clinical activity of imatinib in DFSP.14-16 Here we report a series of 10 patients with DFSP who received imatinib for locally advanced or metastatic DFSP. Strikingly, all eight patients with locally advanced disease responded to imatinib. Unexpectedly, these dramatic clinical responses were seen even in DFSPs expressing relatively low amounts of activated PDGFRB.
Study Design Patients with DFSP were enrolled onto an open-label, phase II study investigating the effects of imatinib in patients with life-threatening diseases known to be associated with one or more imatinib-sensitive tyrosine kinases (Imatinib Target Exploration Consortium Study B2225). The primary aim of the study was to assess activity of imatinib in diseases associated with expression or activation of imatinib-sensitive tyrosine kinases. Patients were treated with 800 mg of imatinib daily (400 mg twice daily) with dose reductions for grade 3 toxicity or for recurrent grade 2 toxicity after an initial 1-week treatment break. The initial dose reduction was to 600 mg, with a second dose reduction to 400 mg allowed for further grade 3 toxicity or recurrent grade 2 toxicity after a 1-week treatment break. Full details of the clinical trial will be published separately. Study investigations were performed after approval by a local human investigations committee and for United States centers in accord with an assurance filed with and approved by the United States Department of Health and Human Services. Informed consent was obtained from each patient.
Eligibility
Efficacy Assessments
All patients were evaluated with either tumor imaging with computed tomography or magnetic resonance imaging scans (patients 9 and 10), clinical photography in the case of locally advanced tumors with superficially visible disease (patients 1, 2, 4, and 6), or clinical examination by palpation with measurements confirmed by ultrasound (patients 3, 5, 7, and 8). Measurable disease in the skin was defined as lesions with at least one diameter
Western Blot Analyses
Karyotyping
Fluorescence In Situ Hybridization
Ten assessable patients were treated with imatinib in the Imatinib Target Exploration Consortium Study B2225. The clinicopathologic and cytogenetic characteristics are listed in Table 1. In each of eight patients with locally advanced disease, the DFSP had noncomplex karyotypes, featuring a typical ring chromosome, and with molecular cytogenetic evidence of t(17;22) by PDGFB FISH and/or painting for chromosomes 17 and 22. By contrast, both patients with metastatic disease had tumors with more complex karyotypes, featuring various numeric and structural chromosomal aberrations. The metastatic DFSP in patient no. 9 was hyperdiploid, and the cytogenetic aberrations included several copies of a ring chromosome (Fig 1C). PDGFB FISH confirmed t(17;22) in the ring chromosomes (Fig 1D). The metastatic DFSP-like tumor in patient 10 lacked a ring chromosome (Fig 1E) and lacked PDGFB rearrangement by FISH in paraffin tumor sections (Fig 1F) and in cytogenetic preparations.
All patients were commenced on imatinib at 800 mg daily (400 mg bid). In this patient population, 800 mg daily was well tolerated, with only one patient requiring dose reduction to 600 mg daily. Four patients received tumor biopsies before and approximately 28 days after commencement of imatinib. Responses were evaluated using traditional Southwest Oncology Group criteria (pre-Response Evaluation Criteria in Solid Tumors). Notably, all eight patients with locally advanced DFSP had partial (four patients) or complete (four patients) responses to imatinib (Table 2). Two patients (patients 7 and 8) who proceeded to surgical resection after initial clinical response had pathologically confirmed complete responses. In four further cases (patients 1, 2, 3, and 5), the patients were rendered disease-free by surgical resection after partial response to imatinib. Of the two patients with metastatic disease and complex cytogenetics, patient 10, whose DFSP lacked the t(17;22), had no clinical response to imatinib by either conventional imaging or fluorodeoxyglucose positron emission tomography scan at day 19 of imatinib therapy. However, follow-up on this patient was quite short because of the patient's death on day 32. By contrast, patient 9, whose metastatic DFSP featured the t(17;22) and other cytogenetic aberrations, had a robust partial response (Table 2 and Fig 2). The clinical responses to imatinib were sustained in all four patients treated with imatinib for greater than 6 months. However, two of these patients subsequently experienced disease progression. Patient 1, with locally advanced DFSP on the cheek, responded for just less than 2 years before the tumor progressed, at which point the patient was rendered disease-free after resection using Mohs surgical technique. Patient 9 with metastatic disease experienced disease progression after 7 months. Patients rendered free of disease by surgical resection had no evidence of progression at 4, 18, 12, and 1 months after the resection.
Several patients had matched pretreatment and on-therapy tumor biopsy specimens. Biopsy specimens obtained during imatinib therapy had reduced cellularity and the development of hyaline changes (Fig 3). PDGFB FISH analysis showed persistence of the t(17;22) in the spindle-cell component of the resected stable disease. However, the t(17;22) was found in only 20% of the spindle cells, suggesting that most of the residual spindle cells were either reactive or were DFSP cells lacking t(17;22).
