Development and validation of a novel clinical fluorescence in situ hybridization assay to detect JAK2 and PD-L1 amplification

A fluorescence in situ hybridization assay for JAK2 and PD-L1 amplification

Meixuan Chen, Mariacarla Andreozzi, Barbara Pockaj, Michael T. Barrett, Idris Tolgay Ocal, Ann E. McCullough, Maria E. Linnaus, James M. Chang, Jennifer H. Yearley, Lakshmanan Annamalai, Karen Anderson

Research output: Contribution to journalArticle

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Abstract

The amplification of chromosome 9p24.1 encoding PD-L1, PD-L2, and JAK2 has been reported in multiple types of cancer and is associated with poor outcome, upregulation of PD-L1, and activation of the JAK/STAT pathway. We have developed a novel fluorescence in situ hybridization assay which combines 3 probes mapping to 9p24.1 with a commercial chromosome 9 centromere (CEN9) probe for detection of the JAK2/9p24.1 amplification. JAK2 fluorescence in situ hybridization was compared with array-based comparative genomic hybridization in 34 samples of triple negative breast cancer tumor. By array-based comparative genomic hybridization, 15 had 9p24.1 copy-number gain (log2rati<40.3) and 19 were classified as non-gain (log2ratio≤0.3). Copy-number gain was defined as JAK2/CEN9 ratio ≥1.1 or average JAK2 signals≥3.0. Twelve of 15 samples with copy-number gain by array-based comparative genomic hybridization were also detected by fluorescence in situ hybridization. Eighteen of 19 samples classified as copy-number non-gain by array-based comparative genomic hybridization were concordant by array-based comparative genomic hybridization. The sensitivity and specificity of the fluorescence in situ hybridization assay was 80% and 95%, respectively (P=0.02). The sample with the highest level of amplification by array-based comparative genomic hybridization (log2ratio=3.6) also scored highest by fluorescence in situ hybridization (ratio=8.2). There was a correlation between the expression of JAK2 and amplification status (Mean 633 vs 393, P=0.02), and there was a trend of association with PD-L1 RNA expression (Mean 46 vs 22, P=0.11). No significant association was observed between PD-L1 immunohistochemistry expression and copy-number gain status. In summary, the novel array-based comparative genomic hybridization assay for detection of chromosome 9p24.1 strongly correlates with the detection of copy-number gain by array-based comparative genomic hybridization. In triple negative breast cancer, this biomarker may identify a relevant subset of patients for targeted molecular therapies.

Original languageEnglish (US)
Pages (from-to)1516-1526
Number of pages11
JournalModern Pathology
Volume30
Issue number11
DOIs
StatePublished - Nov 1 2017

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Comparative Genomic Hybridization
Fluorescence In Situ Hybridization
Triple Negative Breast Neoplasms
Chromosomes
Molecular Targeted Therapy
Chromosomes, Human, Pair 9
Centromere
Tumor Biomarkers
Up-Regulation
Immunohistochemistry
RNA
Breast Neoplasms
Sensitivity and Specificity

ASJC Scopus subject areas

  • Pathology and Forensic Medicine

Cite this

Development and validation of a novel clinical fluorescence in situ hybridization assay to detect JAK2 and PD-L1 amplification : A fluorescence in situ hybridization assay for JAK2 and PD-L1 amplification. / Chen, Meixuan; Andreozzi, Mariacarla; Pockaj, Barbara; Barrett, Michael T.; Ocal, Idris Tolgay; McCullough, Ann E.; Linnaus, Maria E.; Chang, James M.; Yearley, Jennifer H.; Annamalai, Lakshmanan; Anderson, Karen.

In: Modern Pathology, Vol. 30, No. 11, 01.11.2017, p. 1516-1526.

Research output: Contribution to journalArticle

Chen, Meixuan ; Andreozzi, Mariacarla ; Pockaj, Barbara ; Barrett, Michael T. ; Ocal, Idris Tolgay ; McCullough, Ann E. ; Linnaus, Maria E. ; Chang, James M. ; Yearley, Jennifer H. ; Annamalai, Lakshmanan ; Anderson, Karen. / Development and validation of a novel clinical fluorescence in situ hybridization assay to detect JAK2 and PD-L1 amplification : A fluorescence in situ hybridization assay for JAK2 and PD-L1 amplification. In: Modern Pathology. 2017 ; Vol. 30, No. 11. pp. 1516-1526.
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abstract = "The amplification of chromosome 9p24.1 encoding PD-L1, PD-L2, and JAK2 has been reported in multiple types of cancer and is associated with poor outcome, upregulation of PD-L1, and activation of the JAK/STAT pathway. We have developed a novel fluorescence in situ hybridization assay which combines 3 probes mapping to 9p24.1 with a commercial chromosome 9 centromere (CEN9) probe for detection of the JAK2/9p24.1 amplification. JAK2 fluorescence in situ hybridization was compared with array-based comparative genomic hybridization in 34 samples of triple negative breast cancer tumor. By array-based comparative genomic hybridization, 15 had 9p24.1 copy-number gain (log2rati<40.3) and 19 were classified as non-gain (log2ratio≤0.3). Copy-number gain was defined as JAK2/CEN9 ratio ≥1.1 or average JAK2 signals≥3.0. Twelve of 15 samples with copy-number gain by array-based comparative genomic hybridization were also detected by fluorescence in situ hybridization. Eighteen of 19 samples classified as copy-number non-gain by array-based comparative genomic hybridization were concordant by array-based comparative genomic hybridization. The sensitivity and specificity of the fluorescence in situ hybridization assay was 80{\%} and 95{\%}, respectively (P=0.02). The sample with the highest level of amplification by array-based comparative genomic hybridization (log2ratio=3.6) also scored highest by fluorescence in situ hybridization (ratio=8.2). There was a correlation between the expression of JAK2 and amplification status (Mean 633 vs 393, P=0.02), and there was a trend of association with PD-L1 RNA expression (Mean 46 vs 22, P=0.11). No significant association was observed between PD-L1 immunohistochemistry expression and copy-number gain status. In summary, the novel array-based comparative genomic hybridization assay for detection of chromosome 9p24.1 strongly correlates with the detection of copy-number gain by array-based comparative genomic hybridization. In triple negative breast cancer, this biomarker may identify a relevant subset of patients for targeted molecular therapies.",
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AU - Andreozzi, Mariacarla

