Angiomatoid Fibrous Histiocytoma: a Single Institution Case-series
Conditions: Angiomatoid Fibrous Histiocytoma
Sponsor: Istituto Ortopedico Rizzoli

AFH 
 
 
 
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“Angiomatoid Fibrous Histiocytoma: a single institution case-series” 
 
Study code AFH 
Sponsor’s Name and 
Address: 
Istituto Ortopedico Rizzoli 
Via di Barbiano 1/10 
40136 Bologna 
Italy 
Study 
Number/Version/Date: 
Vers 1.0  
03 Sep 2018 
Coordinating Center: IRCCS Istituto Ortopedico Rizzoli 
Pathology Unit 
Via Pupilli 1 
40136 Bologna, Italy 
Coordinating 
Investigator and 
address: 
Marco Gambarotti MD 
Phone: 051-6366.652 
Email: marco.gambarotti@ior.it 
Methodology: Experimental study with biological material 
(Single insitution case series review of clinical and 
histological data) 
Type: Academic 
 
Founding: None 
 
Principal Investigator 
Signature 
I confirm that I’ve read this protocol and I accept to run the study 
in compliance with what is stated in the protocol and with the 
ICh-GCP and all applicable law 
 
Marco Gambarotti MD 
 
Firma 
 
_________________________ 
 
 

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TABLE of CONTENTS 
Background 
 
 
 
 
 
 
 
 
 
pg. 3 
Objective of the study 
 
 
 
 
 
 
 
pg.  4 
Study design   
 
 
 
 
 
 
 
 
pg  5 
Population     
 
 
 
 
 
 
 
 
pg.  5 
Material and Methods   
 
 
 
 
 
 
 
pg.  5 
Statistics 
 
   
 
 
 
 
 
 
 
pg.  5 
Enrolment procedure   
 
 
 
 
 
 
 
pg.  5 
Data Collection            
  
 
 
 
 
 
 
pg.  5 
Ethic and quality assurance   
 
 
 
 
 
 
pg.  6 
Informed Consent   
 
  
 
 
 
 
 
 
pg.  6 
General principles for human biological material (HBM) collection 
 
pg.  7 
Confidentiality                                                                                       
 
 
pg.  67 
Publication of results                                                        
 
 
             pg.  8 
Sponsor role and responsibility                                                        
 
 
 pg.  8 
References 
                                                                                       
 
 
pg.  8 
 
 

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BACKGROUND 
One of the many diagnostic challenges in soft tissue pathology is the AFH – the Angiomatoid 
Fibrous Histiocytoma. It is a rare tumor of intermediate biological potential and uncertain 
differentiation accounting for approximately of 0,3% of all soft tissue tumors (Thway & Fisher, 
2015). The AFH is a histiocytoid cell lesion with primary histological characteristics which include in 
the first place a lymphocytic cuff, a fibrous pseudocapsule and pseudoangiomatoid spaces with 
hemorrhage and hemosiderin deposition. Unusual but often present morphological features are 
also a sclerotic matrix, a myxoid stroma, the presence of osteoclasts and/or a reticulated pattern. 
Nuclear atypia, mitosis and eosinophils can also be found (Bohman, Goldblum, Rubin, Tanas, & 
Billings, 2014; Kao et al., 2014) 
Its name still stems from when it was first described by Enzinger in 1979 as an ‘Angiomatoid 
malignant fibrous histiocytoma' to acknowledge for its potential for metastasis and local 
recurrence and it was first classified as a malignant tumor although later on its proven actual 
relatively low risk for distant metastasis (<5%) was considered and the word 'malignant' was 
removed from its name, hence 'AFH' (Bohman et al., 2014; Thway & Fisher, 2015). Clinically, its 
true potential for local recurrence is being estimated to be as high as 15% (Thway & Fisher, 2015). 
Although the tumor cells stain in a higher percentage for Desmin (50-92%), EMA (50-60%), CD68 
(33-80%) and CD99 (up to a 100%), those markers are very little specific and molecular techniques 
such as FISH and PCR are recommended to make the diagnosis of an AFH with either one of the 
three translocations EWSR1-CREB1 (72%); EWSR1-ATF1 (21%) and FUS-ATF1 (7 %) being almost 
pathognomonic even though other tumors such as Clear cell sarcoma, Primary pulmonary myxoid 
sarcoma or the Hyalinizing clear cell carcinoma of the salivary gland harbor these mutations as well  
(Antonescu et al., 2007; Bohman et al., 2014; Chen et al., 2011; Kao et al., 2014; Rossi et al., 2007; 
Tanas et al., 2010; Thway & Fisher, 2015). There is a certain number of AFH which are negative for 
all rearrangement with FISH and it is being suggested that cryptic rearrangements of these genes 
undetectable by FISH or the existence of other translocations involving other genes account for 
these cases (Tanas et al., 2010; Thway & Fisher, 2015). Most likely FISH doesn't allow for detection 
of rearrangement due to the presence of submicroscopic insertions or because the genetic 
material simply is undetectable to the commercial break-apart probes (Thway & Fisher, 2015). RT-
PCR is recommended to be performed in conjunction with FISH and even though FISH with break 
apart probes is faster and according to some more sensitive RT-PCR is far more specific as it also 

