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In vivo evidence of htid suppressive activity on ErbB-2 in breast cancers over expressing the receptor.

Kurzik-Dumke U, Hörner M, Nicotra MR, Koslowski M, Natali PG - J Transl Med (2010)

Bottom Line: Altered ErbB-2 signalling is associated with an unfavourable prognosis in about 30% of human breast malignancies.A statistically significant inverse correlation between htid and ErbB-2 expression was found in human breast (p < 0,0001) and non-mammary tumors (p < 0,007), and in transgenic mice carrying the rat HER-2/neu oncogene.Our findings provide in vivo evidence that htid is a tissue independent and evolutionarily conserved suppressor of ErbB-2.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Medical Microbiology and Hygiene, Comparative Tumor Biology Group, University Medical Center, Johannes Gutenberg University, Obere Zahlbacher Str 63, 55131 Mainz, Germany. kurzik@uni-mainz.de

ABSTRACT

Background: Htid encoded proteins are physiological partners of a wide spectrum of molecules relevant to neoplastic transformation. One of the molecular ligands of the cytosolic hTid-L and hTid-I forms is the ErbB-2 receptor variably over expressed in diverse solid tumors. Altered ErbB-2 signalling is associated with an unfavourable prognosis in about 30% of human breast malignancies.

Methods: We evaluated htid and HER-2 expression by quantitative real time PCR in tumors of different TNMG status and by immunohistochemistry in a cohort of breast tumors of the Luminal A, B, HER-2 and triple negative subtype.

Results: The RT-PCR analysis revealed that aberrant expression of all three htid forms correlates with malignant transformation. Furthermore, elevated hTid-L expression can be associated with less aggressive tumors. The immunohistochemical testing revealed that tumors of the luminal A subtype are characterized by a high level of htid (81%). In contrast htid expression is significantly lower in tumors of the Luminal B (20%) and HER-2 (18%) subtype over expressing the receptor and in the triple negative (40%) more aggressive malignancies. A statistically significant inverse correlation between htid and ErbB-2 expression was found in human breast (p < 0,0001) and non-mammary tumors (p < 0,007), and in transgenic mice carrying the rat HER-2/neu oncogene.

Conclusions: Our findings provide in vivo evidence that htid is a tissue independent and evolutionarily conserved suppressor of ErbB-2.

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Photographs of simultaneous staining of 30 week old breast tumors arising in HER-2/neu transgenic mice illustrating that over expression of HER-2/neu correlates down regulation of htid. ErbB-2 was detected using a polyclonal chicken anti rat ErbB-2 and a FITC labelled secondary antibody. Tid was detected using the rabbit polyclonal anti hTid antibody [6,7,13] and a Texas Red labelled secondary antibody. The tumor cells over expressing ErbB-2 (A, arrowhead) are characterized by low tid expression (B, arrowhead; cf. Figure 3). In C an overlay of the photographs shown in A and B is presented.
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Figure 4: Photographs of simultaneous staining of 30 week old breast tumors arising in HER-2/neu transgenic mice illustrating that over expression of HER-2/neu correlates down regulation of htid. ErbB-2 was detected using a polyclonal chicken anti rat ErbB-2 and a FITC labelled secondary antibody. Tid was detected using the rabbit polyclonal anti hTid antibody [6,7,13] and a Texas Red labelled secondary antibody. The tumor cells over expressing ErbB-2 (A, arrowhead) are characterized by low tid expression (B, arrowhead; cf. Figure 3). In C an overlay of the photographs shown in A and B is presented.

Mentions: As described above, the inverse correlation of the expression profiles of the two proteins in human breast tumors and non mammary malignancies provides an in vivo proof for the oncosuppressive activity of htid on ErbB-2 described in vitro [15]. Furthermore, since the inverse relationship is highly significant in diverse tissues, this functional link is not cell specific, and therefore of general biological importance. Next, we asked whether the inverse correlation is evolutionarily conserved in the mouse. To answer this question we employed the transgenic mice model of Her-2/neu induced breast tumors [23]. We stained early (22 weeks) and late (30 weeks) breast tumors generated in transgenic mice carrying the rat Her-2/neu oncogene. As shown in Figure 3A, staining of the early tumors with the hTid [7,13] antiserum revealed homogenous (1+) cytoplasmic expression. Differently, using the anti-ErbB-2 antiserum cross reacting with the murine receptor resulted in a faint stain of the cytoplasm and rarely in staining of the membrane of the tumor cells (Figure 3B). In the more advanced 30 weeks tumors the staining patterns changed with regard to both the expression level and distribution. In these lesions, the hTid expression was heterogeneous with alternating areas of moderate to intense (2+) cytoplasmic stain (Figure 3C). The expression of ErbB-2 also appeared heterogenous with only discrete areas of the tumor displaying an intense (2+) staining often cell membrane associated (Figure 3D). This result further suggested that the inverse correlation of the expression levels of the two molecules is indeed detectable also in experimental tumors raised in mice carrying the rat Her-2/neu oncogene. In order to conclusively prove this issue, we submitted the more advanced tumors to double staining using anti-hTid [5,13] and the chicken antibody recognizing the murine TRK receptor molecule. As shown in Figure 4, tumor areas characterized by intense membrane ErbB-2 expression (A) display significantly lower htid level (B, C), thus, demonstrating the inverse correlation of the expression of the two tumor relevant molecules and implying their functional link.


