cancers

Review
Desmoplastic Small Round Cell Tumor: A Review of Main
Molecular Abnormalities and Emerging Therapy
Celso Abdon Mello 1, *, Fernando Augusto Batista Campos 1 , Tiago Goss Santos 2,3 ,
Maria Leticia Gobo Silva 4 , Giovana Tardin Torrezan 3,5 , Felipe D’Almeida Costa 6 , Maria Nirvana Formiga 1 ,
Ulisses Nicolau 1 , Antonio Geraldo Nascimento 6 , Cassia Silva 1 , Maria Paula Curado 7 ,
Suely Akiko Nakagawa 8 , Ademar Lopes 8 and Samuel Aguiar Jr. 8

1 Department of Medical Oncology, A.C.Camargo Cancer Center, Sao Paulo 01509-010, Brazil;
fernando.campos@accamargo.org.br (F.A.B.C.); nirvana.formiga@accamargo.org.br (M.N.F.);
ulisses.nicolau@accamargo.org.br (U.N.); cassia.silva@accamargo.org.br (C.S.)
2 Laboratory of Tumor Biology and Biomarkers, International Center of Research CIPE,
A.C.Camargo Cancer Center, Sao Paulo 01509-010, Brazil; tsantos@accamargo.org.br
3 National Institute of Science and Technology in Oncogenomics and Therapeutic Innovation,
Sao Paulo 05403-010, Brazil; giovana.torrezan@accamargo.org.br
4 Department of Radiation Oncology, A.C.Camargo Cancer Center, Sao Paulo 01509-010, Brazil;
leticia.silva@accamargo.org.br
5 Genomics and Molecular Biology Group, International Center of Research CIPE, A.C.Camargo Cancer Center,
Sao Paulo 01508-010, Brazil
6 Department of Pathology, A.C.Camargo Cancer Center, Sao Paulo 01509-010, Brazil;
felipe.costa@accamargo.org.br (F.D.C.); nascimento.antonio@accamargo.org.br (A.G.N.)
7 Department of Epidemiology, A.C.Camargo Cancer Center, Sao Paulo 01508-010, Brazil;



mp.curado@accamargo.org.br

 8 Department of Surgery, A.C.Camargo Cancer Center, Sao Paulo 01509-010, Brazil;
suely.nakagawa@accamargo.org.br (S.A.N.); ademar.lopes@accamargo.org.br (A.L.);
Citation: Mello, C.A.; Campos,
samuel.aguiar@accamargo.org.br (S.A.J.)
F.A.B.; Santos, T.G.; Silva, M.L.G.; * Correspondence: celso.mello@accamargo.org.br; Tel.: +55-11-2189-2779
Torrezan, G.T.; Costa, F.DA.; Formiga,
M.N.; Nicolau, U.; Nascimento, A.G.; Simple Summary: Desmoplastic small round cell tumor is a rare neoplasm with extremely aggressive
Silva, C.; et al. Desmoplastic Small behavior. Despite the multimodal treatment for newly diagnosed patients with chemotherapy,
Round Cell Tumor: A Review of Main
cytoreductive surgery and radiation, the cure rate is still low. For relapsed or progressive disease,
Molecular Abnormalities and
there is limited data regarding second and third-line therapies. Novel agents have shown only modest
Emerging Therapy. Cancers 2021, 13,
activity. Recent molecular changes have been identified in this disease and this opens opportunities
498. https://doi.org/10.3390/
to be explored in future clinical trials.
cancers13030498

Abstract: Desmoplastic small round cell tumor (DSRCT) is an extremely rare, aggressive sarcoma
Received: 10 December 2020
Accepted: 20 January 2021
affecting adolescents and young adults with male predominance. Generally, it originates from
Published: 28 January 2021 the serosal surface of the abdominal cavity. The hallmark characteristic of DSRCT is the EWSR1–
WT1 gene fusion. This translocation up-regulates the expression of PDGFRα, VEGF and other
Publisher’s Note: MDPI stays neutral proteins related to tumor and vascular cell proliferation. Current management of DSRCT includes a
with regard to jurisdictional claims in combination of chemotherapy, radiation and aggressive cytoreductive surgery plus intra-peritoneal
published maps and institutional affil- hyperthermic chemotherapy (HIPEC). Despite advances in multimodal therapy, outcomes remain
iations. poor since the majority of patients present disease recurrence and die within three years. The dismal
survival makes DSRCT an orphan disease with an urgent need for new drugs. The treatment of
advanced and recurrent disease with tyrosine kinase inhibitors, such as pazopanib, sunitinib, and
mTOR inhibitors was evaluated by small trials. Recent studies using comprehensive molecular
Copyright: © 2021 by the authors. profiling of DSRCT identified potential therapeutic targets. In this review, we aim to describe the
Licensee MDPI, Basel, Switzerland. current studies conducted to better understand DSRCT biology and to explore the new therapeutic
This article is an open access article strategies under investigation in preclinical models and in early phase clinical trials.
distributed under the terms and
conditions of the Creative Commons Keywords: desmoplastic small round cell tumor; treatment; prognosis; surgery; radiotherapy;
Attribution (CC BY) license (https://
chemotherapy; tyrosine kinase receptor; target therapy; rare disease
creativecommons.org/licenses/by/
4.0/).

Cancers 2021, 13, 498. https://doi.org/10.3390/cancers13030498 https://www.mdpi.com/journal/cancers

 Cancers 2021, 13, x 2 of 24

Cancers 2021, 13, 498 2 of 22

1. Introduction
1. Introduction
Desmoplastic small
Desmoplastic small round
round cell
cell tumor
tumor (DSRCT)
(DSRCT) is is an
an extremely
extremely rare,
rare, aggressive
aggressive sar-
sar-
coma. It affects
coma. affects mainly
mainly adolescents
adolescents and and young
young adults
adults and
and originates
originates inin and
andprimarily
primarily
involvesthe
involves theserosal
serosalsurfaces
surfacesofofthe
theabdominal
abdominalcavity.
cavity.ItItwas
wasfirst
firstdescribed
describedby byGerald
Geraldand
and
Rosai in 1989 as a newly characterized clinicopathologic entity [1]. Current
Rosai in 1989 as a newly characterized clinicopathologic entity [1]. Current management of management
of the
the disease
disease includes
includes a combination
a combination of chemotherapy,
of chemotherapy, radiation,
radiation, andand aggressive
aggressive surgi-
surgical
cal resection
resection [2,3][2,3] as summarized
as summarized in Figure
in Figure 1. Despite
1. Despite advances
advances in multimodal
in multimodal therapy,
therapy, the
the outcome
outcome remains
remains poor poor
sincesince the majority
the majority of patients
of patients develop develop highofrates
high rates of disease
disease recur-
recurrence
rence or dieor die within
within three three
years years
[4,5]. [4,5].
Due toDuethetodismal
the dismal survival,
survival, DSRCTDSRCT hasurgent
has an an ur-
unmet need for
gent unmet needmore effective
for more and innovative
effective therapeutic
and innovative options.
therapeutic options.

