Abdelhay, E.

Binato, R.

Bouzas, L.F.

Du Rocher, B.

Dutra, T.

Mencalha, A.

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In Search for Molecules Involved in the Immunossupression Induced by Mesenchymal Stromal Cells 

Poster Session I S265acute GVHD rodent models (Kovacsovics 2008, 2009, Metheny,
2011).
Methods:This is an open label Phase I clinical dose escalation study
with the primary goal to assess safety of MultiStem as an adjunct
therapy for adult hematological malignancy patients shortly after al-
logeneic HSCT. Patients were enrolled for stromal cell administra-
tion as a single dose or in multiple weekly doses. Infusional toxicity
andRRTswas assessed for 30 days following the last dose. Secondary
endpoints included incidence of acute GVHD, infection and survival
through day 100. Dose escalation was guided by the Continual Re-
assessment Method (CRM).
Results: Enrollment was completed for the target of 36 patients
from 5 clinical centers for this Phase I clinical trial. 18 patients re-
ceived a single dose MultiStem IV at 1, 5, or 10 million cells per
kg at day 2 and 18 patients received MultiStem IV at 1 or 5 million
cells per kg at days 2, 9 and 16 or days 2, 9, 16 and 30 after allogeneic
HSCT. There was no observed infusional toxicity in either arm.
Two patients in the single dose arm experienced Bearman RRTs
(Grade 3 mucositis; Grade 3 renal and pulmonary failure). These
events were deemed unrelated to study product. In the multi-dose
cohort, one patient experienced liver GVHD, deemed possibly re-
lated to study product. Engraftment occurred in all 36 patients and
the median time to neutrophil engraftment was 15 days (range, 11-
25 days). The 100-day cumulative incidence of Grade II-IV and
III-IV GVHD was 28% and 6%, respectively in the single dose
arm. Preliminary evaluations of the repeat dose arm showed 21%
and 11% Grade II-IV and III-IV GVHD, respectively. The highest
tested single and repeat dose regimens showed the lowest GVHD
frequencies.
Conclusion: Single dose and repeat dose administration of Multi-
Stem is well-tolerated, without observation of infusional toxicity or
graft failure. The observed low incidence of severe acute GVHD
supports the concept that this stromal stem cell therapy product is
safe and can be harnessed as a novel therapeutic option for GVHD
prophylaxis following HSCT.
166
TREATMENT OF STEROID RESISTANT GRADE II TO IV ACUTE GVHD BY
INFUSION OF MESENCHYMAL STROMA CELLS EXPANDED WITH HUMAN
PLASMA AND PLATELET LYSATE – A PHASE I/II STUDY
Boome, L.C.J.1, Mansilla Puerta, C.M.2, Lindemans, C.A.3,
Slaper, I.C.M.4, Rozenmuller, H.4, Petersen, E.J.1, Bierings, M.3,
Boelens, J.J.3, Wulffraat, N.H.3, Kuball, J.H.E.1,2 1UMCU, Utrecht,
Netherlands; 2UMCU, Utrecht, Netherlands; 3UMCU, Utrecht,
Netherlands; 4UMCU, Utrecht, Netherlands
Introduction: For numerous hematological diseases allogeneic
HSCT is the only curative therapy. Despite multiple improvements
in the last decade in the field of HSCT, aGVHD remains a life-
threatening complication. In particular, the outcome of patients
with severe steroid-resistant aGVHD is very poor. Therefore, it re-
mains important to search for new therapeutic strategies for the
treatment of aGVHD.
Objective: To study the feasibility, safety and efficacy of MSCs ex-
panded with human plasma and platelet lysate (hPPL), in patients
with steroid-refractory aGVHD.
Method: In an open-label, non-randomized prospective phase I/II
study patients with steroid-refractory aGVHD grade II to IV were
treated with2x106/kgMSC. Response rate, TRM, and AEwere as-
sessed for up to 1 yr after inclusion. Serum and bloodsamples from
were collected.
Results: Between January 2009 and December 2010, 20 patients
were included, 2 drop out, and 18 were available for further analysis:
5 children and 13 adults. Median age was 32.5yr (range 1.3-65.9).
Organs involved in aGVHD were skin (67%), GI-tract (83%) and
liver (28%). Overall grade was II for 22%, III for 72%, and IV for
6% patients. 1 patient received one infusion, all other patients re-
ceived two or more infusions. Median follow-up was 5.5m (range
0.33-12). Complete response was observed in 11 patients 61%,
after a median of 65 days (range 10-184 days). The OS was signifi-
cantly better in responders when compared to non-responders
