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Genes that mediate metastasis to the lung in breast cancer: PTK7.

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Shahan Mamoor at Boston College
Shahan Mamoor
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Abstract and Figures

Metastasis to the lung is a medical problem in patients with breast cancer (1-8). The druggability of kinases is an attractive feature for rapid identification and implementation of novel chemotherapies (9, 10). We predict therapeutic index is intrinsically linked to differential expression which can be comprehensively and blindly determined using whole transcriptome data (11). We mined published RNA-sequencing and microarray data (12, 13) using only human tumor tissues to compare primary and metastatic tumor transcriptomes for the discovery of kinases associated with lung metastasis in human breast cancer. We report here differential expression of protein tyrosine kinase 7 (inactive) encoded by PTK7 in the lung metastases of patients with metastatic breast cancer. PTK7 mRNA was present at increased quantities in lung metastatic tissues as compared to primary tumors of the breast. Modulation of PTK7 expression may be relevant to the biology by which tumor cells metastasize from the breast to the lung in humans with metastatic breast cancer.
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Genes that mediate metastasis to the lung in breast cancer: PTK7.
Shahan Mamoor, B.S., M.S., Ph.D cand.1
1shahanmamoor@gmail.com
East Islip, NY USA
Metastasis to the lung is a medical problem in patients with breast cancer1-8. The druggability of kinases is
an attractive feature for rapid identification and implementation of novel chemotherapies9,10. We predict
therapeutic index is intrinsically linked to differential expression which can be comprehensively and
blindly determined using whole transcriptome data11. We mined published RNA-sequencing and
microarray data12,13 using only human tumor tissues to compare primary and metastatic tumor
transcriptomes for the discovery of kinases associated with lung metastasis in human breast cancer. We
report here differential expression of protein tyrosine kinase 7 (inactive) encoded by PTK7 in the lung
metastases of patients with metastatic breast cancer. PTK7 mRNA was present at increased quantities in
lung metastatic tissues as compared to primary tumors of the breast. Modulation of PTK7 expression
may be relevant to the biology by which tumor cells metastasize from the breast to the lung in humans
with metastatic breast cancer.
Keywords: breast cancer, metastasis, lung metastasis, protein tyrosine kinase 7 (inactive), PTK7, systems
biology of breast cancer, targeted therapeutics in breast cancer.
1
In humans with breast cancer, metastasis occurs, among other sites, to the lymph nodes, the brain,
the bone, the liver, and the lung. We performed comparative transcriptome analysis of primary breast and
lung metastatic tumors in patients with metastatic breast cancer using published RNA-sequencing data12.
We also utilized a separate, published microarray gene expression dataset13 to determine if target
differential expression was a shared feature of lung metastasis among patients with breast cancer.
Methods
We used datasets GSE19123012 and GSE5649313 for this global differential gene expression
analysis of lung metastasis in breast cancer in conjunction with GEO2R. GSE191230 was generated using
HiSeq X Ten technology with n=6 primary breast tumors and n=2 lung metastases from patients with
breast cancer; analysis was performed using platform GPL20795; all patients were HER2+. GSE56493
was generated using Rosetta/Merck Human RSTA Custom Affymetrix 2.0 microarray technology with
n=19 primary breast tumors and n=2 lung metastases from patients with breast cancer; analysis was
performed using platform GPL10379. The Benjamini and Hochberg method of p-value adjustment was
used for ranking of differential expression but raw p-values were used to assess statistical significance of
global differential expression. Log-transformation of data was auto-detected, and the NCBI generated
category of platform annotation was used.
Results
We performed comparative transcriptome analysis of metastatic tumor tissues from patients with
metastatic breast cancer using published RNA-sequencing and microarray data12,13 for rapid discovery,
validation and translation of novel therapeutic targets.
PTK7 is differentially expressed in the lung metastases of patients with lung metastatic breast
cancer.
We identified protein tyrosine kinase 7 (inactive), encoded by PTK7, as a differentially expressed
gene in the lung metastatic tissues of humans with breast cancer (Chart 1). When sorting each of the genes
expressed in lung metastases based on significance of difference as compared to primary tumors of the
breast, PTK7 ranked 2560 out of 22997 total transcripts (Chart 1), equating to 88.9% differential
expression. Differential expression of PTK7 in the lung metastases of patients with breast cancer was
statistically significant (Chart 1; p= 1.81E-02).