Figure1.1. Therapeutic
Figure Therapeuticoptions
optionsforfordesmoplastic
desmoplasticsmall
smallround
roundcell
celltumor
tumor(DSRCT).
(DSRCT).Based
Basedon onmultiple
multiple
retrospectives
retrospectives and
and few prospective studies,
studies, the
thebenefit
benefitwas
wasobserved
observedforfortherapies
therapies
in in green,
green, lowlow
evidence of the benefit for therapies in blue. For relapsed or progressive disease, strategies in
evidence of the benefit for therapies in blue. For relapsed or progressive disease, strategies in yellow yel-
lowbeen
had had been
used used
and inand in white
white are perspectives.
are perspectives. HIPECHIPEC (hyperthermic
(hyperthermic intraperitoneal
intraperitoneal chemo-
chemotherapy),
therapy), RTKi (Receptor Tyrosine Kinase inhibition), ARi (Androgen Receptor inhibition),
RTKi (Receptor Tyrosine Kinase inhibition), ARi (Androgen Receptor inhibition), VEGFi (Vascular VEGFi
(Vascular Endothelial Growth Factor inhibition), IO (immune check point inhibition), IGFi (Insulin
Endothelial Growth Factor inhibition), IO (immune check point inhibition), IGFi (Insulin Growth
Growth Factor inhibiotion), mTORi (mammalian Target of Rapamycin inhibition), ABMT (autol-
Factor inhibiotion), mTORi (mammalian Target of Rapamycin inhibition), ABMT (autologous bone
ogous bone marrow transplant).
marrow transplant).
1.1. Demographics
1.1. Demographics of ofDSRCT
DSRCT
Desmoplasticsmall
Desmoplastic smallround
roundcell celltumor
tumorisisaaveryveryrare
raresubtype
subtypeof ofsarcoma.
sarcoma.For Forresearch
research
purposes, DSRCT cases can be searched using the histology
purposes, DSRCT cases can be searched using the histology and behavior (malignant) and behavior (malignant)
classification code
classification code 8806/3.
8806/3. There
There is no uniform
is no uniform definition
definition of of rare
rare sarcoma,
sarcoma, however,
however,the the
burden of rare cancer in our current days is great. The US Orphan
burden of rare cancer in our current days is great. The US Orphan Drug Act of 1983 defined Drug Act of 1983 de-
fined
rare rare diseases
diseases as thoseas affecting
those affecting
less thanless200,000
than 200,000
peoplepeople in the United
in the United States States
[6]. In [6].
2010,In
2010, Greenlee
Greenlee etdescribed
et al. [7] al. [7] described
the US the US burden
burden of rare cancers
of rare cancers according according to the National
to the National Cancer
Cancer Institute
Institute definition definition
as those as thosewith
cancers cancers
fewerwith
thanfewer thanper
15 cases 15 cases
100,000perpeople
100,000 perpeople
year.
More recently, a consortium from the European Union, Surveillance of Rare CancerRare
per year. More recently, a consortium from the European Union, Surveillance of in
Cancer(RARECARE)
Europe in Europe (RARECARE)
[8], described [8], described
a new a new
definition definition
of rare cancerof inrare cancer
Europe in Europe
as those with
as those
fewer thanwith fewer
6 cases perthan 6 cases
100,000 per 100,000
people per year. people per year.
Inaastudy
In studypublished
publishedin in2014
2014[9],
[9],aatotal
totalof of192
192cases
casesof ofDSRCT
DSRCTwere wereidentified
identifiedin inthe
the
SEERdatabase
SEER databasebetween
between 1973
1973 to 2007.
to 2007. The age-adjusted
The age-adjusted incidence
incidence rateonbased
rate based on this
this analysis
analysis
was was 0.3 cases/million,
0.3 cases/million, with a peakwith a peak
incidence incidence
of 0.74 of 0.7420–24
in individuals in individuals
years-old. There20–24
isyears-old.
a predominance
There is of DSRCT in males.
a predominance The age-adjusted
of DSRCT in males. The incidence rates incidence
age-adjusted for males rates
and
females
for maleswereand0.4 and 0.1were
females cases/million,
0.4 and 0.1respectively
cases/million, (p <respectively
0.001) [9]. There is predominance
(p < 0.001) [9]. There is
in African–American
predominance individuals andindividuals
in African–American it is more and common in males.
it is more commonThe inage-adjusted
males. The
incidence
age-adjustedratesincidence
are higherrates
among areAfrican-Americans as compared to Caucasians
higher among African-Americans as compared (0.5to×Cau-
0.2,
pcasians
= 0.037,(0.5
respectively). Out respectively).
× 0.2, p = 0.037, of 192 cases, the Outcommon primary
of 192 cases, sites of disease
the common primary were the
sites of
peritoneum or soft tissue of abdomen and pelvis (42%) and less common primary sites
included the ovary/fallopian tube (6 cases), orbit (1 case), cerebellum (1 case), and cerebral
ventricle (1 case) [9].
 Cancers 2021, 13, x 3 of 24

Cancers 2021, 13, 498 3 of 22

disease were the peritoneum or soft tissue of abdomen and pelvis (42%) and less common
primary sites included the ovary/fallopian tube (6 cases), orbit (1 case), cerebellum (1
case), and cerebral ventricle (1 case) [9].
1.2. Molecular Profile of DSRCT
1.2. Molecular Profile of DSRCT
Cytogenetic and molecular characterization of DSRCT identified a unique chromoso-
Cytogenetic andt(11;
mal rearrangement, molecular
22)(p13; characterization
q12), associated of with
DSRCT thisidentified a unique
tumor [10,11]. Thechromo-
EWS-WT1
somal rearrangement, t(11; 22)(p13; q12), associated with this
is the driver to tumorigenesis of DSRCT and it acts by up-regulating the expression tumor [10,11]. The of
EWS-WT1 is the driver to tumorigenesis of DSRCT and it acts by up-regulating
several genes [12]. The chimeric product of the EWS-WT1 fusion protein acts as a dominant the ex-
pression of several genes [12]. The chimeric product of the EWS-WT1 fusion
transcriptional activator factor that regulates the expression of several growth factor genes, protein acts
as a dominant
including PDGFRA, transcriptional
IGF1R, EGFR, activator factor
IL2, IL15 that
and regulates
also the expression
transcriptional of several
factors such as MYC,
growth factor genes,
PAX2 and WT1 [3,13]. including PDGFRA, IGF1R, EGFR, IL2, IL15 and also transcriptional
factors such as MYC, PAX2 and WT1 [3,13].
The up-regulation of PDGFRα is a hallmark event in the development and DSRCT.
The up-regulation of PDGFRα is a hallmark event in the development and DSRCT.
The role of PDGFRα in the physiologic healing process is well described and is responsible
The role of PDGFRα in the physiologic healing process is well described and is respon-
for collagenous stromal production, inflammatory cell infiltration, especially macrophage
sible for collagenous stromal production, inflammatory cell infiltration, especially mac-
chemotaxis and neo-angiogenesis [14], induces proliferation and is a chemo-attractant
rophage chemotaxis and neo-angiogenesis [14], induces proliferation and is a
to fibroblasts and endothelial cells [14,15]. The development and growth of DSRCT is
chemo-attractant to fibroblasts and endothelial cells [14,15]. The development and
primarily dependent on this translocation product [12]. The EWS-WT1 transcription factor
growth of DSRCT is primarily dependent on this translocation product [12]. The
translocation produces a chimeric protein that induces the expression of PDGFRα that
EWS-WT1 transcription factor translocation produces a chimeric protein that induces the
can explain the histological
expression of PDGFRα characteristics
that can of DSRCT that
explain the histological is markedof
characteristics byDSRCT
profuse stromal
that is
proliferation and increased vascular density [16] (Figure 2).
marked by profuse stromal proliferation and increased vascular density [16] (Figure 2).

Figure2.
Figure 2. Schematic
Schematic representation
representationofofEWS–WT1
EWS–WT1fusion protein
fusion mechanism
protein mechanism of action in desmo-
of action in desmoplas-
plastic
tic smallsmall round
round cellcell tumor.
tumor. Increaseinintyrosine-kinase
Increase tyrosine-kinase receptor
receptorexpression,
expression,modulation
modulation of DNA
of DNA
replication proteins, activation of DNA-Damage Repair (DRR) machinery resulting in proliferation,
replication proteins, activation of DNA-Damage Repair (DRR) machinery resulting in proliferation,
desmoplasia, neo-angiogenesis and drug resistance.
desmoplasia, neo-angiogenesis and drug resistance.

Our group published a study with comprehensive molecular profiling of a patient with
the diagnosis of DSRCT [17]. We identified genetic variants leading to protein alterations
including 12 somatic and 14 germ-line events affecting genes predominantly involved in
mesenchymal cell differentiation pathways. Regarding copy number alterations (CNA) few
events were detected, mainly restricted to gains in chromosomes 5 and 18 and losses at 11p,
Cancers 2021, 13, 498 4 of 22

13q, and 22q. We developed a personalized test to follow up the patient and monitor disease
recurrence by assessing the circulating tumor DNA (ctDNA) in the patient’s plasma. The
genomic breakpoint of the EWS-WT1 gene fusion was tracked for the presence of minimal
residual disease after surgery. This biomarker has been used in four post-treatment blood
samples, three years after surgery, and no trace of EWS-WT1 gene fusion was detected, in
accordance with imaging tests showing no evidence of disease and with the good general
health status of the patient [17]. One interesting finding of our study is the fact that 7 out
of 15 genes harboring somatic mutations (CHL, MEGF10, MEIS2, MYH8, RIMS4, TBPL1
and ZFPM2) are regulated by the same transcription factor, LEF1 (p < 0.001), which, in
turn, is regulated by WT1 [18]. We, therefore, postulate that DSRCT tumors presenting
increased activity of WT1 might up-regulate the expression of several genes mediated by
LEF1. However, the accumulation of mutations in this set of genes regulated by LEF1 and
its relationship with the EWS-WT1 fusion protein remains to be addressed.
More recently, the molecular analysis [19] of 6 patients with DSRCT revealed a total of
137 somatic mutations which were related to specific biological processes: DNA damage-
response (DDR) network and mesenchymal–epithelial reverse transition/epithelial– mes-
enchymal transition (MErT/EMT), reinforcing the relevance of these processes in tumor
heterogeneity, aggressiveness and drug resistance [19].
There are many similarities between DSRCT and Ewing Sarcoma (ES) family tumors.
Most of these tumors carry the EWS translocation. However, one important molecular
aberration that distinguishes DSRCT from ES is the increased Androgen Receptor (AR)
expression [20,21].

