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Sains Malaysiana 52(5)(2023):
http://doi.org/10.17576/jsm-2023-5205-12
Proportion of CD44+ Subset of Tumour Cells in Single Cell Suspension Prepared from FFPET Sections
Directly Correlates with Histological Subtyping of Head and Neck Squamous
Cell Carcinoma
(Bahagian CD44+ Subset Sel Tumor
dalam Suspensi Sel Tunggal Disediakan daripada Bahagian FFPET Berkorelasi Terus
dengan Pengetipan Histologi Karsinoma Sel Skuamosa Kepala dan Leher)
MUHAMMAD
KASHIF1,2,*, SADIA MINHAS3, SHAH JAHAN1, AFRA
SAMAD4, FAHEEM SHAHZAD1, ROMEEZA TAHIR1, ABDUL
HANAN NAGI5 & NADEEM AFZAL1
1Department
of Immunology, University of Health Sciences, 54000, Khyaban e Jamia Punjab,
Lahore, Pakistan
2Department
of Oral Pathology, Bakhtawar Amin Medical and Dental College, Northern Bypass
Road, 60000, Multan, Pakistan
3Department
of Oral Pathology, Akhtar Saeed Medical and Dental College, Bahria Town, 54000,
Lahore, Pakistan
4Department
of Histopathology, Multan Medical and Dental College, 60000, Southern bypass
road, Multan, Pakistan
5Chughtai Laboratory, Lahore, Pakistan
Diserahkan: 19 Disember 2022/Diterima: 17
April 2023
Abstract
CD44 expression in tumours imparts potential to progress,
metastasize, recurrence, and resistance against antineoplastic therapy. In this
study, we sought to describe the variation in the immuno-expression and
numeration of MDR1+ and CD44+ potential cancer stem cells
in different histological grades and subtypes of head and neck squamous cell
carcinoma (HNSCC). Flow-cytometric
analysis was performed on single cell suspension prepared from formalin fixed
paraffin embedded tissue (FFPET) sections of HNSCC using anti-CD44 and
anti-MDR1/ABCB-1 primary monoclonal antibodies. Immunohistochemical (IHC)
staining was also carried out using both of these antibodies on HNSCC tissue
sections mounted on super frosted glass slides. On
immunohistochemical analysis, the mean IRS for CD44 and MDR1 were 8.6364
±3.02114 and 1.5909 ±1.27674 respectively. When mean immune-expression scores
of CD44 antibody and MDR1/ABCB-1 were compared with histological grades and
subtypes of HNSCC, the relationship was found to be statistically
insignificant. Interestingly, a strong statistical difference (p = 0.000) was observed when the mean score of
subset of dysplastic squamous epithelial cells with characteristics of cell
stemness (CD326+CD44+) was compared among different
histological subtypes of HNSCC using flowcytometric analysis. While no
statistically significant association was observed when the mean score for
subset of dysplastic cells with potential of drug resistance (CD44+MDR1+)
was compared among different histological subtypes of HNSCC. Although potential cancer stem cell marker
CD44 and the multidrug resistance maker ABCB-1/MDR1 co-expressed in HNSCC but
the proportion of CD326+CD44+ subset of tumour cells
(potential cancer stem cells/CSCs) significantly correlates with least
aggressive to more aggressive tumour subtypes.
