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The VEGF- and PDGF-family of angiogenic markers have prognostic impact in soft tissue sarcomas arising in the extremities and trunk

  • Thomas K Kilvaer1Email author,
  • Eivind Smeland1,
  • Andrej Valkov2, 3,
  • Sveinung W Sorbye2, 3,
  • Roy M Bremnes1, 4,
  • Lill-Tove Busund2, 3 and
  • Tom Donnem1, 4
BMC Clinical PathologyBMC series ¿ open, inclusive and trusted201414:5

DOI: 10.1186/1472-6890-14-5

Received: 2 November 2013

Accepted: 16 January 2014

Published: 20 January 2014

Abstract

Background

Soft-tissue sarcomas are rare malignant tumors of mesenchymal lineage that can arise in any part of the body. Prognosis, and hence also treatment may vary according to histologic subtype and localization. Angiogenesis is the process of forming new blood vessels from pre-existing ones. The deregulation of this process is thought to be an important step in malignant transformation. This study investigates the prognostic impact of platelet derived growth factor- (PDGF), vascular endothelial growth factor- (VEGF) and fibroblast growth factor (FGF) families in soft-tissue sarcomas of the extremities & trunk (ET) and visceral & retroperitoneal (VR) locations.

Methods

Tumor samples from 181 patients (115 ET and 66 VR) with resected soft tissue sarcomas were collected and tissue microarrays were constructed. Immunohistochemistry was used to evaluate angiogenic marker expression. Recurrence-free survival (RFS), metastasis-free survival (MFS) and disease-specific survival (DSS) were used as endpoints in prognostic impact assessment.

Results

In univariate analyses, almost all investigated angiogenic markers had prognostic impact in the ET group. In contrast, only FGFR-1 showed any significant prognostic impact in the VR group. In the multivariate analyses, PDGF-D (HR = 1.863, 95% CI = 1.057-3.283, P = 0.031), VEGFR-1 (HR = 2.106, 95% CI = 1.038-4.272, P = 0.039) and VEGF-A (HR 2.095, 95% CI 1.028-4.271, P = 0.042) were independent negative prognosticators for DSS, MFS and RFS, respectively, in the ET group. FGFR-1 was an independent positive prognosticator for DSS (HR = 0.243, 95% CI = 0.095-0.618, P = 0.003) in the VR group.

Conclusions

Angiogenic molecules from the PDGF and VEGF families have prognostic impact in soft-tissue sarcomas arising in the ET, but not in VR locations. In the latter histological grade and resection margins are the most important prognostic factors.

Keywords

Angiogenesis Sarcoma Extremity Trunk FGF PDGF VEGF Visceral Retroperitoneal

Background

Soft tissue sarcomas (STS) constitute a highly heterogeneous collection of tumors comprising over 50 histological subtypes, arising from mesenchymal tissue and capable of forming tumors in all parts of the human body [1]. This group amounts to 0.5-1% of the annual tumor burden with a mortality of about 40-60%, resulting in an estimated 11 280 cases and 3 900 deaths in the US in 2012 [2]. It is good practice to distinguish between STSs arising in the extremity & trunk (ET), head & neck (HN) and visceral & retroperitoneal (VR) localizations as treatment and prognosis vary widely according to localization [3]. Further subdivision, according to histological type, malignancy grade, stage and vascular invasion among others, can be conducted [3]. Definitive treatment is radical surgery followed by radiotherapy in case of non-radical surgical margins [4]. Adjuvant chemotherapy for adult STS is still under investigation, and hence the routine use of such treatment is today limited to the palliative setting [5].

Angiogenesis is the process of forming new blood vessels from pre-existing ones. Folkman and coworkers proved this to be a pivotal step in carcinogenesis by showing that tumors would not grow beyond > 2 mm in diameter without forming vasculature [6, 7]. In 2001, Hanahan and Weinberg, suggested angiogenesis as one of the hallmarks of cancer [8] and in the 2011 updated version angiogenesis was still considered one of the most important aspects of cancer progression [9].