2. Clinical Presentation and Diagnosis

The disease predominantly originates from the peritoneum or retroperitoneum and
can invade the omentum with multiple peritoneal implants involving the diaphragm,
splenic hilum, mesentery of the small and large bowel, and the pelvic peritoneum [2,22,23].
Patients can be asymptomatic for long periods of time until symptoms of pain, ascites, con-
stipation, weight loss, distension and jaundice [2,22]. Other sites of the primary tumor are
described in the literature as the thoracic cavity, testicle, head and neck, intracranial, thigh,
axilla/shoulder, intraosseous, uterine corpus, ovary, skull, middle ear, and others [24–28].
About one-half of the patients will present extra-peritoneal metastasis at the time of the
diagnosis [9,22,29,30], although this percentage was lower in the report of Stiles et al. [23].
The liver and lung are the two most common sites for distant metastasis [2,3,23,29].
In most cases, patients with an abdominal disease are diagnosed in an advanced stage,
with large masses and/or extensive seeding in the visceral and parietal peritoneum [30,31].
Symptoms, which are related to the tumor burden and location of the lesions, motivate
investigation by image exams. The most common imaging finding is multiple, lobulated,
low-attenuated, heterogeneous peritoneal, omental and serosal soft tissue masses usually
discrete, round or ovoid, without an apparent primary organ of origin [32–34]. Almost
all patients will present a dominant mass, mainly in the retrovesical or recto-uterine
location, peritoneal or omental [33]. MRI can be helpful in delineating the extent of the
disease, if surgical resection is considered [35] and can reveal lesions with heterogeneous
contrast enhancement [36]. The role for position emission tomography (PET)-CT is not
well established in DSRCT imaging, although it has been used each more as part of staging
evaluation together with a chest CT scan [31,36]. There is no formal staging system for
DSRCT [37]. One was proposed by Hayes-Jordan et al. [38] using the Peritoneal Cancer
Index but it has not yet been validated.
Histologically, the tumor consists of solid sheets, large nests, small clumps, or cords of
cohesive, small, round, ovoid, or spindled cells, with inconspicuous nucleoli, and scant
cytoplasm, lying in a hypocellular, desmoplastic, collagenous stroma [39]. In immuno-
histochemistry, there is an expression of desmin, membrane antigen (EMA), cytokeratins
(AE1/AE3 and CAM5.2) and neural markers (neuron-specific enolase and CD57), and
smaller numbers expressing chromogranin, synaptophysin, CD56, neurofilament protein
Cancers 2021, 13, 498 5 of 22

and S100 protein [40]. DSRCT can be immunoreactive for antibodies selectively directed to
the carboxy terminus of the WT1 protein in more than 90% of cases [41]. It is important to
note that DSRCT can show a polyphenotypic immune profile as well as a marked variation
in morphologic appearances from tumor to tumor and within the same neoplasm [42].

3. Differential Diagnosis
The differential diagnosis of DSRCTs can be made with a spectrum of other round
cell neoplasms, which includes ES, rhabdomyosarcoma, small cell carcinoma and mesothe-
lioma [42]. Since the diagnosis of DSRCT is made by a combination of the histologic
appearance and immunohistochemical staining results, it can be challenging in core biopsy
specimens, once some of the distinctive features such as the prominent stromal pattern
may not be easily appreciable, and atypical immunohistochemical features can be present
due to the limited material [42]. DSRCT is distinguished from the other neoplasms by the
presence of EWS-WT1 translocation, ideally performed by PCR or FISH or by immunohis-
tochemistry staining if the former are not available. Nowadays, large fusion panels using
RT-PCR are able to help in differentiating small round cell sarcomas from the ES family
and new entities are been recognized such as tumors harboring CIC-DUX4, BCOR-CCNB3
and CIC-FOX04 fusions [43].

4. Treatment
4.1. Therapeutic Approach for Newly Diagnosed Patients
Multimodal therapy combining multi-agent intensive chemotherapy, aggressive de-
bulking surgery and adjuvant radiotherapy is considered the standard of care for patients
presenting without extra-abdominal metastases [29,44]. As most of the cases present as
intra-peritoneal, aggressive tumors, the main guidelines recommend initiating treatment
with systemic chemotherapy. Although DSRCT is not specifically addressed by ESMO [45]
or NCCN guideline, this recommendation is stated by the Chicago Consensus on Peritoneal
Malignancies [44] and by the MDAnderson Cancer Center nomogram [46].
The role of surgery in the management of DSRCT is well established in the literature.
In a review of 12 patients treated at Mayo Clinic, Hassan et al. showed the median survival
of patients treated with surgical resection was 34 months, whereas the median survival of
those who underwent biopsy alone was 14 months [47]. In the report of Wong et al., the
median survival for patients who had a resection for their abdominal or pelvic tumors was
47 months, compared to 16 months for those who did not [48]. Complete cytoreductive
surgery is associated with improved survival and should be considered a cornerstone of
treatment together with chemotherapy [46].
The most effective chemotherapeutic regimen with curative intent is still debated,
but most are based on those used to treat other small round cell sarcomas, with a com-
bination of an anthracycline, alkylating agent and vinca alkaloid. DSRCT is somehow
sensitive to chemotherapy, although a transient response followed by disease progres-
sion is common [48]. Farhat et al. [49] reported four patients with the intra-abdominal
disease who experienced disease stabilization lasting 4–9 months after chemotherapy
including cyclophosphamide, etoposide, doxorubicin and cisplatin. Kushner et al. [50]
reported 12 patients with a median survival of 19 months with the P6-protocol, which has
seven courses of chemotherapy consisting of cyclophosphamide, doxorubicin, vincristine
(HD-CAV), etoposide and ifosfamide. This was followed by surgery, radiotherapy, and
myeloablative chemotherapy using thiotepa and carboplatin with stem cell rescue in some
cases. Bertuzzi et al. [51] published a trial that included 7 patients with DSRCT treated with
induction chemotherapy consisting of ifosfamide, epirubicin and vincristine, and those
who responded were then treated with high-dose chemotherapy and autologous bone
marrow transplantation in conjunction with local therapy (surgery and/or radiotherapy).
The authors concluded that high-dose chemotherapy probably has no role in the treatment
of DSRCT [51]. More recently, Scheer et al. [52] found that patients treated with the VAIA
regimen (ifosfamide, vincristine, doxorubicin, actinomycin D) presented longer event-
Cancers 2021, 13, 498 6 of 22

free survival (29.4 months) compared to other protocols, including the P6 protocol. The
interval-compressed regimen of vincristine, irinotecan, temozolamide (VIT) was evaluated
in 6 pediatric patients and showed a tolerable profile with an objective response rate of
50% to the first 2 cycles of VIT [53].

