15018752330
发表时间:2015-11-24 浏览次数:497次
Introduction
Expression or amplification of human epidermal growth-factor receptor 2 (HER2)
frequently occur in primitive neoplastic tissues from patients with breast
carcinoma (BC). However, in recent years, several studies have demonstrated
that HER2 status may vary in the metastatic lesions compared to the primary
tumor, and this discrepancy is more frequently found in distant metastases than
in loco-regional ones. Discordance in HER2 status was not only found between
primary BC and its metastases, but also among the consecutive relapses of the
same tumor, with similar proportions of cases turning from negative to positive
or vice versa and the changes mainly appeared in the second or following
progressions.
HER2 amplification may also be detected in gastric
carcinomas (GCs), with a prevalence ranging between 7.7% and 25% depending on
localization and histology of the cancer, a higher rate of HER2 amplification
occurs in unusual aggressive histology types, such as the hepatoid variant.
However, until date, there were only a few studies reporting HER2 heterogeneity
in paired primary and metastatic GC samples, and demonstrating a low rate of
discordance in HER2 amplification with either positive and negative conversion.
The potential divergence in the HER2 status between the primitive BC/GC
and their metastasized diseases, or among the successive metastases of the same
tumor, has a significant clinical relevance since it may modify the patient's
sensitivity to targeted therapies, which might be appropriate for the primitive
tumor, but not for the metastases or vice versa. For this reason, some
investigators proposed that detection of HER2 status should be re-assessed in
the neoplastic tissues from metastatic BC to establish whether the therapy is
actually appropriate.
Thus, in this study, we evaluated HER2 status in
paired samples of BC/GC and synchronous metastatic lymph nodes that were
collected during the same surgical and tissue processing procedures, thus
limiting and avoiding any potential technical bias due to external factors. Our
aim was to explore the eventual HER2 discordance rate between primary BC and GC
samples and corresponding lymph node metastases.
Methods
This cohort contained 127 surgical BC and GC specimens, together with the
corresponding regional synchronous metastatic lymph nodes. In brief, 65 primary
BC and 62 primary GC (male:female = 39:88; age ranged between 44 and 95 years
with mean age of 69.32 years) were retrospectively collected from the archive of
the Department of Human Pathology at the University of Messina. No patients had
received neo-adjuvant chemotherapy or other therapies before surgery.
The
primary GC was classified for localization and histology type according to WHO
2010, Lauren's classification and HER2 status of the tumor were available for
all cases. Similarly, histology, grade, hormone receptor status, Ki-67, and HER2
status were recorded for all BC cases. Patient identification was not disclosed
in this publication, and all patients had provided written consent to their
medical information being used for research purposes, according with the
Helsinki declaration.
For each case, 3 μm thick tissue sections from two
different formalin-fixed paraffin-embedded representative tissue blocks of the
primary tumor and metastatic lymph nodes (at least four for each case) were
prepared and immunohistochemical stained for HER2 expression. In brief, the
immunohistochemistry was carried out by using a DAKO HercepTest™ kit
(Dako, Glostrup, Denmark) with an automated procedure (DAKO Autostainer Link 48)
according to manufacturer's instructions. Antigen retrieval was performed by 3
cycles in 0.01 mol/L citrate buffer pH 6.0 in a microwave oven at 750 W. For
HER2 score was used to semiquantitatively assess HER2 expression level, that is,
for the primary GC, 0, absent staining; 1+, faint and discontinuous membranous
staining in < 10% of neoplastic elements; 2+, light to moderate lateral,
baso-lateral or complete membranous staining in > 10% of neoplastic elements;
3+, strong, intense lateral, baso-lateral or complete staining in > 10% of
neoplastic elements and for BC, 3+ score was defined when strong membranous
staining was noted in at least 30% cells, 2+ when weak to moderate complete
membranous staining was evidenced in 10-30% of tumors cells, 1+ when a faint or
weak and incomplete membrane staining was observed and 0 when no staining was
observed or when staining was present in < 10% of neoplastic
cells.
Furthermore, fluorescence in situ hybridization (FISH) was
performed using a HER2 FISH PharmDx™ kit (Dako) in those cases with
HER2 immunostaining score for 2+ or more. HER2 amplification was recorded when
HER2 to CEP17 signal ratio was > 2.0.
Fleiss-Cohen weighted K
statistics was used to assess the concordance rate between HER2 status of the
primary carcinomas and metastatic synchronous lesions. K values between 0 and
0.2 were regarded as no agreement, between 0.21 and 0.4 as fair agreement,
between 0.41 and 0.6 as moderate agreement, between 0.61 and 0.8 as substantial
agreement, and between 0.81 and 1 as almost perfect agreement. The statistical
association between HER2 status and the other histopathological parameters was
assessed using Chi-squared test. P < 0.05 was considered statistically
significant. All statistical analyses were performed using the SPSS package
version 6.1.3 (SPSS, Chicago, IL, USA).