Vascular endothelial growth factors (VEGF) and receptors (VEGFR) are pivotal in endothelial cell proliferation and sprouting during angio- and lymphangiogenesis [10]. Platelet-derived growth factors (PDGF) and receptors (PDGFR) play an important part in the regulation of tumor stroma through the recruitment of pericytes and vascular smooth muscle cells helping to stabilize newly formed vessels and through stimulation of stromal cells to produce VEGF-A and thus drive angiogenesis [11, 12]. Fibroblast growth factors (FGF) and receptors (FGFR) drives endothelial cell proliferation and sprouting and activate several molecules involved in extracellular matrix remodelling including matrix metallo-proteinases and urokinase-like plasminogen activator [13].

Our group has previously reported on the expression of VEGF, PDGF and FGF families of growth factors in STSs of all sites [1416]. This report investigates the differential impact of these growth factors in STSs arising in ET versus VR localizations.

Methods

Patients and clinical samples

Primary tumor tissue from anonymized patients diagnosed with STS at the University Hospital of North-Norway and the Hospitals of Arkhangelsk County, Russia, from 1973 through 2006, were collected. In total 496 patients were registered from the hospital databases. Of these, 388 patients were excluded from the study because of: missing clinical data (n = 86), inadequate formalin-fixed paraffin-embedded (FFPE) tissue blocks (n = 161), no surgery performed and/or metastasis present at the time of diagnosis (n = 55) or head and neck sarcomas (n =13). Thus 115 patients with STSs of the extremities and trunk wall and 66 patients with STSs of visceral or retroperitoneal origin, with complete medical records and FFPE tissue blocks were eligible.

This report includes follow-up data as of September 2009. The median follow-up was 53.9 (range 0.5-391.7) months for extremity and trunk patients and 59.4 (range 0.10-366.7) months for visceral and retroperitoneal patients. Complete demographic and clinical data were collected retrospectively. Formalin-fixed and paraffin-embedded tumor specimens were obtained from the archives of the Departments of Pathology at the University Hospital of North-Norway and the Hospitals of Arkhangelsk County, Russia. The tumors were graded according to the French Fédération Nationale des centres de Lutte Contre le Cancer (FNCLCC) system and histologically subtyped according to the World Health Organization guidelines [1, 17]. Wide resection margins were defined as wide local resection with free microscopic margins or amputation of the affected limb or organ.

Microarray construction

All sarcomas were histologically reviewed by two trained pathologists (S. Sorbye and A. Valkov) and the most representative areas of tumor cells (neoplastic mesenchymal cells) were carefully selected and marked on the hematoxylin and eosin (H/E) slide and sampled for the tissue microarray (TMA) blocks. The TMAs were assembled using a tissue-arraying instrument (Beecher Instruments, Silver Springs, MD). The Detailed methodology has been previously reported [18]. Briefly, we used a 0.6 mm diameter stylet, and the study specimens were routinely sampled with four replicate core samples from different areas of neoplastic tissue. Normal tissue from the patients was used as staining control.

To include all core samples, 12 TMA blocks were constructed. Multiple 5-μm sections were cut with a Micron microtome (HM355S) and stained by specific antibodies for immunohistochemistry (IHC) analysis.

Immunohistochemistry

The applied antibodies were subjected to in-house validation by the manufacturer for IHC analysis on paraffin-embedded material. The detailed methodology has previously been reported [1416].

Scoring of immunohistochemistry

The ARIOL imaging system (Genetix, San Jose, CA) was used to scan the slides of antibody staining of the TMAs and the dominant staining intensity was scored as: 0 = negative; 1 = weak; 2 = intermediate; 3 = strong semi-qantitively on computer screen. The detailed methodology has previously been reported and cut-off values chosen were the same as in our previous studies [1416]. High expression in tumor cells were defined as ≥ 1 (VEGF-C), ≥ 1.5 (PDGF-A, PDGF-C, PDGF-B, VEGF-A, VEGF-D, VEGFR-1-2 and -3) and ≥ 2 (PDGF-D, PDGFR-α, PDGFR-β, FGF2 and FGFR-1).

