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Identification of potential therapeutic targets in human head & neck squamous cell carcinoma.

Han J, Kioi M, Chu WS, Kasperbauer JL, Strome SE, Puri RK - Head Neck Oncol (2009)

Bottom Line: QRT-PCR analysis was performed to validate the microarray results.The five candidate genes were further characterized by immunohistochemical technique in surgical samples and tissue arrays.These genes belong to immune response, cell growth, cell cycle regulation, oncogenes, metabolism and others.

View Article: PubMed Central - HTML - PubMed

Affiliation: Tumor Vaccines and Biotechnology Branch, Division of Cellular and Gene Therapies, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD 20892, USA. jing.han@fda.hhs.gov

ABSTRACT

Background: Human head and neck squamous cell carcinoma (HNSCC) is an aggressive and recurrent malignancy. Identification of unique or overexpressed cell-associated or cell surface antigens is critical for diagnosis and development of cancer vaccines and targeted therapies for HNSCC. We have used high throughput microarray technology to search for candidate targets in HNSCC.

Methods: Gene expression profiling in 17 HNSCC tumors and 3 normal tonsil tissues was performed by microarray. QRT-PCR analysis was performed to validate the microarray results. The five candidate genes were further characterized by immunohistochemical technique in surgical samples and tissue arrays.

Results: A total of 192 up-regulated genes at statistical significance of p < 0.01 and log2 ratio > or = 1 were identified in HNSCC tumors compared to normal tissues. These genes belong to immune response, cell growth, cell cycle regulation, oncogenes, metabolism and others. Five potential novel target genes (FABP5, CD24, CD44, CD74, and HSP27) were identified, which were highly expressed in HNSCC tumor samples and tissue arrays. CD24, CD44, and CD74 proteins were expressed on the cell surface, and FABP5 and HSP27 proteins were predominantly expressed in the cytoplasm of HNSCC.

Conclusion: Five genes and their products may serve as a diagnostic biomarker or therapeutic target for HNSCC. While additional work is needed to elucidate the biological significance of these proteins, CD24 and CD74 expressed only in small proportion of cells indicating tumor heterogeneity and subtypes of tumor initiating cells (CD24+/CD44+) present in HNSCC.

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Immunohistochemical analysis of CD24, CD44, CD74, FABP5, and Hsp27 in HNSCC samples. HNSCC tissue arrays contained 16 tumor samples from various locations and different stages. A and B are tumor samples and stained for CD24. D and E stained for CD44, and showed strong positive reactions in most tumor cells. Both CD24+ and CD44+ show cell surface staining, and CD24+ cells present in a small cluster of cells in the large tumor mass. (Magnification: ×200). G and H stained for CD74. Some of tumor cells showed CD74+. J and K stained for FABP5, and M and N stained for Hsp27. FABP5 shows strong cytoplasmic staining as well as Hsp27. C, F, I, L, O: tumor adjacent normal tissue. CD24, CD44, CD74, FABP5, and Hsp27 were negative in normal tissues, although FABP5 shows positive staining in the basal layer of dermis (Fig 2L), but negative in other areas; (Magnification: ×200). IgG was used as a negative control (Data not shown).
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Figure 2: Immunohistochemical analysis of CD24, CD44, CD74, FABP5, and Hsp27 in HNSCC samples. HNSCC tissue arrays contained 16 tumor samples from various locations and different stages. A and B are tumor samples and stained for CD24. D and E stained for CD44, and showed strong positive reactions in most tumor cells. Both CD24+ and CD44+ show cell surface staining, and CD24+ cells present in a small cluster of cells in the large tumor mass. (Magnification: ×200). G and H stained for CD74. Some of tumor cells showed CD74+. J and K stained for FABP5, and M and N stained for Hsp27. FABP5 shows strong cytoplasmic staining as well as Hsp27. C, F, I, L, O: tumor adjacent normal tissue. CD24, CD44, CD74, FABP5, and Hsp27 were negative in normal tissues, although FABP5 shows positive staining in the basal layer of dermis (Fig 2L), but negative in other areas; (Magnification: ×200). IgG was used as a negative control (Data not shown).

Mentions: To narrow down the range of potential therapeutic targets in 192 up-regulated genes identified in our HNSCC samples, we mainly focused on the genes encoding cell surface receptors and some cytoplasmic proteins. Among these genes, we selected five gene products (CD24, CD44, CD74, FABP5, and HSP27) as antibodies to these products were available for paraffin embedded tissue sections. The expression of selected gene products was confirmed by IHC in tissue sections and tissue arrays. Since CD24, CD44, and CD74 are cell surface proteins and function as adhesion molecules, their expression was predominantly on the cell surface. Nine out of 16 HNSCC tumors in tissue arrays were positive for CD24 expression. However, its expression was found only in small clusters of cells within tumors (Fig 2A and 2B). CD44 was strongly expressed on the cell surface of all tumor cells in all cases of HNSCC samples (Fig 2D and 2E). CD74 positive cells were also identified in HNSCC tissue arrays (Fig 2G and 2H). In contrast, CD24, CD44, or CD74 positive cells were not detected in adjacent normal tissues present in tissue arrays (Fig 2C, F, and 2I).