(p\0.001). Of the 11 patients who reached a CR, 8 patients relapsed
approximately 2 months after reaching CR (median 59 days, range:
1-244). Three children relapsed with clinical signs of an allo-
immune-lung, auto-immune-cytopenia or limited cGVHD and all5 adults relapsed with GVHDof the gut (median 98 days after reach-
ing CR, range: 35-302 days). However, GVHD of the gut was then
again sensitive to steroids. Overall, 7 patients died, 4 due to progres-
sion of aGVHD, 1 patient due to abdominal bleeding and 2 due to
sepsis.
Extensive biomarkeranalysis were performed. We found bio-
markers who are associated with clinical response of patients.
Conclusion: Generation and infusion of MSCs in steroid-resistant
aGVHD grade II- IV is feasible, safe and very effective. In addition,
also patients who initially responded toMSCs but develop later a re-
lapse of aGVHDduring tapering or cessation of immunosuppressive
drugs become again sensitive to the treatment with steroids. Clinical
response can be predicted by biomarkers.
167
IN SEARCH FOR MOLECULES INVOLVED IN THE IMMUNOSSUPRESSION
INDUCED BY MESENCHYMAL STROMAL CELLS
Du Rocher, B.1,2, Mencalha, A.1,2, Binato, R.1,2, Dutra, T.1,2,
Bouzas, L.F.1, Abdelhay, E.1,2 1 Instituto Nacional de Cancer, Rio de
Janeiro, Brazil; 2Universidade Federal do Rio de Janeiro, Rio de Janeiro,
Brazil
Background:Mesenchymal stromal cells (MSCs) improve hemato-
poietic recovery, contribute to tissue regeneration and posses immu-
nosuppressive properties. Because of these unique properties their
applicability in the bone marrow transplantation has attracted
much attention, especially as a promising therapy to control graft-
versus-host disease. In this way, a better understanding of how
MSCs induce immunosuppresion can give us the rational needed
for the most appropriate use of these cells.
Objectives:This study aimed to search for soluble factors produced
by MSCs that can potentially be involved in the mechanism of
immunosuppression.
Methods:Mixed leucocyte reactions, were incubated in the absence
or presence of 10% MSCs in a contact independent manner using
a polycarbonate membrane (transwell) between MLRs and MSCs.
After three or seven days, cultures were analysed using flow cytomety
or submited to global gene expression analyses.
Results: We found 672 mRNAs increased and 311 mRNAs de-
creased in at least 2X. 70% of mRNAs increased in MSCs in co-
culture were related to immune response. Of these, it was notorious
the increased in mRNAs associated with antigen presentation via
MHC I and II (8%), chemokine (9%), metabolism / transport of
lipids (9%) and regulatory proteins induced by IFN-y (18 %).
The pathways most likely to be activated inMSCs after co-culture
were IFN-y and IL-17. Among the 672molecules increased inMSCs
we choose COX-2 and the chemokines CCL8 e CXCL8 to evaluate
their functions during the immunosuppression induced by MSCs.
The use of indomethacin, a COX inhibitor, in co-cultures reversed
the inhibitory effect of MSCs. Moreover, when separated from
MLRs by a transwell of 0.5mM, which allows the passage of cells,
we observed a greater suppression of those lymphocytes that mi-
grated to the MSCs’s niche, suggesting that chemokines are impor-
tant in this process.
Conclusions: After co-culture MSCs changed from an inactivated
state, ‘‘steady state’’, to an activated state. This process induced their
immunosuppressive phenotype, which involves COX2 and differents
chemokines. The specific role of CCL8 and CXCL8 is under inves-
tigation.
Financial support:MS, CNPq, and FAPERJ.168
MESENCHYMAL STROMAL CELLS IMPAIR THE DIFFERENTIATION OF
CD14++CD16-CD64+ CLASSICALMONOCYTES INTO CD14++CD16+CD64++
ACTIVATE MONOCYTES
Du Rocher, B.1,2, Mencalha, A.1,2, Gomes, B.E.1,2, Bouzas, L.F.1,
Abdelhay, E.1,2 1 Instituto Nacional de Cancer, Rio de Janeiro, Brazil;
2Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil
Background:Mesenchymal stromal cells (MSCs) possess immuno-
modulatory activity both in vitro and in vivo. However, little
information is available about their function during the initiation
of immunological responses through their interactions with


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In Search for Molecules Involved in the Immunossupression Induced by Mesenchymal Stromal Cells

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