Statistical methods

All statistical analyses were done using the statistical package SPSS (Chicago, IL), version 16. The IHC scores from each observer were compared for interobserver reliability by use of a two-way random effect model with absolute agreement definition. The intraclass correlation coefficient (reliability coefficient) was obtained from these results. The Chi-square test and Fishers Exact test were used to examine the association between molecular marker expression and various clinicopathological parameters. Univariate analyses were done using the Kaplan-Meier method, and statistical significance between survival curves was assessed by the log-rank test. Disease-specific survival (DSS) was determined from the date of diagnosis to the time of cancer related death. Metastasis-free survival (MFS) was defined from the date of diagnosis to the clinical appearance of the first metastasis. Recurrence-free survival (RFS), was defined from the date of diagnosis to the clinical appearance of the first recurrence. To assess the independent value of different pretreatment variables on survival, metastasis and local recurrence, in the presence of other variables, multivariate analyses were carried out using the Cox proportional hazards model. Only variables of significant value from the univariate analyses were entered into the Cox regression analysis. Probability for stepwise entry and removal was set at .05 and .10, respectively. The significance level used for all statistical tests was P < 0.05.

Ethical clearance

The Norwegian National Data Inspection Board and The Regional Committee for Research Ethics (Northern Norway) approved the study.

Results

Clinicopathological variables

The clinicopathological variables are summarized in Table 1. In the ET group, comprising 115 patients, median age was 59 (range 0-89) years, 50% of the patients were male, 67 patients were Norwegian and 48 Russian and 68% of the tumors were located in the extremities. Of the histological subtypes represented, 48 were undifferentiated pleomorphic sarcomas, 18 liposarcomas, 12 fibrosarcomas, 10 synovial sarcomas, 9 leiomyosarcomas, 5 angiosarcomas, 5 rhabdomyosarcomas, 5 malignant peripheral nerve sheath tumors (MPNST) and 3 sarcoma not otherwise specified (NOS).
Table 1

Prognostic clinicopathological variables as predictors for disease-specific survival, metastasis and local recurrence in patients with resected Extremitiy & Trunk and Visceral & Retroperitoneal soft-tissue sarcomas (univariate analyses, log rank test, n = 115 and 66 respectively)

 

Extremity & trunk

Visceral & retroperitoneal

  

Disease-specific survival

Metastasis-free survival

Recurrence-free survival

 

Disease-specific survival

Metastasis-free survival

Recurrence-free survival

Characteristics

Patients (n)

5-Year survival (%)

P

5-Year survival (%)

P

5-Year survival (%)

P

Patients (n)

5-Year survival (%)

P

5-Year survival (%)

P

5-Year survival (%)

P

Age

              

≤ 20 years

12

42

0.431

42

0.129

73

0.690

1

0

<0.001

100

0.112

100

0.786

21-60 years

48

61

 

63

 

69

 

36

71

 

67

 

71

 

> 60 years

55

55

 

69

 

64

 

29

43

 

44

 

62

 

Gender

              

Male

57

56

0.298

66

0.368

66

0.759

15

79

0.039

86

0.022

66

0.799

Female

58

54

 

61

 

67

 

51

51

 

48

 

68

 

Patient nationality

              

Norwegian

67

65

0.004

74

0.008

71

0.249

54

62

0.051

59

0.122

66

0.892

Russian

48

42

 

48

 

57

 

12

36

 

46

 

59

 

Histological entity

              

Pleomorphic sarcoma

48

42

0.004

61

0.001

56

0.664

6

50

0.917

67

0.264

40

0.274

Leiomyosarcoma

9

100

 