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identifies the fusion partner (Noujaim et al., 2017; Thway & Fisher, 2015). Useful negative 
immunohistochemical markers are Cytokeratins, S100, CD31 and Myogenin (Bohman et al., 2014). 
AFH are most often found in the dermis or the subcutaneous tissue of the extremities of children 
and young adults. But it has a wide age range (infants – adults in the 9th decade) and a part from 
other somatic localizations such as trunk, head and neck non-somatic locations have also been 
reported such as lung, mediastinum, retroperitoneum, vulva, omentum, bone and ovary (Chen et 
al., 2011; Shi et al., 2015; Tataroglu, Culhaci, & Cecen, 2015; Thway & Fisher, 2015; Thway, 
Stefanaki, Papadakis, & Fisher, 2013). 
Differential diagnosis for the AFH are tumors or lesions such as inflammatory myofibroblastic 
tumor, follicular dendritic cell sarcoma, poorly differentiated carcinoma, meningioma, lymph node 
metastasis, hematoma, granulomatous inflammation, myoepithelioma, schwannoma, primary 
pulmonary myxoid sarcomas, Ewing sarcoma, rhabdomyosarcoma, malignant rhabdoid tumor or 
aneurysmal fibrous histiocytoma (Chen et al., 2011; Kao et al., 2014; Shi et al., 2015; Thway & 
Fisher, 2015). 
Recent case reports and studies of the metastatic potential of theses tumors have raised the 
question of and the necessity for a molecular or immunohistochemical marker or specific 
histological criteria that could correlate with the worse outcome for this particular group of 
patients. According to those studies metastasis was found in regional lymph nodes, bone and lung 
(Martinez, Moreno, Vinson, Dodd, & Brigman, 2016; Tay et al., 2016; Thway et al., 2013). 
Also, in a recent study Cheah et al. reported on the results of ALK-expression of their series of 
eleven AFHs which showed that ALK expression in those tumors is common which had initially 
caused misdiagnosis as inflammatory myofibroblastic tumor which is one of the differential 
diagnostics of AFH (Cheah et al., 2018). 
 
OBJECTIVE OF THE STUDY  
Primary Objective 
Primary objective is to present our series of AFH's with its clinical, histo-morphological and 
molecular characteristics of these tumors, in order to better understand their characteristics and to 
identify the best therapeutic approach.  
Secondary Objective 
Secondary objective is  to reproduce the results of ALK expression from the study of Cheah et al. on 
our series of 9 AFH's cases. 

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STUDY DESIGN 
This is single institution cases series review of histological and clinical data 
POPULATION 
Inclusion criteria 
1) Male and female patients treated at Rizzoli Institute from 01 January 2006 to 31 
December 2017 
2) Diagnosis of Angiomatoid Fibrous Histiocytoma 
3) Histological slides/formalin-fixed paraffin-embedded tissue tumor (FFPE) blocks from 
archive available to perform the histology analysis 
4) Written informed consent prior to any study-specific analysis and/or data collection  
Exclusion criteria 
1) Patients with histological diagnosis different from Angiomatoid Fibrous Histiocytoma 
 
MATERIAL AND METHODS 
We will retrieve from the archives of the Rizzoli Institute all the cases with a histological diagnosis 
of Angiomatoid Fibrous Histiocytoma. By reviewing our database, we aspect to find approximately 
nine cases (actually all adults). We will review all the medical records, radiological imaging, and 
histological slides of all of these cases.  In all cases with enough available material, 
immunohistochemistry will be performed on FFPE material.  EWS-CREB1 and EWS-ATF1  mutation  
and ALK expression will be analyzed with RT-PCR and/or FISH analysis. 
 
STATISTICS 
To the case series will be applied a descriptive statistic. 
 
