Sabato 27 ottobre 2012 - Pacini Editore

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toplasmic HSCORE 1 for Notch 1 and 3 was evident respectively
in 4 and 5 out of 12 needle biopsies with atrophy. Notch expression
was not evident in the 8 cases with basal cell hyperplasia.
Cytoplasmic HSCORE 1 for Notch-1 and Notch-3 were found in 6
out of 11 high-grade PIN cases adjacent to neoplastic glands, more
intense at the apical pole of the glandular cells. On the contrary,
no expression of Notch-2 was appreciated in negative biopsies and
in high grade PIN. Muscle cells intermingled with non-neoplastic
glands and high-grade PIN, and vascular endothelial cells showed
some degree of Notch 1-3 staining. Expression of Notch receptors
in PC: neoplastic glands showed different HSCORE values and
different immunoreactivity location accordingly to the Gleason
grade and to the antibody evaluated. Notch 1 was expressed in
80% of Gleason grade 3, with 60% showing a HSCORE 1. Only
2 cases Gleason grade 4 (9%) showed Notch 1 positivity, with a
HSCORE 1. None of the Gleason grade 5 was Notch 1 positive.
Only 2 out of 23 high-grade tumors expressed Notch-1, with a
HSCORE 1. Notch 2 expression was observed in 20% of Gleason
grade 3 and in 4.5% of Gleason grade 4, with a HSCORE 1. None
of the Gleason grade 5 was Notch 2 positive. In any case, Notch
1 and Notch 2 staining was confined to the cytoplasm. Higher
Notch-3 expression (HSCORE 3) was observed more frequently
in Gleason high grade cases (121 out of 127 cases) in comparison
with Gleason low grade (20 out of 88 cases) (p < 0.001). Higher
Notch-3 immunostaining (HSCORE 3) occurred more frequently
in tumors with a higher Gleason score (n = 53) in comparison with
low-grade tumors (n = 39) (p < 0.001). Gleason high grade cases
often showed nuclear expression (n = 115) than Gleason low grade
cases (n = 6) (p < 0.001). These data suggest progression of Notch
3 receptor expression and activation with tumor malignancy.
Discussion. It is well-known that androgen sensitivity is a major
determinant of PC outcome. However, there is still insufficient
information as to which signaling pathways are deranged in
more aggressive, clinically localized PC. Identification of these
pathways may result in the design of innovative therapies aimed
at specific molecular targets. Recently, the gene expression profile
of Gleason low grade and high grade tumors was compared
Among the genes that were found to be differentially expressed,
members of the Notch and EGFR signal transduction pathways
appeared to be up-regulated in high grade localized PC. In this
study, we found that the level of Notch 3 receptor protein was
significantly correlated with Gleason grade in human PC biopsies
and that Notch 3 staining was mainly nuclear in high grade
tumors, suggesting receptor activation. We also observed that
Notch 3 was strongly activated in LNCaP cells during hypoxia.
Preliminary data from our group suggest that activation of Notch
3 under hypoxia increases the ability of PC cells to proliferate
and to form colonies in soft agar. On these bases, we propose that
Notch 3 immunostaining of localized PC biopsies may increase
our ability to identify those tumors with worse prognosis that may
require a more aggressive therapy. Moreover, Notch 3 signaling
pathway may be a candidate for an innovative targeted therapy.
references
Andersson ER, Sandberg R, Lendahl U. Notch signaling: simplicity in
design, versatility in function. Development 2011;138:3593-612.
Danza G, Di Serio C, Rosati F, et al. Notch signaling modulates hypoxia-induced
neuroendocrine differentiation of human prostate cancer
cells. Mol Cancer Res 2012 ;10:230-8.
Epstein JI, Netto GJ. Grading of prostatic adenocarcinoma. In: Pine J,
McGough, editors. Biopsy Interpretation of the Prostate. Philadelphia:
Lippincott Williams and Wilkins 2007, pp. 175-202.
Ghafar MA, Anastasiadis AG, Chen MW, et al. Acute hypoxia increases
the aggressive characteristics and survival properties of prostate cancer
cells. Prostate 2003;54:58-67.
Long Q, Johnson BA, Osunkoya AO, et al. Protein-Coding and MicroR-
NA Biomarkers of Recurrence of Prostate Cancer Following Radical
Prostatectomy. Am J Pathol 2011;179:46-54.