78

 

78

 

39

55

 

46

 

77

 

Liposarcoma

18

83

 

94

 

89

 

13

62

 

81

 

54

 

Fibrosarcoma

12

57

 

67

 

55

 

0

      

Angiosarcoma

5

40

 

20

 

67

 

2

50

 

50

 

50

 

Rhabdomyosarcoma

5

60

 

60

 

60

 

1

  

100

 

100

 

MPNST

5

53

 

60

 

60

 

4

67

 

100

 

100

 

Synovial sarcoma

10

13

 

30

 

64

 

1

100

 

0

 

0

 

Sarcoma NOS

3

100

 

67

 

67

 

0

      

Tumor size

              

< 5 cm

38

70

0.048

81

0.053

74

0.085

11

82

0.107

73

0.259

89

0.006

5-10 cm

45

48

 

53

 

61

 

24

62

 

62

 

70

 

> 10 cm

30

49

 

58

 

62

 

31

45

 

45

 

49

 

Missing

2

             

Malignancy grade

              

1

29

89

<0.001

89

0.001

85

0.054

23

78

0.005

77

0.051

86

0.046

2

41

56

 

65

 

60

 

29

46

 

42

 

58

 

3

45

35

 

43

 

57

 

14

46

 

47

 

49

 

Vascular invasion

              

Absent

64

70

<0.001

85

<0.001

79

<0.001

43

59

0.656

60

0.847

66

0.675

Present

50

35

 

33

 

43

 

20

51

 

54

 

78

 

Missing

1

      

3

      

Tumor depth

              

Superficial

12

91

0.010

100

0.012

91

0.041

32

      

Deep

103

51

 

59

 

63

 

34

      

Resection margins

              

Wide

61

66

0.004

72

0.045

82

<0.001

50

65

0.021

59

0.654

90

<0.001

Non-wide

54

44

 

54

 

46

 

16

50

 

53

 

44

 

Abbreviations: MPNST Malingnant peripheral nerve sheat tumor, NOS Not otherwise specified.

In the VR group, median age was 58 (range 13-88) years, 23% of the patients were male and 54 patients were Norwegian and 12 Russian. Of the histological subtypes represented, 39 were leiomyosarcomas, 13 liposarcomas, 6 pleomorphic sarcomas, 4 neurofibrosarcomas/MPNSTs, 2 angiosarcomas, 1 rhabdomyosarcoma and 1 synovial sarcoma.

Interobserver variability

Interobserver scoring agreement was tested for PDGF-B, PDGFR-α, VEGF-C, VEGFR-3, FGF2 and FGFR1 and found to be good (0.77-0.90, P < 0.001) [1416].

Univariate analyses

The impact of the clinicopathological variables on DSS, MFS and RFS in the ET group are summarized in Table 1. Patient nationality (P = 0.004), histological entity (p = 0.004), tumor size (p = 0.048), malignancy grade (P < 0.001), vascular invasion (P <0.001), tumor depth (P = 0.010) and resection margins (P = 0.004) were all prognostic indicators of DSS. Patient nationality (P = 0.008), histological entity (P = 0.001), malignancy grade (P = 0.001), vascular invasion (P < 0.001), tumor depth (P = 0.012) and resection margins (P = 0.045) were prognostic indicators of MFS. Finally, vascular invasion (P < 0.001), tumor depth (P = 0.041) and resection margins (P < 0.001) were prognostic indicators of RFS.