Identification of potential therapeutic targets in human head & neck squamous cell carcinoma.

Han J, Kioi M, Chu WS, Kasperbauer JL, Strome SE, Puri RK - Head Neck Oncol (2009)

Immunohistochemical analysis of CD24, CD44, CD74, FABP5, and Hsp27 in HNSCC samples. HNSCC tissue arrays contained 16 tumor samples from various locations and different stages. A and B are tumor samples and stained for CD24. D and E stained for CD44, and showed strong positive reactions in most tumor cells. Both CD24+ and CD44+ show cell surface staining, and CD24+ cells present in a small cluster of cells in the large tumor mass. (Magnification: ×200). G and H stained for CD74. Some of tumor cells showed CD74+. J and K stained for FABP5, and M and N stained for Hsp27. FABP5 shows strong cytoplasmic staining as well as Hsp27. C, F, I, L, O: tumor adjacent normal tissue. CD24, CD44, CD74, FABP5, and Hsp27 were negative in normal tissues, although FABP5 shows positive staining in the basal layer of dermis (Fig 2L), but negative in other areas; (Magnification: ×200). IgG was used as a negative control (Data not shown).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2719634&req=5

Figure 2: Immunohistochemical analysis of CD24, CD44, CD74, FABP5, and Hsp27 in HNSCC samples. HNSCC tissue arrays contained 16 tumor samples from various locations and different stages. A and B are tumor samples and stained for CD24. D and E stained for CD44, and showed strong positive reactions in most tumor cells. Both CD24+ and CD44+ show cell surface staining, and CD24+ cells present in a small cluster of cells in the large tumor mass. (Magnification: ×200). G and H stained for CD74. Some of tumor cells showed CD74+. J and K stained for FABP5, and M and N stained for Hsp27. FABP5 shows strong cytoplasmic staining as well as Hsp27. C, F, I, L, O: tumor adjacent normal tissue. CD24, CD44, CD74, FABP5, and Hsp27 were negative in normal tissues, although FABP5 shows positive staining in the basal layer of dermis (Fig 2L), but negative in other areas; (Magnification: ×200). IgG was used as a negative control (Data not shown).
Mentions: To narrow down the range of potential therapeutic targets in 192 up-regulated genes identified in our HNSCC samples, we mainly focused on the genes encoding cell surface receptors and some cytoplasmic proteins. Among these genes, we selected five gene products (CD24, CD44, CD74, FABP5, and HSP27) as antibodies to these products were available for paraffin embedded tissue sections. The expression of selected gene products was confirmed by IHC in tissue sections and tissue arrays. Since CD24, CD44, and CD74 are cell surface proteins and function as adhesion molecules, their expression was predominantly on the cell surface. Nine out of 16 HNSCC tumors in tissue arrays were positive for CD24 expression. However, its expression was found only in small clusters of cells within tumors (Fig 2A and 2B). CD44 was strongly expressed on the cell surface of all tumor cells in all cases of HNSCC samples (Fig 2D and 2E). CD74 positive cells were also identified in HNSCC tissue arrays (Fig 2G and 2H). In contrast, CD24, CD44, or CD74 positive cells were not detected in adjacent normal tissues present in tissue arrays (Fig 2C, F, and 2I).

Bottom Line: QRT-PCR analysis was performed to validate the microarray results.The five candidate genes were further characterized by immunohistochemical technique in surgical samples and tissue arrays.These genes belong to immune response, cell growth, cell cycle regulation, oncogenes, metabolism and others.

View Article: PubMed Central - HTML - PubMed

Affiliation: Tumor Vaccines and Biotechnology Branch, Division of Cellular and Gene Therapies, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD 20892, USA. jing.han@fda.hhs.gov

ABSTRACT

Background: Human head and neck squamous cell carcinoma (HNSCC) is an aggressive and recurrent malignancy. Identification of unique or overexpressed cell-associated or cell surface antigens is critical for diagnosis and development of cancer vaccines and targeted therapies for HNSCC. We have used high throughput microarray technology to search for candidate targets in HNSCC.

Methods: Gene expression profiling in 17 HNSCC tumors and 3 normal tonsil tissues was performed by microarray. QRT-PCR analysis was performed to validate the microarray results. The five candidate genes were further characterized by immunohistochemical technique in surgical samples and tissue arrays.

Results: A total of 192 up-regulated genes at statistical significance of p < 0.01 and log2 ratio > or = 1 were identified in HNSCC tumors compared to normal tissues. These genes belong to immune response, cell growth, cell cycle regulation, oncogenes, metabolism and others. Five potential novel target genes (FABP5, CD24, CD44, CD74, and HSP27) were identified, which were highly expressed in HNSCC tumor samples and tissue arrays. CD24, CD44, and CD74 proteins were expressed on the cell surface, and FABP5 and HSP27 proteins were predominantly expressed in the cytoplasm of HNSCC.

Conclusion: Five genes and their products may serve as a diagnostic biomarker or therapeutic target for HNSCC. While additional work is needed to elucidate the biological significance of these proteins, CD24 and CD74 expressed only in small proportion of cells indicating tumor heterogeneity and subtypes of tumor initiating cells (CD24+/CD44+) present in HNSCC.

Show MeSH
Related in: MedlinePlus