We interrogated a second (microarray) dataset13 for target validation, comparing total gene
expression between lung metastases and primary tumors of the breast, again revealing differential
expression of PTK7 in metastasis to the lung in humans with breast cancer (Chart 2). When sorting each of
the genes expressed in lung metastases as compared to primary tumors of the breast, PTK7 ranked 8025
out of 52378 total transcripts, equating to 84.7% differential expression (Chart 2). Differential expression
of PTK7 in the lung metastases of patients with lung metastatic breast cancer was statistically significant
(Chart 2; p=2.12E-01).
Differential expression of PTK7 in the lung metastases of women with metastatic breast cancer,
both identified blindly and through whole transcriptome methodologies, and validated across laboratories,
suggested that differential expression of PTK7 is a transcriptional feature of metastasis to the lungs, rather
than an experimental artifact, a reflection of idiosyncratic behavior in cell culture or animal models, or an
observation from a single laboratory.
2
PTK7 is expressed at higher levels in the lung metastases of patients with metastatic breast cancer.
We obtained exact mRNA expression levels for PTK7, in primary tumors of the breast and in lung
metastasis of patients with metastatic breast cancer to determine direction and statistical significance of
change in PTK7 expression in lung metastases. PTK7 was expressed at higher levels in the lung
metastases of patients with breast cancer as compared to primary tumors of the breast, and this difference
was not statistically significant (Figure 1; p=0.1981). PTK7 was expressed at 15.43 ± 14.43 transcripts
per million in primary tumors of the breast, while it was expressed at 65.64 ± 26.13 transcripts per million
in lung metastasis from patients with breast cancer. We calculated a mean fold change of 4.25 in PTK7
mRNA levels when comparing lung metastases and primary tumors of the breast.
Thus, by mining published RNA-sequencing and microarray data12,13 in an unbiased fashion, we
identified protein tyrosine kinase 7 (inactive), encoded by PTK7, as among the genes whose expression
was most different, transcriptome-wide, in the lung metastases of patients with breast cancer; PTK7
messenger RNA was present at higher quantities in metastasis to the lung.
Discussion
We provided evidence here that protein tyrosine kinase 7 (inactive), encoded by PTK7, is among
the genes whose expression is most different in the lung metastases of patients with lung metastatic breast
cancer, and that PTK7 mRNA is present at increased quantities in lung metastatic tissues as compared to
primary tumors of the breast. Evaluation of the effects of genetic depletion of PTK7 in mouse models of
metastatic breast cancer on metastasis to the lung is merited. Modulation of PTK7 expression may be
relevant to the processes by which breast cancer cells exit the breast, enter the vasculature, evade immune
clearance, and colonize the lung.
3
References
1. Weigelt, B., Peterse, J.L. and Van't Veer, L.J., 2005. Breast cancer metastasis: markers and models.
Nature reviews cancer,5(8), pp.591-602.
2. Eckhardt, B.L., Francis, P.A., Parker, B.S. and Anderson, R.L., 2012. Strategies for the discovery
and development of therapies for metastatic breast cancer. Nature reviews Drug discovery,11(6),
pp.479-497.
3. Minn, A.J., Gupta, G.P., Siegel, P.M., Bos, P.D., Shu, W., Giri, D.D., Viale, A., Olshen, A.B.,
Gerald, W.L. and Massagué, J., 2005. Genes that mediate breast cancer metastasis to lung. Nature,
436(7050), pp.518-524.
4. Wagenblast, E., Soto, M., Gutiérrez-Ángel, S., Hartl, C.A., Gable, A.L., Maceli, A.R., Erard, N.,
Williams, A.M., Kim, S.Y., Dickopf, S. and Harrell, J.C., 2015. A model of breast cancer heterogeneity
reveals vascular mimicry as a driver of metastasis. Nature,520(7547), pp.358-362.