4.1.1. Radiation Therapy

Given the propensity for morbid intra-peritoneal progression, consolidative whole
abdominopelvic radiotherapy (WAP-RT) as part of multimodal treatment (chemotherapy,
surgical debulking and WAP-RT) was first reported using the P6 protocol, in an attempt
to improve local control [50]. Honoré et al. [29], reported in a series of 38 patients with
a median follow-up of almost 5 years, that multimodal treatment combining systemic
chemotherapy, complete macroscopic resection, and postoperative WAP-RT could pro-
long survival in patients without the extra-peritoneal disease (EPM)—median survival of
37.7 months (range 7.9–42.9 months). The factors predictive of 3-year overall survival were
the absence of EPM, complete surgical resection, postoperative WAP-RT and postoperative
chemotherapy [29].
Atallah et al. [54] studied the prognostic role of WAP-RT on oncologic outcomes
as part of multimodal treatment in 103 patients with abdominal DSRCT treated at eight
French centers from 1991 to 2014. Patients were retrospectively divided into three groups
for evaluation: Group A treated with adjuvant RT after cytoreductive surgery, Group B
without RT after cytoreductive surgery, and Group C treated with chemotherapy alone.
Three-year OS was 61.2% (range 41–76%) in Group A, 37.6% (range 22–53.1%) in Group B,
and 17.3% (range 6.3–32.8%) for Group C, respectively (p < 0.001). Peritoneal progression-
free survival (PPFS) and progression-free survival (PFS) also differed significantly between
the three groups (p < 0.001), but not distant progression. They concluded that RT seems
to improve survival after cytoreductive surgery, with better PPFS, PFS and OS for the
patients treated in a multimodal approach, with the limitations of a retrospective study,
lack of statistical power (due to the small number of patients), and the need of randomized
prospective studies to confirm these results [54].
In a more recent publication, Subbiah et al. [46] reported the MDAnderson Cancer
Center experience with the treatment of 187 patients over two decades with the multimodal
approach. The five-year OS rate was substantially improved from 5% to 25% with newer
chemotherapy agents and better surgical and RT techniques. Chemotherapy response
and complete cytoreductive surgery (CCS) were associated with improved survival. Their
results also supported the use of WAP-RT when the time of diagnosis was used as a
reference to estimate OS (univariate analysis, p = 0.01; HR, 0.44) [46]. However, because
RT was given almost exclusively to patients who underwent CCS after chemotherapy,
they removed these confounding factors and assessed the effects of WAP-RT using the
date of the surgery as the start date in a time-variant analysis, and surprisingly, WAP-
RT did not improve OS. This unexpected result conflicts with current clinical practice
of a tri-modality therapy (chemo, surgery and radiation), and updated their treatment
recommendation to consider WAP-RT in highly selected patients that are prospectively
monitored in clinical trials [46]. Desai et al. [55] reported that acute toxicities of WAP-RT
were primarily gastrointestinal and hematologic, and were improved in comparisons of
IMRT against 2D-RT (gastrointestinal grade 2 or higher: 33% × 77%, p = 0.04; and grade
4 hematologic: 33% × 82%, p = 0.02), with no survival differences. Late toxicity (small
bowel obstruction) did not statistically differ between RT modalities [55]. There are two
main indications of radiation therapy in the control of DSRCT. One is in the palliative
setting and the other is in an adjuvant scenario after complete cytoreductive surgery. The
last one is the most recommended and adopted by most of the centers, despite clear
evidence about the real prognostic impact is lacking, as seen in Table 1.
Cancers 2021, 13, 498 7 of 22

Table 1. Impact of Radiation Therapy after surgical resection in the treatment of desmoplastic small round cell tumor (DSRCT).

% Surgery + RdT Dose Comparison Group

Author Year N Outcome with Addition of Radiation
Radiation (N) (Gy) (RdT × no-RdT)
Lal et al. [22] 2005 66 43% (29) 30 yes Improved 3 year OS (55 × 27%, p < 0.02)
Forlenza et al. [56] 2015 19 89% (17) 30 no 5-year OS 16%
Stiles et al. [23] 2018 102 20% (21) NR yes No difference in median OS (27.5 × 28.8 mo, p = 0.32)
Subbiah et al. [46] 2018 165 42% (69) NR yes No improvement in OS (3.6 × 2.9, p = 0.38)
Honore et al. [5] 2019 100 26% (26) 30 yes Improved survival and cure rate (HR = 0.36, p = 0.00013)
Improved median PPFS (22.5 × 14.2 mo, p = 0.024),
Atallah et al. [54] 2016 49 * 55% (27) 20–33 yes
no significant difference in OS (p = 0.40)
Campos et al. [57] 2020 19 21% (4) 30 no 5-year OS 12%

RdT—radiation therapy; NR—not reported; mo—months; OS—overall survival; PPFS—peritoneal progression-free survival; * only patients
achieving complete cytoreductive surgery.

4.1.2. HIPEC
Due to the rarity of this disease and analysis based on retrospective series, the role
of radiotherapy in the management of DSRCT is still controversial. Appropriate patient
selection is critical, as severe toxicities can occur. Despite aggressive multidisciplinary
approaches, patients have a poor prognosis. Prospective randomized multicenter studies
will be needed to evaluate the role of local treatments such as RT in the course of the disease.
Even after chemotherapeutic cytoreduction and surgical resection of gross, visible
disease, the microscopic residual disease is often present [4]. Hence, hyperthermic intra-
peritoneal chemotherapy (HIPEC) has been examined as an adjunctive intraoperative
strategy. In a recent phase 2 trial, 14 DSRCT patients were treated with neoadjuvant
chemotherapy, followed by cytoreductive surgery (CRS), which was complete (CR0) or
near- complete (CR1 ≤ 2.5 cm of tumor remaining) in all patients, with closed technique
HIPEC using 100 mg/m2 of cisplatin for 90 min at 41 degrees Celsius, then followed by
WAP-RT [58]. The 3-year overall survival from time of diagnosis for DSRCT patients was
79%, and the estimated median recurrence-free survival (RFS) was 14.0 months. In 100% of
patients without hepatic or portal metastasis, there was no peritoneal disease recurrence
after CRS-HIPEC. They concluded that CRS, HIPEC and WART are effective local control
therapy in DSRCT patients. Earlier, it was demonstrated that patients who had CR0 or
CR1 and HIPEC had significantly longer median survival compared with patients who
had HIPEC and gross residual disease greater than 2.5 cm after surgical cytoreduction
(63.4 vs. 26.7 months) [59]. In an important retrospective study conducted at MDAnderson
Cancer Center, most of the patients (72%) received combined treatment with chemotherapy,
surgery plus HIPEC and radiation therapy. This study was underpowered to conclude
about the impact of HIPEC on the OS. However, there was not found improvement in the
three- and five-year OS (p = 0.12) of patients treated with HIPEC. Patients with DSRCT
and disease outside the abdomen at the time of surgery do not benefit from HIPEC [59].
Until now, there is no randomized trials designed to evaluate the relative contribution to
improve outcome after complete surgical excision of intra-abdominal implants.
A retrospective study with 187 DSRCT patients confirmed that chemotherapy and
CCS remain the cornerstone of treatment, and suggest that prospective randomized studies
will be required to prove the unequivocal benefit of HIPEC or WAP RT in the management
of DSRCT [46].

4.2. Prognosis
Despite multimodal treatment, DSRCT has a poor prognosis, and approximately
60–70% of patients die due to disease progression usually within 3 years after diagno-
sis [5,23,58,60]. The median overall survival varies between 28–60 months, with a median
disease-free survival between 10–15.5 months [23,29,57,58,60]. Table 2 shows a comparison
of multimodal treatment in DSRCT.
Cancers 2021, 13, 498 8 of 22

Table 2. Summary of studies with multimodal therapy for first-line treatment of patients with desmoplastic small round
cell tumor.

Author Year N Study Design/Region Therapy Overall Survival Relapse

Lal et al. [22] 2005 66 Retrospective, single center, USA ChT, CRS, RdT 5-year 15% NR
Forlenza et al. [56] 2015 19 Prospective, single center, USA ChT, CRS, BMT, RdT 5-year 16% 3-year EFS 11.0%
Osborne et al. [60] 2015 32 Retrospective, Single center USA ChT, CRS, RdT 5-year 38% 3-year EFS 9.9%
Honore et al. [61] 2017 48 Retrospective ChT, CRS, HIPEC, RdT 5-year 19% 5-year DFS 12%
Prospective, multicenter, Germany,
ChT, CRS, BMT,
Scheer et al. [52] 2018 60 Poland, Austria, 3-year 30% 3-year EFS 11.0%
RdT HIPEC
Sweden, Switzerland
ChT, CRS, BMT
Stiles et al. [23] 2018 125 Retrospective, multicenter, USA 5-year 10% NR
HIPEC RdT
ChT, CRS, BMT,
Subbiah et al. [46] 2018 165 Retrospective, single center, USA 5-year 25% NR
HIPEC, RdT
Retrospective, multicenter,
Honore et al. [5] 2019 100 ChT, CRS, HIPEC, RdT 5-year 5% 3-year DFS 7.0%
France
Median DFS 10
Campos et al. [57] 2020 19 Retrospective, single center, Brazil ChT, CRS, HIPEC RdT 5-year 12%
months
ChT—chemotherapy, CRS—cytoreductive surgery, BMT—autologous bone marrow transplant, HIPEC—hyperthermic intraperitoneal
chemotherapy, RdT—radiation therapy, DFS—disease-free survival, EFS—event-free survival, NR—not reported.

One study proposed that the absence of extra-peritoneal metastasis, complete surgical
resection and postoperative WAP-RT are factors predictive of 3-year overall survival [29].
The multimodality treatment combining chemotherapy, cytoreductive surgery, HIPEC and
WAP-RT can warrant local control of the disease, but patients will still present distant
metastasis during follow-up, meaning that more effective chemotherapy is necessary to
improve long-term outcomes [58].

