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The Emerging Role of Angiogenesis Inhibitors in Hematologic Malignancies

The Emerging Role of Angiogenesis Inhibitors in Hematologic Malignancies

ABSTRACT: Angiogenesis is an important component of the pathogenesis of hematologic malignancies. A negative prognostic implication of increased angiogenesis has been established for acute and chronic myeloid and lymphocytic leukemias, myeloproliferative diseases, multiple myeloma, non-Hodgkin’s lymphoma (NHL), and hairy cell leukemia. An association between the return of increased marrow vascularity to normal levels and durability of response has been established in some of these diseases. Elevated levels of proangiogenic factors have been associated with a poor prognosis in the acute and chronic leukemias, multiple myeloma, and NHL. These data lend support to the reduction of activity of proangiogenic factors as a therapeutic modality. Vascular endothelial growth factor (VEGF) has been implicated as the major proangiogenic factor that regulates multiple endothelial cell functions, including mitogenesis. A direct relationship between VEGF and leukemic blasts and malignant plasma cells has been established, but VEGF may have a function distinct from its role in angiogenesis. Current protocols with anti-VEGF agents in patients with hematologic malignancies involve the use of monoclonal antibody, blockers of the VEGF-receptor tyrosine kinase pathway, thalidomide (Thalomid) and its analogs, and cyclooxygenase inhibitors. The receptor tyrosine kinase inhibitors also affect platelet-derived growth factor, c-kit, and Flt-3 to varying degrees, considerably broadening their potential efficacy. This review will summarize several angiogenesis inhibitors in clinical development. [ONCOLOGY 16(Suppl 4):23-29, 2002]

Data on the significant role of
angiogenesis in the hematologic disorders—including acute myeloid leukemia
(AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), chronic
lymphocytic leukemia (CLL), the myelodysplastic syndromes (MDS), non-Hodgkin’s
lymphomas (NHL), hairy cell leukemia, multiple myeloma, and agnogenic myeloid
metaplasia—is growing in an exponential manner (Table
1
).[1-26] Bone marrow
microvessel density (MVD) is significantly greater in patients with advanced MDS
(ie, refractory anemia with excess blasts [RAEB] or RAEB in transformation
[RAEB-t]) compared with normal individuals. Patients with AML also have a
greater bone marrow MVD,[10] and successful induction chemotherapy has resulted
in a significant decrease in bone marrow MVD in these patients.[3] In children
with ALL, a complete remission (CR) associated with a return to normal bone
marrow MVD is more durable than a CR associated with a persistent increase in
MVD.[4] In patients with AML, ALL, and MDS, in addition to the relative increase
in MVD, there is a shift from the normal bone marrow predominance of straight,
nonbranching microvessels to vessels with a complex, arborizing architecture. In
patients with multiple myeloma, increased MVD has been directly correlated with
the plasma cell labeling index.[27]

The Proper Angiogenesis Modulator to Target

As summarized in Table 2, there are numerous molecules to target
within the angiogenesis cascade. However, one factor, vascular endothelial
growth factor (VEGF), has emerged as the prime target in treating hematologic
malignancies. Vascular endothelial growth factor is pivotal in the angiogenic
process and regulates several endothelial cell functions, including mitogenesis,
permeability, vascular tone, and the production of vasoactive molecules.[28]
Transcription of the VEGF gene (located on the short arm of chromosome 6 at
6p21.3) is physiologically regulated by hypoxia. Production of VEGF falls under
the control of alternate mRNA splicing and proteolytic processing. Alternate
splicing is responsible for the production of the isoforms VEGF 189, VEGF 165,
VEGF 121, and VEGF 205.[16]

The activity of VEGF is stimulated by three receptor tyrosine
kinases: VEGFR-1 (Flt-1), VEGFR-2 (KDR/Flk-1), and VEGFR-3 (Flt-4). Primarily
expressed on endothelial cells and monocytes, VEGFR-1 mediates cell
motility.[28] The proliferative and mitogenic activities of VEGF and increases
in vascular permeability are mediated primarily via VEGFR-2.[29] Finally,
VEGFR-3, homologous to the neurophilin-1-receptor, is involved in
lymphoangiogenesis.[30]