In vivo evidence of htid suppressive activity on ErbB-2 in breast cancers over expressing the receptor.

Kurzik-Dumke U, Hörner M, Nicotra MR, Koslowski M, Natali PG - J Transl Med (2010)

Photographs of simultaneous staining of 30 week old breast tumors arising in HER-2/neu transgenic mice illustrating that over expression of HER-2/neu correlates down regulation of htid. ErbB-2 was detected using a polyclonal chicken anti rat ErbB-2 and a FITC labelled secondary antibody. Tid was detected using the rabbit polyclonal anti hTid antibody [6,7,13] and a Texas Red labelled secondary antibody. The tumor cells over expressing ErbB-2 (A, arrowhead) are characterized by low tid expression (B, arrowhead; cf. Figure 3). In C an overlay of the photographs shown in A and B is presented.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2909173&req=5

Figure 4: Photographs of simultaneous staining of 30 week old breast tumors arising in HER-2/neu transgenic mice illustrating that over expression of HER-2/neu correlates down regulation of htid. ErbB-2 was detected using a polyclonal chicken anti rat ErbB-2 and a FITC labelled secondary antibody. Tid was detected using the rabbit polyclonal anti hTid antibody [6,7,13] and a Texas Red labelled secondary antibody. The tumor cells over expressing ErbB-2 (A, arrowhead) are characterized by low tid expression (B, arrowhead; cf. Figure 3). In C an overlay of the photographs shown in A and B is presented.
Mentions: As described above, the inverse correlation of the expression profiles of the two proteins in human breast tumors and non mammary malignancies provides an in vivo proof for the oncosuppressive activity of htid on ErbB-2 described in vitro [15]. Furthermore, since the inverse relationship is highly significant in diverse tissues, this functional link is not cell specific, and therefore of general biological importance. Next, we asked whether the inverse correlation is evolutionarily conserved in the mouse. To answer this question we employed the transgenic mice model of Her-2/neu induced breast tumors [23]. We stained early (22 weeks) and late (30 weeks) breast tumors generated in transgenic mice carrying the rat Her-2/neu oncogene. As shown in Figure 3A, staining of the early tumors with the hTid [7,13] antiserum revealed homogenous (1+) cytoplasmic expression. Differently, using the anti-ErbB-2 antiserum cross reacting with the murine receptor resulted in a faint stain of the cytoplasm and rarely in staining of the membrane of the tumor cells (Figure 3B). In the more advanced 30 weeks tumors the staining patterns changed with regard to both the expression level and distribution. In these lesions, the hTid expression was heterogeneous with alternating areas of moderate to intense (2+) cytoplasmic stain (Figure 3C). The expression of ErbB-2 also appeared heterogenous with only discrete areas of the tumor displaying an intense (2+) staining often cell membrane associated (Figure 3D). This result further suggested that the inverse correlation of the expression levels of the two molecules is indeed detectable also in experimental tumors raised in mice carrying the rat Her-2/neu oncogene. In order to conclusively prove this issue, we submitted the more advanced tumors to double staining using anti-hTid [5,13] and the chicken antibody recognizing the murine TRK receptor molecule. As shown in Figure 4, tumor areas characterized by intense membrane ErbB-2 expression (A) display significantly lower htid level (B, C), thus, demonstrating the inverse correlation of the expression of the two tumor relevant molecules and implying their functional link.

Bottom Line: Altered ErbB-2 signalling is associated with an unfavourable prognosis in about 30% of human breast malignancies.A statistically significant inverse correlation between htid and ErbB-2 expression was found in human breast (p < 0,0001) and non-mammary tumors (p < 0,007), and in transgenic mice carrying the rat HER-2/neu oncogene.Our findings provide in vivo evidence that htid is a tissue independent and evolutionarily conserved suppressor of ErbB-2.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Medical Microbiology and Hygiene, Comparative Tumor Biology Group, University Medical Center, Johannes Gutenberg University, Obere Zahlbacher Str 63, 55131 Mainz, Germany. kurzik@uni-mainz.de

ABSTRACT

Background: Htid encoded proteins are physiological partners of a wide spectrum of molecules relevant to neoplastic transformation. One of the molecular ligands of the cytosolic hTid-L and hTid-I forms is the ErbB-2 receptor variably over expressed in diverse solid tumors. Altered ErbB-2 signalling is associated with an unfavourable prognosis in about 30% of human breast malignancies.

Methods: We evaluated htid and HER-2 expression by quantitative real time PCR in tumors of different TNMG status and by immunohistochemistry in a cohort of breast tumors of the Luminal A, B, HER-2 and triple negative subtype.

Results: The RT-PCR analysis revealed that aberrant expression of all three htid forms correlates with malignant transformation. Furthermore, elevated hTid-L expression can be associated with less aggressive tumors. The immunohistochemical testing revealed that tumors of the luminal A subtype are characterized by a high level of htid (81%). In contrast htid expression is significantly lower in tumors of the Luminal B (20%) and HER-2 (18%) subtype over expressing the receptor and in the triple negative (40%) more aggressive malignancies. A statistically significant inverse correlation between htid and ErbB-2 expression was found in human breast (p < 0,0001) and non-mammary tumors (p < 0,007), and in transgenic mice carrying the rat HER-2/neu oncogene.

Conclusions: Our findings provide in vivo evidence that htid is a tissue independent and evolutionarily conserved suppressor of ErbB-2.

Show MeSH
Related in: MedlinePlus