The impact of the angiogenic markers on DSS, MFS and RFS in the ET group are summarized in Table 2. PDGF-A (P = 0.035), PDGF-B (P = 0.006), PDGF-C (P = 0.032), PDGF-D (P = 0.003), PDGFR-α (P = 0.002), PDGFR-β (P = 0.029), VEGF-A (P = 0.001), VEGFR-1 (P = 0.001) and FGF2 (P = 0.033) were prognostic indicators of DSS. PDGF-A (P = 0.007), PDGF-B (P = 0.003), PDGFR-α (P = 0.002), PDGFR-β (P = 0.002), VEGF-A (P = 0.001), VEGFR-1 (P < 0.001) and VEGFR-3 (P = 0.008) were prognostic indicators of MFS. PDGF-A (P = 0.012), PDGF-B (P = 0.015), PDGFR-α (P = 0.011), VEGF-A (P = 0.002) and VEGFR-1 (P = 0.036) were prognostic indicators of RFS.
Table 2

Angiogenic markers as predictors for disease-specific survival, metastasis and local recurrence in patients with resected soft-tissue sarcomas of the extremities or trunk (univariate analyses, log rank test, n = 115)

  

Disase-specific survival

Metastasis-free survival

Recurrence-free survival

Marker expression

Patients (n)

5-Year survival (%)

P

5-Year survival (%)

P

5-Year survival (%)

P

PDGF-A

       

Low

54

60

0.035

74

0.007

79

0.012

High

58

52

 

51

 

55

 

Missing

3

      

PDGF-B

       

Low

44

68

0.006

78

0.003

82

0.015

High

68

48

 

52

 

56

 

Missing

3

      

PDGF-C

       

Low

31

71

0.032

68

0.214

68

0.564

High

80

50

 

61

 

65

 

Missing

4

      

PDGF-D

       

Low

73

67

0.003

70

0.051

77

0.002

High

40

34

 

49

 

42

 

Missing

2

      

PDGFR-α

       

Low

69

67

0.002

74

0.002

77

0.011

High

43

38

 

42

 

45

 

Missing

3

      

PDGFR-β

       

Low

85

64

0.029

72

0.002

69

0.825

High

24

32

 

35

 

58

 

Missing

6

      

VEGF-A

       

Low

60

65

0.001

75

0.001

77

0.002

High

51

43

 

48

 

51

 

Missing

4

      

VEGF-C

       

Low

69

55

0.476

68

0.083

68

0.232

High

38

60

 

56

 

63

 

Missing

8

      

VEGF-D

       

Low

84

57

0.131

67

0.081

70

0.177

High

29

50

 

51

 

55

 

Missing

2

      

VEGFR-1

       

Low

67

63

0.002

77

<0.001

72

0.036

High

44

46

 

43

 

58

 

Missing

4

      

VEGFR-2

       

Low

78

58

0.332

67

0.189

71

0.240

High

28

52

 

53

 

57

 

Missing

9

      

VEGFR-3

       

Low

75

60

0.053

70

0.008

70

0.159

High

34

45

 

46

 

57

 

Missing

6

      

FGF2

       

Low

75

61

0.033

66

0.214

70

0.648

High

35

49

 

56

 

67

 

Missing

6

      

FGFR-1

       

Low

83

58

0.460

64

0.411

66

0.768

High

26

47

 

55

 

71

 

Missing

6

      

Abbreviations: PDGF Platelet-derived growth factor, PDGFR Platelet-derived growth factor receptor, VEGF Vascular endothelial growth factor, VEGFR Vascular endothelial growth factor receptor, FGF Fibroblast growth factor, FGFR Fibroblast growth factor receptor.

The impact of the clinicopathological variables on DSS, MFS and RFS in the VR group are summarized in Table 1. Age (P < 0.001), gender (P = 0.039), malignancy grade (P = 0.005) and resection margins (P = 0.021) were prognostic indicators of DSS. Gender (P = 0.022) was a prognostic indicator of MFS and tumor size (P = 0.006), malignancy grade (P = 0.046) and resection margins (P < 0.001) were prognostic indicators of RFS.