VEGF Activity in Hematologic Malignancies

Significantly elevated levels of VEGF are observed in a variety
of hematologic malignancies (Table 3). In a study of 417 patients, a number of
patients with AML (n = 115) and advanced MDS (n = 40) had similar elevated
plasma levels of VEGF.[31] In stored serum samples, increasing levels of
cellular VEGF had a positive correlation with a lower CR rate, shorter CR
duration, and shorter overall and disease-free survival times.[9] These data are
in contrast to those reported for the receptor tyrosine kinase Tie-1, in which
we used Western blot analysis to confirm and radioimmunoassay to quantify Tie-1
protein expression in bone marrow samples obtained from untreated patients with
AML (n = 66) or MDS (n = 29).[11] Tie-1 protein levels were elevated in all
disease specimens and were significantly higher in patients with AML than in
patients with MDS. However, Tie-1 levels did not correlate with CR or survival
duration in patients with either AML or MDS.

This contrast with respect to VEGF results is particularly
interesting when one considers the relative roles of Tie-1 and VEGF in marrow
angiogenesis. Tie-1 is expressed on both vascular endothelial cells and immature
hematopoietic cells, as is VEGF. Although Tie-1-deficient mice with a targeted
insertional mutation in their germ line have defects in endothelial cell
integrity, resulting in edema and hemorrhage, data indicate that Tie-1 is not
critical for either bone marrow stem cell engraftment or self-renewal.[32]
Interactions between proangiogenic molecules and marrow stromal elements may
also contribute to AML growth (Figure 1). For example, incubation of human
umbilical vein endothelial cells with VEGF results in increased secretion of
granulocyte-macrophage colony-stimulating factor (GM-CSF), an AML blast growth
factor, by the endothelium.[33]

We have recently evaluated the clinical significance of VEGFR-1
and VEGFR-2 protein levels in patients with untreated AML and MDS to identify
any relationship between these receptors and VEGF levels in each disease. Levels
of VEGFR-1 were significantly higher in AML than in MDS patients, and VEGFR-2
levels were equivalent. There was no correlation between VEGFR levels and
survival. Levels of VEGF were significantly higher in MDS than in AML patients,
and levels correlated with poorer survival in MDS patients. There was a
significant correlation between VEGF and VEGFR-2 levels in both AML and MDS
patients, but not between VEGF and VEGFR-1 levels in either disease. These data
suggest that VEGF expression, not VEGFR expression, is of prognostic relevance
in AML and MDS. The significance of the differential expression of VEGFR in AML
and MDS is unclear.

Chronic Lymphocytic Leukemia

Intracellular levels of VEGF have also been correlated with
prognosis in patients with CLL.[1] In an analysis of samples collected from
patients (n = 225) with B-cell CLL, the median intracellular VEGF level was 7.26
times higher than that of normal peripheral blood mononuclear cells. Patients
with lower levels of VEGF protein showed a trend toward a relatively reduced
rate of survival. In a subgroup of patients with Rai stages 0 to II, Binet stage
A and B disease, and good prognostic features (eg, beta-2-microglobulin of 2.8
mg/dL or less), lower levels of VEGF were also associated with shorter survival times.
However, no overall correlation among VEGF, disease stage, and beta-2-microglobulin
levels was demonstrated in the cohort.

In a separate report, there was an indirect correlation between
elevations in serum VEGF and the duration of progression-free survival (PFS) in
patients with Binet stage A disease.[15] Molica et al combined the patients’
Rai classification with serum VEGF levels, and identified two groups with
different PFS within stages I and II.[15] Patients with Rai stage I to II with
elevated VEGF levels had very short PFS (median, 9.5 months) compared with
patients with Rai stage I to II with low VEGF levels, who had longer PFS (median
not reached at 15 months).