4.3. Current and Emerging Therapy for Relapsed or Progressive Disease

DSRCT is characterized by poor response to conventional chemotherapy and early
relapse after radical surgery. Second-line treatment is ineffective in most cases. In our
cohort, out of 19 patients treated with first-line chemotherapy, 13 received the second-line
and the progression-free survival was only 3.9 months [57]. This short survival time high-
lights the aggressiveness of this disease and the challenge in developing new therapeutic
strategies to treat these young patients. Despite the development of new regimens for ES
and other soft tissue and bone sarcoma in recent years, DSRCT is underrepresented or were
not included in the trials that lead to the drug approval [62]. As a result, the evidence to use
second-line therapy is very limited. It is paramount to develop active cooperative groups
to quickly collect data and propose new strategies for the treatment of DSRCT. Moreover,
patients outside Europe and North America are almost never offered the opportunity to
participate in clinical trials for rare diseases. The SELNET (https://selnet-h2020.org/),
that is a Horizon 2020/EU project, aims to create a network between European and Latin
American countries to improve diagnosis and treatment of sarcomas and eventually to
develop clinical trials in these centers across the Atlantic Ocean. Table 3 displays the
outcome of main studies with medical treatment and explores the response rate according
to the line of treatment.
Cancers 2021, 13, 498 9 of 22

Table 3. Selected studies evaluating the efficacy of systemic treatments in patients with DSRCT according to the line of
treatment.

Median Age/Range
Author, Year Line of Treatment N Systemic Therapy Response Survival
(Years)
80% SD
Farhat et al., 1996 [49] 1st line 5 22/16–26 PA(E)VEP * mRFS: 6 mo
20% CR
50% PR
Bertuzzi et al., 2003 [63] 1st line 10 29/NA IVE * 20% SD NA
30% PD
78% SD
mPFS = 12.8 mo
Forlenza et al., 2015 [56] 1st line 19 18.5/10–42 P6-protocol * 11% MR
3-year OS = 26 ± 10%
11% PD
mTTP = 18.1 mo
Magnan et al., 2017 [64] 1st line 15 NA ITB RR = 27%
3-year OS = 61%
P6-protocol * P6-protocol: EFS = 12.9 mo
Scheer et al., 2019 [52] 1st line 60 14.5/6–38 VAIA * NA ** VAIA: EFS = 29.4 mo
CEVAIE * CEVAIE: EFS = 12 mo
Kushner et al., 1996 [50] 1st and 2nd line 12 14/7–22 P6-protocol * 83% PR mPFS > 12 mo
Bond et al., 2008 [65] 1st line and beyond 10 16/3–29 Imatinib Mesylate 100% PD NA
6% PR
Tap et al., 2012 [66] 1st and beyond 16 33/19–63 Ganitumab mPFS = 19 mo
63% SD
Vinorelbine/
Casanova et al., 2004 [67] 2nd line 1 17 PR mPFS > 6 mo
Cyclophosphamide
Bisogno et al., 2006 [68] 2nd line and beyond 3 10.6/1–18.5 Irinotecan 100% PD NA
25% PR
Italiano et al., 2013 [69] 2nd line and beyond 8 23/14–58 Sunitinib 37.5% SD mPFS > 4 mo
37.5% PD
33% SD mPFS = 3.2 mo
Verret, B. et al., 2017 [70] 2nd line and beyond 6 25/19–52 Trabectedin
67% PD mOS = 4 mo
3% CR
3% PR mPFS = 5.6 mo
Menegaz et al., 2018 [71] 2nd line and beyond 38 25/5–48 Pazopanib
55% SD mOS = 15.7 mo
38% PD
80% PR
Tarek et al., 2018 [72] 3rd line and beyond 5 15/11–28 VCT mTTP = 8.5 mo
20% SD
PFS = 10 mo
Thijs et al., 2010 [73] 4th line 1 21 Temsirolimus SD
OS = 13 mo

Chemotherapeutic regimens: PA(E)VEP (cyclophosphamide, etoposide, doxorubicin or epirubicin, and cisplatin); IVE (ifosfamide,
vincristine, and epirubicin); P6-protocol (high-dose cyclophosphamide, doxorubicin, and vincristine, alternating with ifosfamide, and
etoposide); ITB (irinotecan, temozolomide, and bevacizumab); VAIA (ifosfamide, vincristine, adriamycin, actinomycin D); CEVAIE
(ifosfamide, vincristine, actinomycin D, carboplatin, epirubicin and etoposide); VCT (vinorelbine, cyclophosphamide, and temsirolimus).
NA: not available; CR: complete response; PR: partial response; MR: mixed response; SD: stable disease; PFS: progression-free survival; OS:
overall survival; RFS: relapse-free survival; TTP: time-to-progression; EFS: event–free survival; RR: response rate; mo: months; m: median.
* After induction chemotherapy, patients underwent additional therapies, which could include myeloablative chemotherapy followed
by autologous stem cell transplant, surgery, and/or radiotherapy. ** VAIA scheme correlated with an increased chance of R0 or R0/R1
resection.

4.3.1. The Importance of Pre-Clinical Models to Drug Development for Rare Sarcoma
The use of preclinical models is an important step in the development of new ther-
apies for tumors in general and is crucial for rare tumors such as DSRCT. Due to the
rarity of DSRCT, conducting clinical trials with new drugs is extremely difficult for many
reasons [74]. First, the fact that genetic and functional comprehensive analyses of these
tumors are limited. Second, the aggressive behavior and chemotherapy resistance makes
the inclusion in target therapy clinical trials difficult. Accrual of patients in clinical trials is
difficult due to the limited number of individuals affected yearly. As a result, the use of
pre-clinical models is an important step in the development of novel drugs since a higher
number of mechanisms can be modulated and faster therapeutic targets can be explored.
Modeling these tumors with experimental models allows the investigation of the molecular
mechanisms that underlie tumor origin and progression. The fidelity of the model is also
related to the predictive capacity to anticipate eventual effects of drugs which will help to
determine the efficiency and efficacy of anti-tumoral drugs. In the case of rare tumors, the
possibility of scaling-up is crucial to translate technology from the bench to the bedside.
Cancers 2021, 13, 498 10 of 22

In 2002, Nishio and collaborators [75] reported for the first time the development of
the DSRCT cell line, named JN-DSRCT-1, derived from pleural effusion from a 7-year-old
patient with lung metastasis. JN-DSRCT-1 cells are small, round or spindle-shaped with
oval nuclei and were maintained continuously in vitro for more than 190 passages for more
than 40 months. The cell has a tumorigenic capacity and the histology of heterotransplanted
tumors in SCID mice maintains the characteristics of the original DSRCT, including the
expression of immunohistochemical markers (vimentin, desmin, CD57, among others), a
t(11; 22) translocation (p13; q12) and presence of EWS-WT1 fusion [75]. JN-DSRCT-1 cells
are being used in several studies that help to unveil DSRCT biology, especially the role of
EWS-WT1 fusion protein and also to identify targets for therapeutic interventions [12,76].
As mentioned before, EWS-WT1 translocation is the major driver in DSRCT and plays
many roles in tumor biology that have the potential to be used as therapeutic targets. As al-
ready described in other studies, EWS-WT1 gene underwent RNA splicing and one variant
lacks three amino-acids and was named EWS-WT1(-KTS) due to the absence of Lys-Thr-Ser
residues [77]. This isoform activates a gene encoding a tetraspanin-family protein, T-cell
acute lymphoblastic leukemia-associated antigen 1 (TALLA-1) [78]. TALLA-1 is part of
multi-protein family involved in several processes, such as cell adhesion, migration and
metastasis, and this gene could be a candidate for diagnostic marker and a putative target
for therapy [78]. Another target for EWS-WT1 fusion gene is ENT4 (equilibrative nucleoside
transporter 4) which encodes a pH-dependent adenosine transporter [79]. Neural gene
induction is also triggered by EWS-WT1 in JN-DSRCT-1 cells and neural reprogramming
factor ASCL1 is an important player in mediating multiple WT1-responsive elements,
suggesting that neural differentiation pathway could be tested as therapeutic agents for
DSRCT [80]. Recent work described the dependence of EWS-WT1 in DSRCT survival [12].
Silencing EWS-WT1 causes proliferation loss, growth arrest and gene expression analysis
indicates repression of estrogen signaling and highlights therapeutic genetic vulnerabilities,
such as FGFR4, JAK3, mTOR, PDGF, ERG, and TGFB1 genes [12]. Another study that eval-
uated potential therapeutic targets performed RNA sequencing of 12 tumor samples from
pediatric patients with DSRCT found high expression of IGF2, FGFR4, CD200 and CD276,
the latter two molecules are candidates for immune checkpoint inhibitor therapy [81].
In terms of therapy, the first use of JN-DSRCT-1 cells was to test the effect of rapamycin-
induced apoptotic death [82]. The mechanism involves the up-regulation of Bax concomi-
tant Bcl-xL down-regulation. Rapamycin also down-regulates EWS-WT1 and 26S p44.5
proteasome subunit, suggesting that rapamycin induces apoptosis by preventing the degra-
dation of the Bax protein by the proteasome, and that this process is independent of
mTOR inhibition. Furthermore, these results strongly support the introduction of the use
of rapamycin as a cytotoxic agent for the treatment of DSRCT. JN-DSRCT-1 cells were
tested to verify the effectiveness of the TRAIL receptor agonist (apoptosis inducer), called
ONC201 [83]. In this study, it was found that the induction of TRAIL decreases prolifer-
ation and induces apoptosis in vitro and decreases tumor growth in vivo. The potential
of anti-angiogenic agents in decreasing tumor growth of the JN-DSRCT-1 cell was also in-
vestigated [74]. Animals with JN-DSRCT-1 cell xenografts were treated with bevacizumab
and showed a prolongation of the time to progression and the long-term regressions were
marked after treatment with the combination of irinotecan and bevacizumab compared
to irinotecan alone [74]. Interestingly, there is recent evidence that indicates that the use
of other anti-angiogenic agents may be effective in the treatment of DSRCT, as in the case
report of a patient with advanced DSRCT, in the second-line of treatment, refractory to
cisplatin, was treated with apatinib (VEGFR-2 inhibitor drug) and had a positive response,
with a significant reduction in tumor mass [84]. JN-DSRCT-1 cells are sensitive to alkylating
agent trabectedin and the mechanism of action involves the expression of genes involved
with proliferation and apoptosis [85]. An alternative mechanism of action of trabectedin is
the impairment of transactivation of FUS-CHOP fusion protein in liposarcoma [86]. This
activity is also observed in EWS-WT1 fusion protein, trabectedin reduces its binding on
its target gene promoters and, thus, affects EWS-WT1—dependent gene expression in
Cancers 2021, 13, 498 11 of 22