Results
Thirty GC cases (48.40%) were localized in the lower third of the stomach, 22 (35.48%) in the middle third and 10 (16.12%) in the upper-third (four of which were localized at gastro-esophageal junction). Thirty-five GC cases (56.45%) were diagnosed histopathologically according to the WHO criteria as adenocarcinoma (tubular, papillary, tubulo-papillary, and mucinous), 20 cases (32.25%) as poorly cohesive carcinoma, and 7 cases (11.30%) were mixed both. According to Lauren's classification, 35 cases ( 56.45%) were classified as intestinal type, 20 cases (32.25%) as diffuse and 7 cases (11.30%) as mixed. Thirty-two of these 62 primary GC (51.61%) were recorded as low-grade tumors, while 30 cases were high grade (48.39%). HER2 immunohistochemical staining showed that 11 primary GCs (17.74%) were scored for 3+ HER2 expression, while 4 cases were 2+ (6.42%), 5 cases 1+ (8.10%), and 42 cases (67.74%) were not expressed HER2 at all. FISH analysis revealed no amplification in all of these cases with HER2 scores of 2+ or more. Taken together, in primary GC, HER2 was overexpressed in 11 cases (17.74%) but there was no HER2 amplification in 51 cases (82.26%). The overall concordance rate of HER2 status in primary GC between corresponding synchronous metastases was 90.32%, whereas a change in HER2 status was observed in 6 (9.68%) [Table 1], e.g. 4 cases with HER2 amplification in the primary GC but no amplification in the metastasized tumors [negative conversion; [Figure 1]a and b, two of these discordant cases did not show HER2 amplifications in the primitive tumor but amplified in the lymph node metastases [positive conversion; [Figure 1]c and d and [Table 2].
In the primary BC, the most frequent histology type was ductal invasive carcinomas with the following grading: 4 G1 (6.25%), 28 G2 (43%), and 33 G3 (50.75%). HER2 overexpression occurred in 14 (21.53%) of primary BC, 4 (6.15%) of which exhibited a score 2+, 2 (3.09%) a score 1+, while 45 (69.23%) cases didn't express HER2 at all. FISH analysis was conducted in those cases with the HER2 score of 2+ or more and the data revealed no HER2 amplification in these cases. Among 1+ cases, FISH was carried out in only two selected carcinomas showing high grade, high Ki-67 value, N+ status, and the absence of endocrine receptors expression, but no HER2 amplification was identified. HER2 was amplified in 14 BC cases (21.54%) but there was no HER2 amplification in these 51 cases (78.46%). The overall concordance rate was 95.39%, whereas changes in HER2 status between primary carcinoma and corresponding synchronous metastases were evidenced in 3 (4.61%) cases [Table 3]. Two of the discordant cases were HER2 negative in the primitive tumor but positive in the metastasized tumors [Figure 2]a and b, whereas one case was HER2 positive in the primary BC and turned to negative in the metastatic tumor [Figure 2]c and d and [Table 4].
After that, we performed statistical analyses and found that the K value for the concordance rate in the HER2 status between primitive tumors and metastases was 0.651 (substantial agreement). HER2 amplification was significantly more frequent in the intestinal-type GC than that of diffuse-type while no significant differences in HER2 expression were noted among BC histology types. No statistical significant correlation emerged between HER2 and clinicopathological parameters (hormone receptors, growth fraction, pT, pN, and grade) either in GC as well as BC.
Discussion
In the current study, we retrospectively analyzed HER2 expression in surgical GC
and BC specimens versus the corresponding metastatic lymph nodes. Our results
firstly confirmed the presence of a high level of concordance in HER2 status
between the primary GC/BC and their corresponding lymph node metastases (90.32%
and 95.39% respectively), which is consistent with previous observations of
metachronous metastases (87.5-94.9%). Moreover, we also found evidence of HER2
differences between primary carcinomas and their nodal metastases, that is,
9.68% GC cases and 4.61% BC cases did have the discordance between the primary
and secondary tumors. Specifically, four cases had HER2 amplifications in the
primary GC but there were no HER2 amplifications in the metastatic tumors. In
contrast, two of the gastric discordant cases showed no HER2 amplifications in
the primitive tumor but amplified in the lymph node metastatic tumors.
Similarly, there were two of the discordant BC cases showed negative HER2 in the
primitive tumor but became positive in the metastatic tumors, whereas one case
was from positive HER2 in the primary BC to negative in the metastases.
Therefore, a positive or negative conversion was encountered in either GC or BC
cases, although with a different discordance rate. A possible explanation for
the discordance observed in GC than in BC cases could be attributed to the most
frequent occurrence of a heterogeneity in GC cases, compared to BC. Hence, the
biopsies or tissue microarray assays do not seem adequate for assessment of HER2
expression, in contrast to that elsewhere reported. In addition, the
multisampling method performed in this study using at least two tissue blocks of
primary tumors and four of metastatic lymph nodes could identify more discordant
cases and compensate a potential heterogeneous HER2 expression. The possible
explanation of HER2 positive conversion may be related to the selection of a new
HER2 positive clone in metastatic lymph nodes as a result of disease
progression. Loss of HER2 amplification (negative conversion) in metastatic
tumors could not be only attributed to the development of resistance to
trastuzumab therapy since our patients had not been subjected to any
neo-adjuvant treatment.
Changes in HER2 status between primary GC/BC and
synchronous lymph node metastases may have relevant clinical impact. For
example, only HER2 positive GC and BC currently support the use of trastuzumab
in these patients; thus, our present finding suggests a need to reassess HER2
status before trastuzumab treatment. As a matter of fact, assessment of HER2
expression in the primary GC and BC may exclude from the targeted treatment a
significant percentage of patients with a negative primary tumor, but positive
metastases. Finally, the influence of discordant HER2 status in the therapeutic
management as well as in the prognostic impact of patients affected by GC and BC
should be greatly considered in order to correctly identify possible eligible
candidates for trastuzumab-based therapy, even among patients with HER2 negative
primary carcinomas.
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