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CONGRESSO aNNualE di aNatOmia patOlOGiCa SiapEC – iap • fiRENzE, 25-27 OttOBRE 2012
GDP-4-keto-6-deoxy-mannose-3,5-epimerase-4reductase
as a novel diagnostic and prognostic
marker in prostate carcinoma
M.R. Ambrosio, P. Guazzi, B.J. Rocca, S. Bartolommei, A. Sacchini,
M.T. del Vecchio, S.A. Tripodi, F. Arcuri
Department of Human Pathology and Oncology, Pathological Anatomy
Section, University of Siena, Italy
Background. It is well known that the structure of carbohydrates
produced by cells changes during carcinogenesis; in this frame,
alterations in fucosylation patterns, as the result of different levels
of expression for various fucosyltransferase have been studied.
Overexpression of fucosylation enzymes and high production of
GDP-fucose is common in cancer, promoting metastatic potential.
The regulation of fucosylation is a complex phenomenon, involving
several systems that control fucosyltransferases or substrate
availabilities. The level of fucosyltransferases and intracellular
GDP-L-fucose, a sugar nucleotide and a common donor substrate
for all fucosyltransferases, may regulate that of fucosylated glycoproteins.
In fucosylation process, L-fucose (6-deoxy-L-galactose)
monomers are transferred to target GDP-L-fucose in the cytosol
via two different pathway, named the salvage pathway and the
de novo pathway. In the de novo pathway, GDP-L-fucose is synthesized
from GDP-mannose via an oxidation, an epimerization,
and a reduction. These steps are catalyzed by two enzymes, GDPmannose-4,6-dehydratase
and GDP-4-keto-6-deoxy-mannose-3,5epimerase-4-reductase.
In humans, the latter enzyme is designated
as FX. It has been reported that FX-knockout mice exhibit a complete
deficiency of cellular fucosylation, demonstrating that the
de novo pathway is the major route for cellular GDP-L-fucose
biosynthesis in vivo. Recent studies on prostatic carcinoma cell
lines (LNCaP) have shown that PSA derived from LNCaP cells
contain more fucosylated and N-acetylgalactosaminylated residues
than those from normal and benign prostatic hyperplasia (BPH)
tissues, suggesting a role of cellular fucosylation in prostatic adenocarcinoma.
The aim of this study is to elucidate whether FX is
involved in prostatic carcinogenesis, and to evaluate its potential
role as diagnostic and prognostic marker.
Materials and methods. Western blotting and Real Timequantitative
PCR analysis were carried out on BPH and prostatic
carcinoma (PCa) specimens obtained from patients undergoing
adenomectomy and radical prostatectomy at the Urology Division
of Siena University Hospital. Immediately after surgery,
specimens were rinsed in sterile PBS at room temperature, blotted
dry, cut with a razor blade. Aliquots for western blotting analysis
and RNA extraction were snap-frozen and stored in liquid nitrogen.
For immunohistochemical evaluation, the material consisted
of 120 core needle biopsy specimen from 70 prostate cancer
patients, and from 50 cancer-free patients, collected between
January and June 2011. The mean age of the patients was 70.4
(range: 54 to 86 years). None of the patients received adjuvant
therapy. Core needle biopsy specimens were immediately fixed
in 10% buffered formalin and embedded in paraffin. From each
block, 4 µm-thick sections were cut and stained with hematoxylin
and eosin. Tumor grade was established according to the Gleason
grading system and the tumors classified as low grade when the
score was < 7 and as high grade when the score was > 8. Foci of
high-grade intraepithelial neoplasia (PIN) were also identified.
Cancer-free core needle biopsy specimens may contain basal cell
hyperplasia, atrophy, acute or chronic inflammation. Immunohistochemical
staining was performed on 4 + 0.5µm-thick sections
of each block employing the Ultravision Detection System
Anti-Polyvalent HRP and semiquantitatively scored by HSCORE
(HSCORE = ∑ i i*P i), using the following categories: 1 (no staining
or weak, but detectable staining), 2 (moderate staining) and
3 (intense staining). Statistical analysis was performed using a
statistical software package (SYSTAT-7). A P value < 0.05 was
considered statistically significant.