JN-DSRCT-1 cells [85]. Recently, the combination of PARP inhibitor olaparib with the
alkylating agent temozolomide was tested in JN-DSRCT-1 cells in vitro and in vivo and the
results indicates that the combination has synergistic effects upon cell viability, inducing
cell cycle arrest which progress to apoptosis induction, causing tumor reduction [76].
Additionally to JN-DSRCT-1 cells, Markides and collaborators established more
DSRCT cell lines, including BER lineage that also presents EWS-WT1 fusion protein and
have similar behavior of JN-DSRCT-1 [12,87]. Together, this evidence shows the impor-
tance of obtaining tumor models to accelerate preclinical research, bring possibilities for
investigating new therapeutic approaches for this rare but lethal malignancy.

4.3.2. Targeting Angiogenesis and Other TKR

As a hypervascular tumor, DSRCT is characterized by an overexpression of proteins
that promote and maintain the angiogenic process necessary for continued tumor growth
and proliferation. EWS-WT1 is able to induce PDGFA expression [21] and activation of the
IGF1R gene [14] (Figure 2). Other tyrosine kinase receptors (TKR) expression have been
found to be disrupted in DSRCT and are related to proliferation and angiogensis. VEGFR-2
and VEGFA expression was found to be markedly increased in the DSRCT tumor sample
and in the human DSRCT cell line, JN-DSRCT [88].
The use of tyrosine kinase inhibitors (TKI) for VEGF, VEGFR, PDGFRα and other
proteins involved in tumoral vascular proliferation has been explored in the clinical sce-
nario [84,89,90]. Pazopanib, apatinib and sunitinib inhibit angiogenesis by abrogating
the VEGF-induced phosphorylation of VEGF receptors as well as other TKRs including
PDGFR, FGFR, and c-KIT, affecting downstream activation of the PI3K/AKT, PKC, and
other pathways that mediate cell proliferation, migration, and survival [91].
In the PALETTE study, 369 patients were randomized to receive pazopanib 800 mg/day
versus placebo [62]. Median PFS was 4.6 months (95%CI: 3.7–4.8) for pazopanib compared
with 1.6 months. A combined analysis of patients with the diagnosis of DSRCT treated
in the EORTC phase II study 62,043 (3 patients), EORTC phase III 62,072 (3 patients) and
in a UK Pazopanib expanded access program (3 patients) was performed [89]. Data from
nine patients included in this analysis revealed a median age of 30 years and all patients
were males with widespread metastatic DSRCT. Four patients had one previous chemother-
apy line (44%), four had 2 previous chemotherapy lines (44%) and one patient 3 (12%).
The response rate was partial response (PR) in 2/9 (22%) patients, stable disease (SD)
in 5/9 patients (56%) and progressive disease (PD) in 2/9 (22%) with a clinical benefit
rate (PR + SD > 12 weeks) of 78%. Median PFS and OS were 9.2 (95% CI: 0–23.2) and
15.4 (95% CI: 1.5–29.3) months respectively [89]. In a relatively large, retrospective study
with 29 patients treated with pazopanib, clinical benefit was observed in 62% (18/29) of
patients with DSRCT (CR in 1 patient, PR in 1 patient, SD in 16 patients) and the median
progression-free survival was 5.4 months [71].
Sunitinib was one of the first generation of TKIs with great inhibition of VEGF receptor
2 among other TKRs. In vascular sarcomas, sunitinib showed early and promising activity
in alveolar sarcoma, a chemo-resistant subtype of sarcoma [92]. In a retrospective analysis
of patients with DSRCT, sunitinib showed clinical benefit in 8 patients evaluated [69].
Partial response was observed in two patients and SD in 3 patients treated in the second-
and-beyond-line of therapy. Sorafenib was used in 2 patients [90], both in the fifth-line of
therapy. The best response to sorafenib was stable disease and the median progression-free
survival of 3.5 and 4 months. On the other hand, as first-line treatment, apatinib was used
in only one patient [84]. Apatinib is another VEGFR-2 inhibitor with demonstrated activity
in gastric and other tumors [93]. Clinical benefit and tumor shrinkage were reported in one
patient treated with apatinib in the first-line. The patient had not received chemotherapy
previously [84]. Another report showed partial response with apatinib in combination with
chemotherapy as second-line treatment [94]. Ramucirumab, a VEGFR inhibitor is been
tested in combination with cyclophosfomide and vinblastine in patients with relapsed and
refractory DSRCT (ClinicalTrials.gov Identifier: NCT04145349).
Cancers 2021, 13, 498 12 of 22

Bevacizumab, a VEGF-A inhibitor, was combined with irinotecan and temozolamide

(ITB regimen) in the first-line treatment of DSRCT. In this single-arm pilot study, 14 out of
15 patients completed the planned treatment that comprised two cycles of ITB followed by
the conventional trial P6 with VAC and IE. The response rate to ITB was 27% and no major
unexpected adverse event was observed [64].
Most of the trials with novel agents are designed to treat a myriad of histologies,
including DSRCT. It is difficult to identify trials accruing only patients with this disease.
We provide a summary with selected ongoing trials accruing patients with, but not limited
to, DSRCT, displayed in Table 4.

4.3.3. Targeting Androgen Receptor Pathway

The increased prevalence of DSRCT in young males motivated the investigation of the
testosterone synthesis pathway in tumorigenesis of this disease. In 2007, Fine et al. [95] first
demonstrated AR expression in DSRCT. In a cohort of 27 heavily pretreated patients, 37%
stained positive for AR. The functionality of the pathway was demonstrated by in vitro
assay that showed growth of tumor cells when stimulated by di-hydro-testosterone, and
inhibition of growth by flutamide [95]. Another study performing single-sample gene set
enrichment analysis found that the majority of DSRCTs were enriched for the AR signature
when compared to other sarcomas, such as ES and alveolar rhabdomyosarcoma [3,20].
In the previous reported Fine et al. [95] study, six patients with AR-positive DSRCT
received combined androgen blockade (CAB) with bicalutamide and leuprorelin. Three
patients had clinical tumor benefits for a period lasting 3 to 4 months. All of them had
normal testosterone levels at the initiation of CAB therapy, while the other three non-
responders had castrated levels. In another report, a patient with strong AR expression
received anti-androgen therapy with bicalutamide, presenting progressive disease 2 months
later [73]. Negri et al. [21], using whole genome gene expression profiling and a cancer stem
cell gene array, showed that AR-positive DSRCT cells harbor characteristics of stemness,
which could explain the limited effectiveness of targeting this pathway.