5. Landemaine, T., Jackson, A., Bellahcene, A., Rucci, N., Sin, S., Abad, B.M., Sierra, A., Boudinet,
A., Guinebretiere, J.M., Ricevuto, E. and Nogues, C., 2008. A six-gene signature predicting breast cancer
lung metastasis. Cancer research,68(15), pp.6092-6099.
6. Wculek, S.K. and Malanchi, I., 2015. Neutrophils support lung colonization of
metastasis-initiating breast cancer cells. Nature,528(7582), pp.413-417.
7. Fischer, K.R., Durrans, A., Lee, S., Sheng, J., Li, F., Wong, S.T., Choi, H., El Rayes, T., Ryu, S.,
Troeger, J. and Schwabe, R.F., 2015. Epithelial-to-mesenchymal transition is not required for lung
metastasis but contributes to chemoresistance. Nature,527(7579), pp.472-476.
8. Ye, X., Brabletz, T., Kang, Y., Longmore, G.D., Nieto, M.A., Stanger, B.Z., Yang, J. and
Weinberg, R.A., 2017. Upholding a role for EMT in breast cancer metastasis. Nature,547(7661),
pp.E1-E3.
9. Cohen, P., Cross, D. and Jänne, P.A., 2021. Kinase drug discovery 20 years after imatinib:
progress and future directions. Nature Reviews Drug Discovery,20(7), pp.551-569.
10. Attwood, M.M., Fabbro, D., Sokolov, A.V., Knapp, S. and Schiöth, H.B., 2021. Trends in kinase
drug discovery: targets, indications and inhibitor design. Nature Reviews Drug Discovery,20(11),
pp.839-861.
11. Muller, P.Y. and Milton, M.N., 2012. The determination and interpretation of the therapeutic index
in drug development. Nature Reviews Drug Discovery,11(10), pp.751-761.
12. Ong, L.T., Lee, W.C., Ma, S., Oguz, G., Niu, Z., Bao, Y., Yusuf, M., Lee, P.L., Goh, J.Y., Wang, P.
and Yong, K.S.M., 2022. IFI16-dependent STING signaling is a crucial regulator of anti-HER2 immune
response in HER2+ breast cancer. Proceedings of the National Academy of Sciences,119(31),
p.e2201376119.
13. Tobin, N.P., Lundberg, A., Lindström, L.S., Harrell, J.C., Foukakis, T., Carlsson, L., Einbeigi, Z.,
Linderholm, B.K., Loman, N., Malmberg, M. and Fernö, M., 2017. PAM50 provides prognostic
information when applied to the lymph node metastases of advanced breast cancer patients. Clinical
Cancer Research,23(23), pp.7225-7231.
4
Rank: 2560
p-value: 1.81E-02
stat: -2.364198
log2FoldChange: -1.9924487
baseMean: 1719.85
Gene: PTK7
Gene name: protein tyrosine kinase 7 (inactive)
Chart 1: PTK7 is differentially expressed in lung metastatic breast cancer when comparing lung
metastases to primary tumors of the breast.
The rank of global differential expression, the p-value with respect to differential expression
transcriptome-wide, stat (the Wald statistic), fold change in gene expression expressed as log2, basemean,
the average of normalized counts among all samples analyzed the gene and gene name are listed in this
chart.
5
Rank: 8025
probe ID: 100136051_TGI_at
p-value: 2.12E-01
t: -1.29
B: -4.6513
Gene: PTK7
Gene name: protein tyrosine kinase 7 (inactive)
Chart 2: PTK7 is differentially expressed in lung metastatic breast cancer when comparing lung
metastases to primary tumors of the breast.
The rank of global differential expression, probe/transcript ID, the p-value with respect to differential
expression transcriptome-wide, t, a moderated t-statistic, B, the log-odds of differential expression
between the groups compared, the gene and gene name are listed in this chart.
6
Figure 1: PTK7 is expressed at higher levels in the lung metastases of patients with metastatic breast
cancer when compared to primary tumors of the breast.
The mRNA expression level of PTK7 in primary tumors of the breast (left) and in lung metastases of
patients with metastatic breast cancer (right) is graphically depicted; the result of a statistical test
evaluating significance of difference in PTK7 expression between primary tumors and lung metastases is
listed above.
7
ResearchGate has not been able to resolve any citations for this publication.
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