In a recent examination of the role of VEGFR-2 in CLL, patients
with elevated VEGFR-2 levels had greater elevations in lymphocyte counts, severe
anemia, elevated serum beta-2-microglobulin levels, and advanced-stage
disease.[19] Elevated VEGFR-2 levels were associated with reduced survival.
Therefore, further studies on the interactive effects of VEGF and VEGFR-2 on CLL
proliferation are warranted. An analysis of Flt-1 and Tie-1 protein levels in
this same cohort of CLL patients also showed that these protein levels correlate
with white blood cell counts and absolute lymphocyte counts.[34] Flt-1 protein
levels correlated with levels of cellular VEGF, whereas levels of Tie-1 did not.
Furthermore, neither correlated with survival in the overall cohort; however,
higher levels of Tie-1, but not Flt-1, correlated with reduced survival.

Although high levels of both VEGF and basic fibroblast growth
factor (bFGF) have been documented in patients with CLL, bone marrow MVD is not
increased in most of these patients.[31] As observed in AML with VEGF levels,
aside from any proangiogenic role, VEGF and bFGF have significant, independent
roles in the pathophysiology of CLL.

Lymphomas

Elevated levels of VEGF are also associated with an adverse
prognosis in patients with NHL.[25] However, the highest prognostic power was
observed when VEGF and serum bFGF levels were examined in combination. The risk
of death in patients whose VEGF and bFGF levels were both within the highest
quartiles was greater than in other patients, independent of the prognostic
variables in the International Prognostic Index.[25] In collaboration with the
Omaha group, we have recently analyzed the clinical significance of serum levels
of VEGF, bFGF, hepatocyte growth factor (HGF), and angiogenin in untreated
patients with NHL or Hodgkin’s disease (HD). In patients with HD or NHL, VEGF
and HGF levels were significantly increased (personal communication, M. Albitar,
2002). In contrast, angiogenin levels were significantly decreased in patients
with either NHL or HD.

As reported by Salven et al, higher levels of VEGF and bFGF in
NHL patients correlated with more advanced disease stage and with higher lactate
dehydrogenase levels.[25] In addition, elevated levels of VEGF correlated with
shorter survival in patients with HD. Elevated baseline VEGF levels tended to
return to normal in patients with NHL who responded to therapy, and no
significant changes in the levels of other factors were observed. In posttherapy
samples collected from patients with HD, HGF and bFGF returned to normal levels,
whereas VEGF levels remained elevated. Patients with NHL with higher posttherapy
VEGF levels had relatively reduced survival and VEGF levels after therapy
remained predictive of survival in patients with HD. Angiogenic factors appear
to have distinct roles in the biology of HD and NHL, and a better appreciation
of these differences may aid in the diagnosis of some patients.

Multiple Myeloma

In patients with multiple myeloma, levels of VEGF, bFGF, and HGF
parallel disease activity, and VEGF levels correlate with features of aggressive
disease, including levels of serum C-reactive protein and beta-2-microglobulin
l.[7] Multiple myeloma patients have significantly higher levels of VEGF in bone
marrow than in peripheral blood—malignant plasma cells have been shown to
express and secrete VEGF. Although marrow MVD parallels disease activity in MM,
and it is thus reasonable to postulate that VEGF is acting in an autocrine
fashion, multiple myeloma cells have a low level of VEGFR expression. Thus, VEGF
may act in a paracrine fashion in multiple myeloma—both by its interactions
with interleukin-6, a known myeloma growth factor, and with bFGF. Levels of bFGF
have been reported to correlate with response rates to thalidomide (Thalomid)
treatment in multiple myeloma patients.[35] Because HGF is overproduced in
multiple myeloma and malignant cells express the HGF receptor c-met, this may be
the basis for another autocrine loop in multiple myeloma.[36]

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