Tumor tissue was examined for expression and activation of PDGFRA and PDGFRB by immunoblot using antisera to the active phosphorylated forms of the PDGF-receptor. Interestingly, PDGFR activation was low in five DFSPs evaluated before imatinib therapy (two examples are shown in Fig 4). Indeed, PDGFR activation was at least 10-fold lower than in gastrointestinal stromal tumors (GISTs) expressing PDGFR with an intrinsic gain-of-function mutation (Fig 4). The weak baseline activation in the DFSP precluded accurate quantification of the reduction in activated PDGFRB inhibition in response to imatinib.
This report details the clinical response to imatinib of 10 patients with locally advanced or metastatic DFSP. All tumors were characterized by cytogenetic and in some cases immunoblotting methods. In our series, there was a good clinical response of all eight localized DFSP with an associated t(17;22). Although most localized DFSP can be managed with local surgery,1 our findings demonstrate that imatinib may assist in disease control in patients with locally advanced or metastatic disease. In four patients, imatinib therapy accomplished substantial regression of locally advanced tumors before surgical resection. All four of these patients remain disease-free after surgery, including two that have been followed up for more than a year after surgical resection. Although Mohs surgery may improve local control,5 and surgery remains the treatment modality of choice for localized DFSP, our results suggest that medical therapy with imatinib may be indicated in following situations: (1) locally advanced disease that is inoperable, (2) locally advanced disease where reduction in the size of the tumor would aid in surgical resection, and (3) metastatic disease not amenable to surgical resection. It is not clear whether imatinib therapy before surgery reduces the need for wide excision margins or Mohs surgery. Therefore, we would continue to recommend wide excision margins or Mohs surgery if definitive surgery follows imatinib therapy. Imatinib therapy may be the treatment of choice in patients with locally advanced disease not amenable to surgery, as response rates to radiotherapy or cytotoxic chemotherapy seem to be inferior.1,19,20 The high response rate to imatinib therapy in DFSP supports the hypothesis that DFSP cells are dependent on aberrant activation of PDGFRB for cellular proliferation and survival. Our clinical results are consistent with studies of DFSP cell culture and animal models, in which t(17;22)-mediated PDGFRB autocrine activation is effectively inhibited by imatinib. Unexpectedly, the level of PDGFRB activation as assessed by receptor autophosphorylation was substantially lower than PDGFR activation in a GIST with intrinsic PDGFRA oncogenic mutation. Therefore, although mutation of a serine/threonine kinase such as BRAF in melanoma21 or mutation or amplification of a receptor tyrosine kinase (RTK) such as PDGFRA in GIST tumors,22 epidermal growth factor receptor in epithelial tumors,23 or ERB-B2 in breast cancer24 leads to strong activation of the involved oncogenic RTKs, it seems that autocrine/paracrine oncogenic mechanisms—as seen in DFSP—can be associated with substantially lower levels of RTK activation. These novel observations indicate that neither high levels of RTK activation nor RTK overexpression are required for clinical response to therapeutic inhibition of receptor signaling. Rather, inhibition of low-level RTK activation can be clinically effective, providing that the tumor cells are dependent on that signaling mechanism. Our findings demonstrating low activation of PDGFRB in DFSP tumors contrast with those of Sjöblom et al,12 who readily detected phosphorylation of PDGFRB in cultured cells derived from DFSP tumors. We propose two possible explanations for this apparent discrepancy: first, the culture conditions used by Sjöblom et al included the use of fetal calf serum, a source of exogenous PDGFs. It is possible that these culture conditions induced greater phosphorylation of PDGFRB than we observed in tumor biopsy specimens. Second; Sjöblom et al used immunoprecipitation rather than direct immunoblotting to detect phosphorylation of PDGFRB. It is likely that immunoprecipitation is more sensitive than immunoblotting at detecting phosphorylated PDGFRB. Nonetheless, there is a clear and substantial difference between receptor phosphorylation in GISTs or PDGFA/B-stimulated NIH3T3 cells compared with the DFSP tumors (Fig 4). Moreover, in our experience, only weak activation of PDGFRB is detectable using immunoprecipitation of protein lysates purified from fresh DFSP tumors (J.A.F., unpublished data). We treated two patients with metastatic disease; both patients had fibrosarcomatous histology associated with complex karyotypes. In one case (patient 10), both the locally recurrent and metastatic lesions lacked t(17;22), suggesting that this DFSP-like tumor was not necessarily dependent on signaling through PDGFRs. Consistent with this prediction, the second metastatic DFSP (patient 9) had an associated t(17;22) and had a partial clinical response to imatinib, although the DFSP progressed after 7 months of therapy. Four other patients with metastatic DFSP have been reported in the literature with clinical responses observed in all of these cases,14-16 although one patient had only a transient response of some but not all lesions.14 Notably, this patient had fibrosarcomatous histology with an associated complex karyotype but no evidence of t(17;22).14 Only one of the three other patients whose metastatic DFSP had a significant clinical response to imatinib had cytogenetic evaluation of their tumor; this case had evidence of PDGFB rearrangement by FISH. However, neither a ring chromosome or t(17;22) were present, and the PDGFB rearrangement involved an unidentified translocation partner. The limited clinical experience in metastatic DFSP suggests that imatinib therapy has a role in the management of advanced disease. Given the ineffectiveness of cytotoxic chemotherapy for this disease, we conclude that a trial of imatinib therapy is clinically indicated in patients with metastatic disease. Further studies may be helpful in determining the usefulness of cytogenetics and/or PDGFB FISH in predicting the likelihood of clinical response of metastatic DFSP to imatinib therapy.