AU - Pockaj, Barbara

AU - Barrett, Michael T.

AU - Ocal, Idris Tolgay

AU - McCullough, Ann E.

AU - Linnaus, Maria E.

AU - Chang, James M.

AU - Yearley, Jennifer H.

AU - Annamalai, Lakshmanan

AU - Anderson, Karen

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N2 - The amplification of chromosome 9p24.1 encoding PD-L1, PD-L2, and JAK2 has been reported in multiple types of cancer and is associated with poor outcome, upregulation of PD-L1, and activation of the JAK/STAT pathway. We have developed a novel fluorescence in situ hybridization assay which combines 3 probes mapping to 9p24.1 with a commercial chromosome 9 centromere (CEN9) probe for detection of the JAK2/9p24.1 amplification. JAK2 fluorescence in situ hybridization was compared with array-based comparative genomic hybridization in 34 samples of triple negative breast cancer tumor. By array-based comparative genomic hybridization, 15 had 9p24.1 copy-number gain (log2rati<40.3) and 19 were classified as non-gain (log2ratio≤0.3). Copy-number gain was defined as JAK2/CEN9 ratio ≥1.1 or average JAK2 signals≥3.0. Twelve of 15 samples with copy-number gain by array-based comparative genomic hybridization were also detected by fluorescence in situ hybridization. Eighteen of 19 samples classified as copy-number non-gain by array-based comparative genomic hybridization were concordant by array-based comparative genomic hybridization. The sensitivity and specificity of the fluorescence in situ hybridization assay was 80% and 95%, respectively (P=0.02). The sample with the highest level of amplification by array-based comparative genomic hybridization (log2ratio=3.6) also scored highest by fluorescence in situ hybridization (ratio=8.2). There was a correlation between the expression of JAK2 and amplification status (Mean 633 vs 393, P=0.02), and there was a trend of association with PD-L1 RNA expression (Mean 46 vs 22, P=0.11). No significant association was observed between PD-L1 immunohistochemistry expression and copy-number gain status. In summary, the novel array-based comparative genomic hybridization assay for detection of chromosome 9p24.1 strongly correlates with the detection of copy-number gain by array-based comparative genomic hybridization. In triple negative breast cancer, this biomarker may identify a relevant subset of patients for targeted molecular therapies.

AB - The amplification of chromosome 9p24.1 encoding PD-L1, PD-L2, and JAK2 has been reported in multiple types of cancer and is associated with poor outcome, upregulation of PD-L1, and activation of the JAK/STAT pathway. We have developed a novel fluorescence in situ hybridization assay which combines 3 probes mapping to 9p24.1 with a commercial chromosome 9 centromere (CEN9) probe for detection of the JAK2/9p24.1 amplification. JAK2 fluorescence in situ hybridization was compared with array-based comparative genomic hybridization in 34 samples of triple negative breast cancer tumor. By array-based comparative genomic hybridization, 15 had 9p24.1 copy-number gain (log2rati<40.3) and 19 were classified as non-gain (log2ratio≤0.3). Copy-number gain was defined as JAK2/CEN9 ratio ≥1.1 or average JAK2 signals≥3.0. Twelve of 15 samples with copy-number gain by array-based comparative genomic hybridization were also detected by fluorescence in situ hybridization. Eighteen of 19 samples classified as copy-number non-gain by array-based comparative genomic hybridization were concordant by array-based comparative genomic hybridization. The sensitivity and specificity of the fluorescence in situ hybridization assay was 80% and 95%, respectively (P=0.02). The sample with the highest level of amplification by array-based comparative genomic hybridization (log2ratio=3.6) also scored highest by fluorescence in situ hybridization (ratio=8.2). There was a correlation between the expression of JAK2 and amplification status (Mean 633 vs 393, P=0.02), and there was a trend of association with PD-L1 RNA expression (Mean 46 vs 22, P=0.11). No significant association was observed between PD-L1 immunohistochemistry expression and copy-number gain status. In summary, the novel array-based comparative genomic hybridization assay for detection of chromosome 9p24.1 strongly correlates with the detection of copy-number gain by array-based comparative genomic hybridization. In triple negative breast cancer, this biomarker may identify a relevant subset of patients for targeted molecular therapies.

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