The impact of angiogenic markers on DSS, MFS and RFS in the VR group is summarized in Table 3. FGRF-1 (P = 0.023) was the only prognostic indicator for DSS and PDGF-C (P = 0.045) for RFS.
Table 3

Angiogenic markers as predictors for disease-specific survival, metastasis and local recurrence in patients with resected visceral & retroperitoneal soft-tissue sarcomas (univariate analyses, log rank test, n = 66)

  

Disase-specific survival

Metastasis-free survival

Recurrence-free survival

Marker expression

Patients (n)

5-Year survival (%)

P

5-Year survival (%)

P

5-Year survival (%)

P

PDGF-A

       

Low

23

49

0.473

63

0.593

65

0.315

High

39

63

 

54

 

73

 

Missing

4

      

PDGF-B

       

Low

14

54

0.604

82

0.088

61

0.291

High

48

59

 

51

 

73

 

Missing

4

      

PDGF-C

       

Low

20

39

0.297

59

0.986

53

0.045

High

41

65

 

56

 

76

 

Missing

5

      

PDGF-D

       

Low

48

52

0.078

52

0.197

70

0.343

High

15

80

 

73

 

59

 

Missing

3

      

PDGFR-α

       

Low

41

56

0.672

61

0.527

71

0.761

High

21

61

 

51

 

66

 

Missing

4

      

PDGFR-β

       

Low

58

57

0.360

54

0.532

70

0.766

High

4

75

 

75

 

75

 

Missing

4

      

VEGF-A

       

Low

34

51

0.326

64

0.719

64

0.054

High

29

68

 

53

 

79

 

Missing

3

      

VEGF-C

       

Low

34

66

0.402

69

0.071

62

0.051

High

29

50

 

45

 

84

 

Missing

3

      

VEGF-D

       

Low

34

60

0.856

62

0.388

63

0.116

High

30

55

 

51

 

78

 

Missing

2

      

VEGFR-1

       

Low

37

55

0.724

63

0.358

70

0.510

High

25

63

 

49

 

71

 

Missing

3

      

VEGFR-2

       

Low

44

55

0.858

63

0.446

69

0.821

High

19

67

 

48

 

75

 

Missing

3

      

VEGFR-3

       

Low

36

54

0.552

59

0.821

65

0.220

High

25

61

 

52

 

76

 

Missing

5

      

FGF2

       

Low

39

56

0.805

51

0.214

74

0.748

High

20

65

 

67

 

66

 

Missing

7

      

FGFR-1

       

Low

43

45

0.023

56

0.385

68

0.448

High

20

89

 

63

 

78

 

Missing

3

      

Abbreviations: PDGF Platelet-derived growth factor, PDGFR Platelet-derived growth factor receptor, VEGF Vascular endothelial growth factor, VEGFR Vascular endothelial growth factor receptor, FGF Fibroblast growth factor, FGFR Fibroblast growth factor receptor.

Multivariate cox proportional hazards analysis

Table 4 presents multivariate analyses of clinicopathological and angiogenic marker variables with respect to DSS, MFS and RFS in the ET and VR groups, respectively.
Table 4

Multivariate analyses of clinopathological variables and angiogenic markers as prognostic values for disease-specific survival, metastasis and local recurrence in patients with resected soft-tissue sarcomas of the trunk or extremities (cox proportional hazards test)

 

Disase-specific survival

 

Metastasis-free survival

 

Recurrence-free survival

 

Variable

HR

95% CI

P

HR

95% CI

P

HR

95% CI

P

Extremity & trunk

Malignancy grade

         

1

1.000

 

<0.001*

      

2

4.066

1.389-11.901

0.010

      

3

6.025

2.058-17.634

0.001

      

Vascular invasion

         

Absent

1.000

  

1.000

  

1.000

  

Present

2.141

1.188-3.859

0.011

5.284

2.418-11.544

<0.001

2.135

1.019-4.475

0.045

Resection margins

         

Wide

1.000

     

1.000

  

Non-wide

1.818

1.032-3.203

0.039

   

2.687

1.289-5.602

0.008

PDGF-B

         