4.3.4. Targeting PI3K/AKT/mTOR Pathway

Activation of the phosphatidylinositol-3-kinase (PI3K)-protein kinase B (Akt)-mammalian
target of rapamycin (mTOR) pathway is proposed to be implicated in the development of
a variety of sarcomas [96–98]. There is emerging data indicating the possible involvement
of the mTOR pathway in DSRCT. A single-center small study attempted to evaluate the
morphoproteomic profiling of the mTOR pathway in DSRCT, ES and Wilms’ tumor, and
showed that the PI3K/Akt/mTOR pathway is constitutively activated in DSRCT [99]. Another
study described a patient with DSRCT harboring a secondary somatic mutation in the PIK3CA
gene [100].
In vitro study demonstrated that rapamycin, an mTOR inhibitor, induced the apoptotic
death of DSRCT line cells [82]. There are few data on the clinical efficacy of inhibition of
PI3K/AKT/mTOR pathway. In one case report, a 21-year-old man with DSRCT achieved
stable disease with temsirolimus for 40 weeks [73]. Tarek et al. [72] reported their experience
with five patients with relapsed DSRCT treated with vinorelbine, cyclophosphamide and
temsirolimus, all of them presenting partial response, with median time to progression of
8.5 months (range 7–16 months). A phase I trial evaluated the combination of cixutumumab
(an IGFR antibody) with temsirolimus, which resulted in stable disease lasting longer than
5 months in two of the three patients with DSRCT of the study [101]. In a retrospective
series of patients with high-grade STS treated with pazopanib plus sirolimus following
progression on pazopanib, one patient with DSRCT had stable disease for 11 months with
the combination treatment [102]. Recently, a trial was designed to estimate the response
rate to two initial courses of temsirolimus, temozolomide and irinotecan (window therapy)
in previously untreated patients with high-risk ES family of tumors, including DSRCT
(ClinicalTrials.gov Identifier: NCT01946529). The interim analysis determined the window
therapy did not meet the anticipated response, and trial accrual was stopped.
Cancers 2021, 13, 498 13 of 22

Table 4. Selected trials including desmoplastic small round cell tumor (DSRCT) patients.

Phase of Trial Design Primary Outcome ClinicalTrials.gov Identifier

Phase 1/2 Ramucirumab IV + Cyclophosphamide p.o. + Vinorelbine IV (experimental arm), versus Cyclophosphamide p.o. + Vinorelbine IV 1. Progression-fFree survival NCT04145349
2 cycles of the investigational combination irinotecan, temozolomide and bevacizumab, will be given followed by conventional
1. Tolerability
Phase 1 chemotherapy with a modified P6 approach and surgical local control. Completion of modified P6 chemotherapy will be followed by NCT01189643
2. Adverse event profile
a second-look surgery.
Experimental arm A: Single dose of IP RIT administered through an IP catheter with 131 I-omburtamab at 80 mCi/m2 , followed by
WA-IMRT approximately 2–4 weeks after completing IP RIT
Phase 2 1. Progression-free survival NCT04022213
Experimental arm B: Single dose of IP RIT administered through an IP catheter with 131 I-omburtamab at 80 mCi/m2
Experimental arm C: Single dose of IP RIT administered through an IP catheter with 131 I-omburtamab at 80 mCi/m2
1. Recommended phase II does of Prexasertib
Phase 1/2 Dose Escalation/Dose Expansion Study of Prexasertib in Combination with Irinotecan 15 mg/m2 IV daily × 10 days in 21 day cycles NCT04095221
2. Response
Nab-paclitaxel will be administered as follows:
Age ≥ 21: 125 mg/m2 days 1, 8 and 15 in cycles of 28 days 1. Overall response rate
Phase 2 NCT03275818
Age ≥ 6 months and ≤ 20 years: 240 mg/m2 (for patients weighing > 10 kg) and 11.5 mg/kg (for patients weighing ≤ 10 kg) on days 2. Objective response rate
1, 8 and 15 in cycles of 28 days
Participants will receive vincristine, doxorubicin, cyclophosphamide, ifosfamide, etoposide, irinotecan, temozolomide, temsirolimus,
Participants with DSRCT will not be included in the
Phase 2 bevacizumab, and sorafenib. Depending on the size and location of the participant’s tumor, they will have surgery alone, radiation NCT01946529
analysis of primary outcome
alone or surgery followed by radiation.
1. Transplant-related mortality
Phase 2 Allogeneic Hematopoietic Stem Cell Transplantation 2. Rate of grade III or higher organ toxicity attributable NCT04530487
to conditioning
1. To assess the feasibility of HIPEC with doxorubicin
Patients undergo cytoreduction and HIPEC over 60 min consisting of doxorubicin and cisplatin. Patients then receive sodium and cisplatin after surgical resection.
Phase 1 NCT04213794
thiosulfate IV over 12 h. 2. To assess morbidity, hospital length of stay and
peri-operative mortality outcome.
1. Safety and tolerability
2. Determine the MTD
Experimental arm A: participants will receive B7H3-specific CAR T cells only
Phase 1 3. Assess the DLT and describe the full toxicity profile NCT04483778
Experimental arm B: participants will receive CAR T cells directed at B7H3 and CD19
4. Assess the feasibility of manufacturing B7H3 and
B7H3xCD19 specific CARs
1. Estimate the MTD and DLT
Experimental arm A: participants will receive EGFR-specific CAR T cells only. 2. Assess the number of successfully manufactured
Phase 1 NCT03618381
Experimental arm B: participants will receive CAR T cells directed at EGFR and CD19 EGFR806 and EGFR806xCD19 CAR T cell products
3. Safety
Phase 2 Nivolumab 240 mg IV every 2 weeks plus Ipilimumab 1 mg/m2 IV every 6 weeks 1. Response to therapy as evaluated by RECIST 1.1 NCT02982486
Phase 2 Reduced-intensity chemotherapy, haploidentical bone marrow, post-transplant cyclophosphamide and shortened duration tacrolimus 1. Safety NCT01804634
Phase 1 CLR 131 intravenous administration 1. Number of participants with DLT NCT03478462

IV: intravenous; IP RIT: intraperitoneal radioimmunotherapy; MDT: maximum tolerated dose; DLT: dose limiting toxicity.
Cancers 2021, 13, 498 14 of 22

4.3.5. Targeting DNA Damage Repair (DDR) Proteins

Studies using next-generation sequencing characterized a subgroup of DSRCT with
secondary genomic alterations in genes associated with DNA damage repair (DDR), in-
cluding ATM, RAD50, BARD1, BRCA1/2, PALB2 and CHEK2 [19,103]. It is still unknown if
those genomic alterations act as driver mutations in DSRCT tumorigenesis [103].
Since poly(ADP-ribose) polymerases (PARPs) perform an important role in DDR,
specifically in the base excision repair of single-strand DNA breaks, PARP inhibitors
recently emerged as a new treatment for cancer-based on synthetic lethality concept,
particularly in BRCA-mutant cancers defective in homologous repair [104,105]. DSRCT has
high-level of PARP1, the most abundant enzyme of the PARP family, and a combination
of olaparib and temozolamide has demonstrated enhanced antitumor effects in vitro and
in vivo [76,106,107].
It was described that EWS-FLI1 in ES and EWS-WT1 in DSRCT might share common
mechanisms of gene expression de-regulation [100]. EWS-WT1 up-regulates the expression
of ERG, an ETS family member of FLI1. It is possible that ERG may drive the expression of
these targets, making this tumor an ETS-like tumor [15]. Another characteristic observed
in vitro is that the tumor modulates the DNA damage response, both suppressing p53 sig-
naling and driving the expression of gene sets associated with the DNA damage response,
suggesting a direct link to the resistance to chemotherapy in DSRCT [12]. Deregulation
of DNA damage response is another important feature of the other FET family fusions
such as EWS-FLI1 in ES. This characteristic makes the use of PARP inhibitor an attractive
therapeutic strategy in both ES and DSRCT. The combination of trabectedin and PARP
inhibitor is under investigation in clinical trials. Trabectedin is an intercalating DNA agent
that promotes DNA damage and deregulate several repair pathways, inhibits transcriptor
factors such as FUS-CHOP factor a exert cytotoxic activity in certain subtypes of cells such
as the tumor-associated macrophages (TAM), myxoid liposarcoma and ES [108,109]. The
combination of trabectedin and olaparib has shown robust inhibition of tumor proliferation
in sarcoma mouse models [110]. The heavy damage in the single strand or double-strand
DNA was not repaired by olaparib in the experimental model [110]. A phase Ib trial
(TOMAS) conducted by the Italian Sarcoma Group was designed to explore the synergistic
effect of trabectedine and olaparib in patients with advanced sarcoma and showed promis-
ing results [111]. Out of 50 accrued, 11 had a diagnosis of bone tumor and of these only
4 with ES and no activity was observed in this group of patients, despite the biological
rationale for this combination in ES. The efficacy of this combination is under evaluation
by the phase II trial TOMAS2 (ClinicalTrials.gov Identifier: NCT03838744).
In an ongoing trial, prexarsetinib, an inhibitor of checkpoint kinase 1 (chk1) in combi-
nation with irinotecan and temozolamide is currently under evaluation in an early phase
trial (ClinicalTrials.gov Identifier: NCT0409522).