ENROLLMENT PROCEDURE 
Patients considered eligible will be included in the study, after providing a written informed 
consent.  
Due to the incidence of mortality of the disease under investigation, it would be possible that 
some eligible subjects will be deceased. 
 
DATA COLLECTION 
Clinical data will be retrieved by patient charts. 

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A protocol-specific CRF reporting the results of the review will be provided. 
A CRF is required and should be completed for each included subject. 
 
ETHICS AND QUALITY ASSURANCE 
The clinical trial protocol and its documents will be sent before initiating the study to the 
competent Authorities and Ethics Committees of each participating country for its approval. 
The responsible investigator will ensure that this study is conducted in agreement with either the 
most updated Declaration of Helsinki and all the international and local  laws that apply to clinical 
trials and to patient protection.  
The protocol has been written, and the study will be conducted according to the principles of the 
ICH Harmonized Tripartite Guideline for Good Clinical Practice  
(ref: http://www.emea.eu.int/pdfs/human/ich/013595en.pdf).  
 
INFORMED CONSENT 
All patients will be informed, by the investigator, of the aims of the study, the possible risks and 
benefits that will derive from the study participation.  
The Investigator must clearly inform that the patient is free to refuse participation in the study and 
that can withdraw consent at any time and for any reason.  
They will be informed as to the strict confidentiality of their patient data, but that their medical 
records may be reviewed for trial purposes by authorized individuals other than their treating 
physician.  
The informed consent procedure must conform to the ICH guidelines on Good Clinical Practice. 
This implies that "the written informed consent form should be signed and personally dated by the 
patient or by the patient's legally acceptable representative".   
The Investigator must also sign the Informed Consent form, and will keep the original at the site 
and a copy of the original must be handed to the patient.  
The competent ethics committee for each Institution participating to the study must validate local 
informed consent documents before the study can be opened. It will be emphasized that the 
participation is voluntary and that the patient is allowed to refuse further participation in the 
study whenever he/she wants. This will not prejudice the patient's subsequent care.  
Due to the high incidence of mortality of the disease under investigation, it would be possible that 
some potential eligible subjects will be deceased.  

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GENERAL PRINCIPLES FOR HUMAN BIOLOGICAL MATERIAL (HBM) COLLECTION 
Human biological material (HBM) collection involves the collection and storage of biological 
material, residual biological material or derivatives in compliance with ethical and technical 
requirements.  
Biological material (FFPE blocks of tumor sample) are already stored in the archive of the Istituto 
Ortopedico Rizzoli, Pathology Department. 
The biological material will be used and stored according with the sample characteristic and 
applicable regulation. 
 The Istituto Ortopedico Rizzoli will have a designated person responsible for collection and 
will act as a communication point  
 The collected HBM should be documented, i.e. the amount remaining and its location. act 
as a communication point  
 
CONFIDENTIALITY 
In order to ensure confidentiality of clinical trial data as disposed the national and European 
applicable regulation, data will be only accessible for the trial Sponsor and its designees, for 
monitoring/auditing procedures, the Investigator and collaborators, the Ethics Committee of each 
corresponding site and the Health Authority. 
Investigator and the Institution will allow access to data and source documentation for 
monitoring, auditing, Ethic Committee revision and inspections of Health Authority, but 
maintaining at all times subject personal data confidentiality as specified in the “Directive 
95/46/EC of the European Parliament and of the Council of 24 October 1995”.  
 The Investigator must guarantee that patient anonymity is kept at all times and their identity 
must be protected from unauthorized persons and institutions.  
All patients included in the study will be identified with a numeric code, so that no identifiable 
personal data will be collected  (pseudo anonymization) 
The Investigator must have and conserve a patients’ inclusion registry where it figures the 
personal data of the patient: name, surname, address and corresponding identification code into 
the study, this register will be kept on the Investigator File. 
 
 
 

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PUBBLICATION OF RESULTS  
The results from this study will be published or shown at scientific conferences. The final 
publication of the study results will be written by the Principal Investigator. 
 
