Amplification of c-myc gene and overexpression of c-Myc protein in breast cancer and adjacent non-neoplastic tissue

doi:10.1016/S0009-9120(01)00260-0

Clinical Biochemistry

Volume 34, Issue 7, October 2001, Pages 557-562

https://doi.org/10.1016/S0009-9120(01)00260-0 Get rights and content

Abstract

Background: Deregulated c-Myc expression and alterations of c-myc oncogene have been reported to play an important role in breast cancer tumorigenesis. We examined the relationship between c-Myc protein level, amplification of c-myc oncogene and commonly used clinical and pathologic factors.

Methods: The studies were conducted on 94 ductal and lobular cancers. Amplification of c-Myc was assessed by the semiquantitative multiplex PCR assay. The amount of c-Myc protein was estimated by the densitometry analysis of Western blots.

Results: Amplification of c-Myc was found in 21% of examined cancers. There was no association of c-myc amplification with established risk factors. Overexpression of c-Myc protein without c-myc amplification was associated with negative status of axillary lymph node. The size of lobular carcinoma displaying overexpression of c-Myc and the normal copy number of c-myc gene was significantly smaller than the size of tumor with elevated c-Myc and amplification of c-myc gene (p < 0.01). Within tumors displaying overexpression of c-Myc protein and c-myc gene amplification the size of ductal carcinoma was smaller than the size of lobular carcinoma (p < 0.007).

Conclusion: Data presented in this study suggest that alterations of c-myc gene and c-Myc protein level might be related to breast cancer progression. The prognostic utility of elevated level of c-Myc protein associated with normal status of c-myc gene for patients with lobular carcinoma requires further studies.

Introduction

Breast cancer is one of the most common tumors in women, and the number of new cases increases continuously worldwide. The clinical course of this disease is highly variable. This includes cases with rapid progression and a short time of survival, but also patients with completely disease free interval of more than 10 yr. The reason for such variability is not fully understood and clinicians continuously search for prognostic parameters that can accurately predict prognosis, and indicate a suitable adjuvant therapy for each patient.

Accumulation of various genetic damage is a hallmark of human cancers. Many of these genetic alterations now have been identified. Over the past several years a vast number of experimental data on involvement of c-myc gene alterations in human breast cancer accumulated. c-Myc protein is a product of c-myc proto-oncogene and exerts diverse effects on cell behavior. Data gathered to date indicate that c-Myc protein plays a critical role in normal cell-cycle progression, inhibition of terminal differentiation and induction of programmed cell death [1], [2], [3]. On the other hand, deregulated c-Myc expression as a consequence of genetic changes has been observed in a variety of neoplasms [4]. Several reports indicate that genetic alterations of c-myc oncogene play an important role in induction and progression of human breast cancer [5], [6], [7], [8], [9]. Occurrence of c-myc oncogene amplification in breast cancer has been related to poor prognosis [10], [11], [12]. Overexpression of c-Myc protein to some extend correlates with gene amplification but not all tumors with c-myc amplification displayed increased level of this oncoprotein and some tumors displayed c-Myc overexpression without gene amplification [13], [14]. Despite over 15 yr continuous clinical work, the prognostic utility of c-myc gene alterations in breast cancer is still uncertain.

The level of c-myc expression in breast cancer was mostly investigated by measuring the level of mRNA [15], [16], [17]. Such a measurement does not necessary correlate with the protein level. The previous immunohistochemical examination of 206 breast cancers indicated that long recurrence-free survival of the patients was related to overexpression of c-Myc protein [18]. In search for ways to improve the characterization of breast cancer it would be important to examine the relationship between c-myc amplification and overexpression of c-Myc protein and widely used clinical and pathologic prognostic factors. In this report, we investigate amplification of c-myc and c-Myc protein level in primary breast cancer and adjacent nonneoplastic specimens. To gain insight into value of these parameters we analyzed the relation of overexpression and amplification of c-myc to several prognostic factors such as tumor size, estrogen and progesterone receptor status, mitotic index, local metastases and histologic grading.