Low

      

1.000

  

High

      

2.099

0.937-4.706

0.072

PDGF-D

         

Low

1.000

     

1.000

  

High

1.863

1.057-3.283

0.031

   

1.844

0.931-3.653

0.079

VEGF-A

         

Low

      

1.000

  

High

      

2.095

1.028-4.271

0.042

VEGFR-1

         

Low

   

1.000

     

High

   

2.106

1.038-4.272

0.039

   

Visceral & retroperitoneal

Gender

         

Male

   

1.000

     

Female

   

4.612

1.089-19.536

0.038

   

Malignancy grade

         

1

1.000

 

0.003*

   

1.000

 

0.061

2

4.812

1.823-12.705

0.002

   

2.069

0.630-6.794

0.231

3

5.646

1.790-17.804

0.003

   

5.665

1.330-24.123

0.019

Resection margins

         

Wide

1.000

     

1.000

  

Non-wide

2.712

1.222-6.018

0.014

   

11.996

3.128-46.005

<0.001

PDGF-C

         

Low

      

1.000

  

High

      

0.413

0.157-1.089

0.074

FGFR-1

         

Low

1.000

        

High

0.243

0.095-0.618

0.003

      

Abbreviations: HR Hazard ratio, CI Confidence interval, VEGF Vascular endothelial growth factor, VEGFR Vascular endothelial growth factor receptor, PDGFR Platelet-derived growth factor receptor, FGFR Fibroblast growth factor receptor, *overall significance as prognostic factor.

In the ET group, high malignancy grade (P < 0.001), the presence of vascular invasion (P = 0.011), non-wide resection margins (P = 0.039) and high expression of PDGF-D (HR = 1.863, 95% CI = 1.057-3.283, P = 0.031) were significant independent prognostic indicators of DSS. Further, the presence of vascular invasion (P < 0.001) and high expression of VEGFR-1 (HR = 2.106, 95% CI = 1.038-4.272, P = 0.039) were significant independent prognostic factors of MFS, while the presence of vascular invasion (P = 0.045), non-wide resection margins (P = 0.008) and high expression of VEGF-A (HR 2.095, 95% CI 1.028-4.271, P = 0.042) were significant independent prognostic factors of RFS.

In the VR group, high malignancy grade (P = 0.003) and non-wide resection margins (P = 0.014) were significant independent adverse prognostic indicators of DSS whereas high FGFR-1 expression (HR = 0.243, 95% CI = 0.095-0.618, P = 0.003) was an independent positive prognostic indicator of DSS. Female gender (P = 0.038) was an independent negative prognostic indicator of MFS while non-wide resection margins (P < 0.001) was an independent negative prognostic indicator of RFS.

Discussion and conclusions

In our univariate analyses high expression of most examined angiogenic markers were prognosticators of DSS and/or MFS and/or RFS in the ET group. Further, PDGF-D was an independent negative prognostic indicator of DSS, VEGFR-1 an independent negative prognostic indicator of MFS and VEGF-A an independent negative prognostic indicator of RFS. In contrast, only FGFR-1 was a prognosticator of DSS in both the univariate and multivariate analyses of the VR group. To our knowledge, this is the first comparison of the expression of angiogenic molecules in ET versus VR STSs.