SPONSOR ROLE AND RESPONSIBILITY 
The sponsor is the sole owner of the data and is responsible of all the clinical trial activities from 
study design, development, data collection, management, analysis, interpretation of data, writing 
and the decision to submit the report for publication written by the Principal Investigator,  
 
REFERENCES 
Antonescu, C. R., Dal Cin, P., Nafa, K., Teot, L. A., Surti, U., Fletcher, C. D., & Ladanyi, M. (2007). EWSR1-
CREB1 is the predominant gene fusion in angiomatoid fibrous histiocytoma. Genes, Chromosomes & 
Cancer, 46(12), 1051–1060. https://doi.org/10.1002/gcc.20491 
Bohman, S. L., Goldblum, J. R., Rubin, B. P., Tanas, M. R., & Billings, S. D. (2014). Angiomatoid fibrous 
histiocytoma: An expansion of the clinical and histological spectrum. Pathology, 46(3), 199–204. 
https://doi.org/10.1097/PAT.0000000000000073 
Cheah, A. L., Zou, Y., Lanigan, C., Billings, S. D., Rubin, B. P., Hornick, J. L., & Goldblum, J. R. (2018). ALK 
Expression in Angiomatoid Fibrous Histiocytoma: A Potential Diagnostic Pitfall. The American Journal 
of Surgical Pathology. https://doi.org/10.1097/PAS.0000000000001103 
Chen, G., Folpe, A. L., Colby, T. V., Sittampalam, K., Patey, M., Chen, M. G., & Chan, J. K. C. (2011). 
Angiomatoid fibrous histiocytoma: Unusual sites and unusual morphology. Modern Pathology, 24(12), 
1560–1570. https://doi.org/10.1038/modpathol.2011.126 
Kao, Y. C., Lan, J., Tai, H. C., Li, C. F., Liu, K. W., Tsai, J. W., … Huang, H. Y. (2014). Angiomatoid fibrous 
histiocytoma: Clinicopathological and molecular characterisation with emphasis on variant 
histomorphology. Journal of Clinical Pathology, 67(3), 210–215. https://doi.org/10.1136/jclinpath-
2013-201857 
Martinez, S. J., Moreno, C. C., Vinson, E. N., Dodd, L. G., & Brigman, B. E. (2016). Angiomatoid fibrous 
histiocytoma: novel MR imaging findings. Skeletal Radiology, 45(5), 661–670. 
https://doi.org/10.1007/s00256-016-2344-4 
Noujaim, J., Jones, R. L., Swansbury, J., Gonzalez, D., Benson, C., Judson, I., … Thway, K. (2017). The spectrum 
of EWSR1-rearranged neoplasms at a tertiary sarcoma centre; Assessing 772 tumour specimens and 
the value of current ancillary molecular diagnostic modalities. British Journal of Cancer, 116(5), 669–
678. https://doi.org/10.1038/bjc.2017.4 
Rossi, S., Szuhai, K., Ijszenga, M., Tanke, H. J., Zanatta, L., Sciot, R., … Hogendoorn, P. C. W. (2007). EWSR1-
CREB1 and EWSR1-ATF1 fusion genes in angiomatoid fibrous histiocytoma. Clinical Cancer Research, 
13(24), 7322–7328. https://doi.org/10.1158/1078-0432.CCR-07-1744 
Shi, H., Li, H., Zhen, T., Zhang, F., Dong, Y., Zhang, W., & Han, A. (2015). Clinicopathological features of 
angiomatoid fibrous histiocytoma: A series of 21 cases with variant morphology. International Journal 
of Clinical and Experimental Pathology, 8(1), 772–778. 

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Tanas, M. R., Rubin, B. P., Montgomery, E. A., Turner, S. L., Cook, J. R., Tubbs, R. R., … Goldblum, J. R. (2010). 
Utility of FISH in the diagnosis of angiomatoid fibrous histiocytoma: A series of 18 cases. Modern 
Pathology, 23(1), 93–97. https://doi.org/10.1038/modpathol.2009.138 
Tataroglu, C., Culhaci, N., & Cecen, E. (2015). Angiomatoid fibrous histiocytoma: case report and review of 
the literature. The Turkish Journal of Pediatrics, 57(1), 102–104. 
Tay, C. K., Koh, M. S., Takano, A., Aubry, M. C., Sukov, W. R., & Folpe, A. L. (2016). Primary angiomatoid 
fibrous histiocytoma of the lung with mediastinal lymph node metastasis. Human Pathology, 58, 134–
137. https://doi.org/10.1016/j.humpath.2016.07.022 
Thway, K., & Fisher, C. (2015). Angiomatoid fibrous histiocytoma: The current status of pathology and 
genetics. Archives of Pathology and Laboratory Medicine, 139(5), 674–682. 
https://doi.org/10.5858/arpa.2014-0234-RA 
Thway, K., Stefanaki, K., Papadakis, V., & Fisher, C. (2013). Metastatic angiomatoid fibrous histiocytoma of 
the scalp, with EWSR1-CREB1 gene fusions in primary tumor and nodal metastasis. Human Pathology, 
44(2), 289–293. https://doi.org/10.1016/j.humpath.2012.08.008