4.3.6. Targeting c-MET and Insulin Growth Factor Pathway

c-Met (mesenchymal-epithelial transition factor) has been found to be overexpressed
in a variety of solid tumors, including sarcomas [112–115], but the involvement of this
receptor in DSRCT development is still scarce [100]. In the largest DSRCT comprehensive
genomic profile study, no secondary mutation on c-MET was found [103]. There is a
case report of a patient with intra-abdominal DSRCT who received anlotinib, a multi-
kinase inhibitor that targets c-Met, for the progressive disease after surgery and first-line
chemotherapy showing stable disease for 4 months [116].
The oncogenic fusion product EWSR1-WT1 in DSRCT was reported to activate the
IGF-1R gene promoter, providing the basis to test the activity of anti-IGF-1R antibodies
in the metastatic setting [117]. IGF2 has been up-regulated by the fusion product and is a
potential target [81]. In a phase II study, ganitumab administered to 16 metastatic DSRCT
patients determined one PR (6%) and 10 (63%) SD, with a median PFS of 15 months [66].
Cancers 2021, 13, 498 15 of 22

4.3.7. Cancer Vaccines

Although initial studies on cancer vaccines had shown disappointing results with low
response rates, a better understanding of the interaction between tumor, microenvironment,
and immune system in the last decades have opened new perspectives for this therapy,
including in the sarcoma field [118,119]. Cancer vaccines seek to induce tumor immune
responses through antigen presentation and stimulation of new T cell responses [118]. Few
studies have explored vaccines in DSRCT treatment. A phase I study using a vaccine of
tumor lysate-pulsed autologous dendritic cells in the treatment of pediatric patients with
solid tumors demonstrated feasibility for generating specific T-cell responses and regression
or stabilization of metastatic disease in some patients, but failed to prevent progressive
disease in the patient with DSRCT included in the trial [120]. A more recent trial evaluated
the efficacy of an adjuvant dendritic cell vaccine administered three to eight weeks after
completion of standard treatment in pediatric patients with high-risk sarcomas [121]. The
survival advantage was demonstrated for patients with ES and rhabdomyosarcoma, but
no clinical benefit was seen in the two patients with DSRCT in the study [121].

4.3.8. Perspectives with Novel Targets (Immune Checkpoint and NTRK Inhibitors)
In general, sarcomas are not considered good candidates for immune therapy the
way this therapy has currently been applied for other tumors [122]. Around one-third
of sarcomas are characterized by single gene translocation that acts as a driver mutation.
Moreover, sarcomas are amongst the neoplasms with the lowest tumor mutational burden,
a recognized predictor of response to immune check point inhibitors (IO) [123].
Initial data using the combination of anti-PD1 +/− anti-CTLA-4 have shown promis-
ing results, particularly in alveolar soft part sarcoma (ASPS), undifferentiated pleomorphic
sarcoma (UPS) and dedifferentiated lipossarcoma [124,125]. The ALLIANCE trial demon-
strated that nivolumab monotherapy is not effective with only 5% of objective response
rate against 16% for the combination of nivolumab and ipilimumab [125]. In the SARC28
trial, pembrolizumab induced objective response rate in 5 UPS patients [124] and a post
hoc analysis of this trial demonstrated that patients with a specific sarcoma immunolog-
ical classification (SIC) based on gene expression analysis derived the greatest benefit
from pembrolizumab. This immune “high” or “hot” signature was associated with the
structure of T lymphocyte, follicular dendritic cells and enriched B cells [126]. The combi-
nation of immune check point inhibitor and TKIs was evaluated and showed a safe profile
and promising activity in a phase II trial with predominant ASPS patients [127]. Out of
11 patients with ASPS, six achieved a partial response (54.5%, 95% CI 24.6–81.9), and two
(18%) of 11 achieving stable disease with a median PFS of 12 months [127].
It is difficult to predict that IO will be active in DSRCT based on preliminary data about
immune regulation and biomarkers expression in this rare disease. Gene expression analy-
sis revealed that DSRCT is characterized by a signature of immunological ignorance [21].
However, a study with samples of paraffin embed tumor sample showed elevated PD1
expression by tumor cells [128]. The expression of PD1, PDL-1, and CD8 was analyzed
in a cohort of 11 patients with DSRCT and it was observed a high rate of PD1 (81%),
CD8 (64%) and low PDL-1 (18%) expression [128]. Additionally, in vitro assay using the
JN-DSRCT-1 cells culture to test the activity of nivolumab showed no effect in decreasing
tumor cell proliferation. It is in line with the most recent data that demonstrated the
single-agent nivolumab was ineffective for most sarcoma subtype [125]. On the other hand,
the combination of an anti-PD1 and anti-CTLA4 (ipilimumab) resulted in a better out-
come [129]. However, preclinical data using gene expression analysis showed that DSRCT
overexpresses immune regulatory proteins such as CD200 and CD276 (B7H3), which is not
regulated by the EWS-WT1 fusion protein [81]. Currently, enoblituzumab is being tested
in many solid tumors (ClinicalTrials.gov Identifier: NCT02982941), including pediatric
patients with DRSCR, the results of this trial is pending. It has previously described that
DSRCT shows high expression of CD276 (B7H3) [81] and it was the basis for a phase I trial
designed to evaluate the safety, pharmacokinetics and bio-distribution of intra-peritoneal
Cancers 2021, 13, 498 16 of 22

radio-immunotherapy with a monoclonal antibody anti-B7H3 131 I-omburtamab in patients

with DSRCT or other B7H3 positive neoplasms with peritoneal involvement. The results
with 48 treated patients with DSRCT showed this approach is tolerable and the maximum
tolerated dose was not reached [130]. A phase II trial was advocated based on these results.
The neurotrophic tyrosine kinase receptor (NTKR) fusions act as driver mutation is
a myriad of neoplasms. The use of NTRK inhibitors has demonstrated robust activity
across many histologies [131]. Sarcoma was one of the most common tumor type included
in the larotrectinib trials [132] and the search for NTRK fusion in sarcoma patients are
recommended based on priority criteria [131]. Recently, it was demonstrated that EWS-
WT1 promotes direct NTRK3 transcription and in vivo and in vitro inhibition of NTRK3
cells with entrectinib resulted in significant decrease in cell growth. This finding opens the
possibility for future trials with NTRK inhibitor in DSRCT [133].

5. Conclusions
In summary, DSRCT is a rare and aggressive disease and fatal for the majority of the
patients. A modest improvement in survival has been observed in more recent studies. For
second-line treatment, there is no standard of treatment and the results with conventional
chemotherapy and novel agents are disappointing. A better understanding of disease biology
has identified potential targets to be explored in future clinical trials. It is paramount the work
of the cooperative group to organize prospective databanks and conduct clinical trials.

Author Contributions: Conceptualization, C.A.M., F.D.C., T.G.S.; methodology, writing—original

draft preparation, C.A.M., T.G.S., F.D.C., M.L.G.S.; writing—review and editing, C.A.M., F.A.B.C.,
G.T.T., T.G.S., F.D.C., M.N.F., U.N., A.G.N., S.A.N., A.L., S.A.J., C.S., M.P.C.; supervision, C.A.M. All
authors have read and agreed to the published version of the manuscript.
Funding: This study was supported by grants from the National Institute of Science and Technology
in Oncogenomics and Therapeutic Innovation (INCITO) funded by the São Paulo Research Foun-
dation (FAPESP—grant number 2014/50943—1), National Council for Scientific and Technological
Development (CNPq–grant number 465682/2014—6) and Coordination for the Improvement of
Higher Education Personnel (CAPES—88887.136405/2017—00). TGS is supported by the FAPESP
grant (2018/25541—8) and GTT is supported by the FAPESP grant (2018/06269—5). ACCCC is a
member of the TELNET consortium, sponsored by Horizon2020/EU (EU proposal 825806).
Institutional Review Board Statement: Not applicable.
Informed Consent Statement: Not applicable.
Data Availability Statement: No new data were created or analyzed in this study. Data sharing is
not applicable to this article.
Acknowledgments: We would like to thank Sarcoma European & Latin American Network (SEL-
NET), a consortium granted by European Commission (number: SEP-210512885), for supporting the
idea of this special issue.
Conflicts of Interest: C.A.M. declares advisory board and speaker fee from MSD, Roche, Bayer, Lilly,
Servier, Merck Serono, Amgen. Clinical Trial honoraria from BMS, Servier, Roche.

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