Current knowledge of the importance of tumor localization (ET versusVR tumors) when it comes to the prognostic impact of angiogenic markers in STSs is limited. Yudoh et. al. investigated the level of VEGF-A in tissue from ET patients and found high levels to predict survival, local recurrence and metastasis [18]. We have previously reported on the expression of PDGFs, VEGFs and FGFs in a larger cohort of STS of mixed sites and histology and found high expression of VEGFR-3, PDGF-B and FGF2 to have independent negative prognostic impact on DSS [1416]. When comparing the expression of angiogenic markers based on tumor location, it becomes apparent that these variables almost exclusively have prognostic impact in STS arising in the ET group (Tables 2, 3 and 4). This difference could to some extent be due to a smaller number of patients in the VR group, with a resulting increased risk of false negative results. However, near all angiogenic markers showed significant prognostic impact in the univariate analyses of the ET group, whereas only FGFR-1 showed prognostic impact in the VR group. Table 1 summarizes the clinopathological values in the ET and VR groups and it is apparent that the VR group contains a higher percentage of leiomysarcomas and liposarcomas. The different distribution of histologies between the ET and VR groups might suggest that angiogenic markers have higher impact in STSs arising in ET locations. Another explanation may be that ET tumors, even the slow growing ones, will produce symptoms when they reach a certain size due to limits created by connective and muscle tissue and blood and lymph vessels. VR tumors could in contrast grow to significant size before producing symptoms. This may explain our results as VR tumors in many cases only are found after the angiogenic switch have occurred, thus the impact of angiogenic markers have been negated in these tumors.

In the PDGF-axis, all markers were prognosticators of DSS, all but PDGF-C were prognosticators of MFS and all but PDGF-C and PDGFR-β were prognosticators of RFS in the ET group (Table 2), while none of the PDGFs were prognosticators in the VR group. Further, PDGF-D was found to be an independent negative prognostic factor for DSS in the ET group. In our previous study, PDGF-B was an independent prognosticator of DSS [15], and in this study PDGF-D is an independent prognosticator of DSS. PDGF-B binds all PDGFRs while PDGF-D binds PDGFR-αβ and-ββ [11]. Both PDGF-B and PDGF-D has been shown to exhibit similar and extensive angiogenic and transforming abilities [19, 20]. Although our results cannot distinguish whether PDGF signalling drives tumor development through angiogenesis or other pathways, they strongly suggest PDGF signalling to be an important part of STS growth and progression.

In the VEGF-axis, VEGF-A, and VEGFR-1 were prognosticators of DSS, MFS and RFS in the ET group, while none of the VEGFs were prognosticators in the VR group (Table 2). Further, VEGFR-1 was an independent prognostic indicator of MFS and VEGF-A was an independent prognostic indicator of RFS in the ET group. VEGF-A signalling is the major angiogenic pathway, and high tumor expression and availability in serum has previously been associated with malignancy grade, metastasis, local recurrence and worse overall survival in STS patients [18, 2126]. VEGFR-1 is thought to modulate VEGF-A signalling through VEGFR-2, has anti-angiogenic properties in its soluble form, and has been linked to metastasis in experimental studies suggesting a feasible biological link for our finding in these STS patients [27, 28]. This latter finding is quite interesting as antibodies and small-molecules targeting VEGFR-1 are being developed [29, 30].

In the FGF-axis, FGF-2 was an unfavorable prognostic indicator of DSS in ET group. FGF2 is thought to drive cell-cycling, activate extracellular matrix remodelling and to rescue PDGF-B and VEGF-A driven angiogenesis in the presence of their respective inhibitors [13, 31, 32]. Surprisingly, FGFR-1 was an independent positive indicator of DSS in the VR group. To our knowledge these are new data, but these results have to be validated before a firm conclusion may be drawn due to the low number of patients.

This study enhances our current knowledge on angiogenic prognosticators in STSs, strongly indicates the involvement of the PDGF and VEGF pathways in ET STS development and adds to the growing body of evidence suggesting that STSs of different sites and histology should be analyzed independently in future studies. Further emphasis should also be put on validating VEGFR-1 as a predictor of MFS in ET STS patients, as these patients may benefit from adjuvant therapy targeting VEGFR-1.

Declarations

Authors’ Affiliations

(1)
Department of Oncology, University Hospital of North Norway
(2)
Institute of Medical Biology, University of Tromso
(3)
Department of Clinical Pathology, University Hospital of North Norway
(4)
Institute of Clinical Medicine, University of Tromso

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  33. Pre-publication history

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This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

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