Section snippets

Materials and methods

The studies were conducted on 94 ductal and lobular cancers selected from a pool consecutively accessioned cases. Patients with primary breast cancer had either an excision biopsy or a modified radical mastectomy. All specimens were measured and serially sectioned. All tissues were reviewed by a single pathologist (A. K.). Infiltrating cancers were considered ductal or lobular based upon their predominant cell type. Each specimen was dissected in a 5 mm bread-loaf fashion and tumor was measured

Results

The status of c-myc gene was studied on DNA isolated from 94 human breast cancers (lobular carcinoma and ductal carcinoma) and corresponding adjacent nonneoplastic tissues. To ruled out aneuploidy frequently occurring in developing tumors we used the gene for tissue plasminogen activator (TPA) located on chromosome 8 as a reference gene. Amplification of c-myc gene was considered to occur when the products of multiplex PCR displayed the intensity ratio (c-myc: TPA) equal or higher to that

Discussion

Data published to date indicate that oncogene amplification is one of the most common genetic alteration found in human cancers [22]. Amplification of c-myc has been demonstrated in 10 to 40% of breast cancers [6], [7], [8], [11], [12], [13]. The presence of c-myc amplification has also been shown to be a strong indicator of poor prognosis [7], [12]. We have studied amplification of c-myc gene in human breast cancers and adjacent nonneoplastic tissues by means of the semiquantitative multiplex

Acknowledgements

The authors wish to thank dr. R. Kowara for helpful discussion and dr. G. Kobierska for assistance in pathomorphological examination.

References (36)

A. Leder et al.
Consequences of widespread deregulation of the c-myc gene in transgenic micemultiple neoplasms, and normal development
Cell
(1986)
M. Roux-Dosseto et al.
c-myc gene amplification in selected node-negative breast cancer patients correlates with high rate of early relapse
Eur J Cancer
(1992)
O.H. Lowry et al.
Protein measurements with the folin phenol reagent
J Biol Chem
(1951)
R.A. Maas et al.
Growth arrest associated changes of mRNA levels in breast cancer cells measured by semi-quantitative RT-PCRpotential early indicators of treatment response
Cancer Lett
(1995)
P.W. Brandt-Rauf et al.
Molecular markers of carcinogenesis
Pharmacol Ther
(1998)
D.S. Askew et al.
Activation of apoptosis associated with enforced myc expression in myeloid progenitor cells is dominant to the suppression of apoptosis by interleukin-3 or erythropoietin
Blood
(1993)
C. Grandori et al.
Myc target genes
TIBS
(1997)
B.E. Thompson
The many roles of c-Myc in apoptosis
Annu Rev Physiol
(1998)
L.M. Facchini et al.
The molecular role of Myc in growth, and transformationrecent discoveries lead to new insights
FASEB J
(1998)
K.B. Marcu et al.
myc function, and regulation
Annu Rev Biochem
(1992)

P.H. Watson et al.

Relationship of c-myc amplification to progresion of breast cancer from in situ to invasive tumor, and lymph node metastasis

J Nat Cancer Inst

(1993)

P.H. Watson et al.

Influence of c-myc on the progression of human breast cancer

Curr Top Microbiol Immunol

(1996)

A. Sierra et al.

Synergistic cooperation between c-Myc, and Bcl-2 in lymph node progression of T1 human breast carcinomas

Breast Cancer Res Treat

(1999)

A. Borg et al.

c-myc amplification is an independent prognostic factor in postmenopausal breast cancer

Int J Cancer

(1992)

E.M. Berns et al.

c-myc amplification is a better prognostic factor than HER2/neu amplification in primary breast cancer

Cancer Res

(1992)

E.M. Berns et al.

TP53, and MYC gene alterations independently predict poor prognosis in breast cancer patients

Genes Chromosom Cancer

(1996)

L.P. Pertschuk et al.

Steroid hormone receptor immunohistochemistry, and amplification of c-myc protooncogene. Relationship to disease-free survival in breast cancer

Cancer

(1993)

C.S. Bruggers et al.

Expression of the c-Myc protein in childhood medulloblastoma

J Pediatr Hematol Oncol

(1998)

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Amplification of c-myc gene and overexpression of c-Myc protein in breast cancer and adjacent non-neoplastic tissue

Abstract

Introduction

Section snippets

Materials and methods

Results

Discussion

Acknowledgements

Cell

Eur J Cancer

J Biol Chem

Cancer Lett

Pharmacol Ther

Blood

Myc target genes

TIBS

The many roles of c-Myc in apoptosis

Annu Rev Physiol

The molecular role of Myc in growth, and transformationrecent discoveries lead to new insights

FASEB J

myc function, and regulation

Annu Rev Biochem