The following authors or their immediate family members have indicated a financial interest. No conflict exists for drugs or devices used in a study if they are not being evaluated as part of the investigation. Employment: Zariana Nikolova, Novartis; Sasa Dimitrijevic, Novartis. Consultant/advisory role: Grant A. McArthur, Novartis; George D. Demetri, Novartis; Allan van Oosterom, Aventis, Eli Lilly, Novartis, Pfizer, Pharmanaut, Roche; Michael C. Heinrich, Novartis; Christopher L. Corless, Novartis; Jonathan A. Fletcher, Novartis. Stock ownership: Zariana Nikolova, Novartis; Sasa Dimitrijevic, Novartis. Honoraria: George D. Demetri, Novartis; Michael C. Heinrich, Novartis. Research funding: George D. Demetri, Novartis; Allan van Oosterom, Amgen, Aventis, Novartis, Pfizer, Pharmanaut, Roche, Schering-Plough; Michael C. Heinrich, Novartis; Jonathan A. Fletcher, Novartis. Expert testimony: George D. Demetri, Novartis.
Presented in part at the 39th Annual Meeting of the American Society of Clinical Oncology, Chicago, IL, May 31-June 3, 2003. Authors' disclosures of potential conflicts of interest are found at the end of this article.
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11. Shimizu A, O'Brien KP, Sjoblom T, et al: The dermatofibrosarcoma protuberans-associated collagen type Ialpha1/platelet-derived growth factor (PDGF) B-chain fusion gene generates a transforming protein that is processed to functional PDGF-BB. Cancer Res 59:3719-3723, 1999
12. Sjoblom T, Shimizu A, O'Brien KP, et al: Growth inhibition of dermatofibrosarcoma protuberans tumors by the platelet-derived growth factor receptor antagonist STI571 through induction of apoptosis. Cancer Res 61:5778-5783, 2001
13. Linn SC, West RB, Pollack JR, et al: Gene expression patterns and gene copy number changes in dermatofibrosarcoma protuberans. Am J Pathol 163:2383-2395, 2003 14. Maki RG, Awan RA, Dixon RH, et al: Differential sensitivity to imatinib of 2 patients with metastatic sarcoma arising from dermatofibrosarcoma protuberans. Int J Cancer 100:623-626, 2002[CrossRef][Medline]
15. Rubin BP, Schuetze SM, Eary JF, et al: Molecular targeting of platelet-derived growth factor B by imatinib mesylate in a patient with metastatic dermatofibrosarcoma protuberans. J Clin Oncol 20:3586-3591, 2002 16. Labropoulos S, Papadopoulos S, Hadjiyiassemi L, et al: Response of metastatic dermatofibrosarcoma protuberans to imatinib mesylate. Proc Am Soc Clin Oncol 830:830, 2003 (abstr 3334) 17. Fletcher JA, Kozakewich HP, Hoffer FA, et al: Diagnostic relevance of clonal cytogenetic aberrations in malignant soft-tissue tumors. N Engl J Med 324:436-442, 1991[Abstract] 18. Hsi BL, Xiao S, Fletcher JA: Chromogenic in situ hybridization and FISH in pathology. Methods Mol Biol 204:343-351, 2002[Medline] 19. Suit H, Spiro I, Mankin HJ, et al: Radiation in management of patients with dermatofibrosarcoma protuberans. J Clin Oncol 14:2365-2369, 1996[Abstract] 20. Ballo MT, Zagars GK, Pisters P, et al: The role of radiation therapy in the management of dermatofibrosarcoma protuberans. Int J Radiat Oncol Biol Phys 40:823-827, 1998[CrossRef][Medline] 21. Wan PT, Garnett MJ, Roe SM, et al: Mechanism of activation of the RAF-ERK signaling pathway by oncogenic mutations of B-RAF. Cell 116:855-867, 2004[CrossRef][Medline]
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23. Lynch TJ, Bell DW, Sordella R, et al: Activating mutations in the epidermal growth factor receptor underlying responsiveness of non-small-cell lung cancer to gefitinib. N Engl J Med 350:2129-2139, 2004 24. Segatto O, Lonardo F, Pierce JH, et al: The role of autophosphorylation in modulation of erbB-2 transforming function. New Biol 2:187-195, 1990[Medline] Submitted July 20, 2004; accepted November 2, 2004.
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Copyright © 2005 by the American Society of Clinical Oncology, Online ISSN: 1527-7755. Print ISSN: 0732-183X
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ABSTRACT
INTRODUCTION
50% decrease from baseline in the sum of products of perpendicular diameters of all measurable lesions. Complete response was defined as complete disappearance of all measurable and nonmeasurable but assessable disease. 
