Page 23 - Read Online

Page 2 of 16 Merhi et al. J Cancer Metastasis Treat 2021;7:42 https://dx.doi.org/10.20517/2394-4722.2021.80

Methods: Blood and bone marrow samples were collected in 45 patients with distinct subtypes defined by French
American British classification, i.e., M0, M1, M2, M3, M4, and M5. Levels of MMPs and VEGF-A in leukemic blood
cells and culture supernatants were determined by RT-PCR, ELISA, and gelatin zymography (MMPs). The balance
between cell death and survival was assessed by flow cytometry with analysis of phosphatidylserine
externalization and caspase-3 activation.

Results: The administration of HF promoted a caspase-associated apoptosis in primary AML blasts (from blood
and bone marrow), but not normal blood cells and monocytes. In addition, HF inhibited the levels of secreted
proMMP-2, proMMP-9, and VEGF-A without altering transcripts. The induction of apoptosis by HF significantly
paralleled the inhibition of MMP-2/9 and VEGF-A release by HF. No differences were seen in response to the
deleterious effects of HF between AML cells of distinct subtypes.

Conclusion: Our results suggest that HF, through its proapoptotic and potential antiangiogenic properties (by
inhibiting MMP-2/9 and VEGF-A) on primary AML cells, might be a useful experimental agent, in combination
with existing drugs, for new therapeutic approaches in the treatment of this incurable disease.

Keywords: Acute myeloid leukemia, apoptosis, hyperforin, matrix metalloproteinase, VEGF, secretion

INTRODUCTION
Angiogenesis, characterized by the formation of new vessels from preexisting blood vessels, participates in
the progression of many malignant tumors by supplying oxygen and nutrients . Angiogenesis induction is
[1]
[2-4]
observed in several hematologic malignancies including acute myeloid leukemia . Endothelial cells (ECs)
[5,6]
and tumor cells secrete proangiogenic molecules . Among the positive regulators of angiogenesis, vascular
endothelial growth factors (VEGFs), and more specifically VEGF-A, play a prominent role to sustain
angiogenesis [1,5,6] . By binding to VEGF receptors at the surface of ECs, VEGF activates cell signaling
pathways involved in the activation of the proliferation and survival of ECs [1,2,7] . In addition to VEGF, matrix
metalloproteinases (MMPs) are regulators of pathologic angiogenesis [8-10] . MMPs enable extravasation and
migration of newly formed ECs and release VEGF bound to the extracellular matrix [8-11] . In particular, the
proteolytic activities of MMP-2 and MMP-9 are implicated in tumor-associated processes such as cell
growth, survival, migration, invasion, and angiogenesis [8,12,13] . Moreover, by binding cell surface proteins, the
proforms of MMP-2 and MMP-9 can directly trigger intracellular signaling pathways involved in the
modulation of cell growth and survival, migration, or angiogenesis [8,14] .

Of particular interest, some natural products display antiangiogenic effects and represent potential interest
for the search of novel anti-cancer drugs [15-18] . Among these biologically active compounds, the
phloroglucinol hyperforin (HF) isolated from the plant St John’s wort (Hypericum perforatum, Figure 1)
acts as a multi-targeting agent [15,19-21] . HF displays anti-depressant, antibacterial, antioxidant, and anti-
inflammatory properties [15,19-22] . Moreover, HF exhibits anti-proliferative and proapoptotic activities towards
a number of mammalian cancer cell lines in vitro [15,23-28] . The antiangiogenic properties of HF have been
demonstrated in vitro and in vivo [29,30] . HF’s targets include MMP-2, MMP-9, and VEGF-A, all involved in
cell survival and migration, and angiogenesis [31-33] .

Acute myeloid leukemia (AML) is a heterogeneous hematopoietic cancer characterized by the accumulation
of malignant precursors of the myeloid lineage in the bone marrow (BM), interfering with the production of
[34]
normal blood cells . The latter are used to define distinct AML subfamilies . Human AML cells show
[34]
abnormally high levels of proliferation and survival, disseminating from bone marrow into peripheral blood
and extramedullary organs . The major cause of mortality of AML patients after allogeneic transplantation
[34]