Keywords: CD44; CSCs; FFPET; flowcytometry;
HNSCC; immunohistochemistry; multidrug resistance; OSCC
Abstract
Pengekspresan CD44 dalam tumor
memberikannya potensi untuk berkembang, bermetastasis, berulang dan berintangan
terhadap terapi antineoplastik. Dalam kajian ini, kami ingin menerangkan variasi dalam
pengekspresan imun dan penbilangan sel stem kanser berpotensi MDR1+ dan CD44+ dalam gred histologi yang berbeza dan pengetipan karsinoma
sel skuamosa kepala dan leher (HNSCC). Analisis aliran-sitometri dilakukan pada penggantungan
sel tunggal yang disediakan daripada bahagian tisu terbenam parafin tetap
formalin (FFPET) HNSCC menggunakan antibodi monoklonal utama anti-CD44 dan
anti-MDR1/ABCB-1. Pewarnaan imunohistokimia (IHC) juga dilakukan
menggunakan kedua-dua antibodi ini pada bahagian tisu HNSCC yang dipasang pada
slaid kaca super beku. Pada analisis imunohistokimia, purata IRS untuk CD44 dan
MDR1 masing-masing ialah 8.6364 ±3.02114 dan 1.5909 ±1.27674. Apabila min
skor pengekspresan imun antibodi CD44 dan MDR1/ABCB-1 dibandingkan dengan gred
histologi dan pengetipan HNSCC, hubungan itu didapati tidak signifikan secara
statistik. Menariknya, perbezaan statistik yang kuat (p = 0.000)
diperhatikan apabila skor min subset sel epitelium skuamosa displastik dengan
ciri-ciri pensteman sel (CD326+CD44+) dibandingkan antara
pengetipan histologi HNSCC yang berbeza menggunakan analisis sitometri aliran. Walaupun tiada
perkaitan yang signifikan secara statistik, diperhatikan apabila skor min bagi subset
sel displastik dengan potensi rintangan drug (CD44+MDR1+)
dibandingkan antara pengetipan histologi HNSCC yang berbeza. Walaupun
penanda sel stem kanser berpotensi CD44 dan pembuat rintangan pelbagai drug
ABCB-1/MDR1 dinyatakan bersama dalam HNSCC tetapi perkadaran subset CD326+CD44+ sel tumor (sel stem berpotensi kanser/CSC) berkorelasi dengan ketara dengan
tumor yang paling tidak agresif kepada pengetipan tumor yang lebih agresif.
Kata
kunci: CD44; CSCs; FFPET; HNSCC;
imunohistokimia; OSCC; rintangan
pelbagai ubat; sitometri aliran
RUJUKAN
Adelstein, D., Gillison, M.L.,
Pfister, D.G., Spencer, S., Adkins, D., Brizel, D.M., Burtness, B., , , Cmelak, A.J., Colevas, A.D., Eisele, D.W., Fenton, M., Foote, R.L., Gilbert, J., Haddad, R.I., Hicks, W.L., Hitchcock, Y.J., Jimeno, A., Leizman, D., Lydiatt, W.M., Maghami, E., Mell, L.K., Mittal, B.B., Pinto, H.A., Ridge, J.A., Rocco, J., Rodriguez, C.P., Shah, J.P., Weber, R.S., Witek, M., Worden, F., Yom, S.S., Zhen, W., Burns, J.L. & Darlow 2017. NCCN guidelines insights: Head and neck cancers,
Version 2. Journal of the National
Comprehensive Cancer Network 15(6): 761-770. https://doi.org/10.6004/jnccn.2017.0101
Akhter, M., Hossain, S., Rahman,
Q.B. & Molla, M.R. 2011. A study on histological grading of oral squamous
cell carcinoma and its co-relationship with regional metastasis. Journal of Oral and Maxillofacial Pathology 15(2): 168. https://doi.org/10.4103/0973-029x.84485
Arantes, L.M.R.B., De Carvalho,
A.C., Melendez, M.E. & Carvalho, A.L. 2017. Serum, plasma and saliva
biomarkers for head and neck cancer. Expert
Review of Molecular Diagnostics 18(1): 85-112. https://doi.org/10.1080/14737159.2017.1404906
Becker, M. & Levy, D. 2017. Modeling the transfer of
drug resistance in solid tumors. Bulletin
of Mathematical Biology 79(10): 2394-2412. https://doi.org/10.1007/s11538-017-0334-x
Bhave, S.L., Teknos, T.N. & Pan,
Q. 2011. Molecular parameters of head and neck cancer metastasis. Critical Reviews in Eukaryotic Gene
Expression 21(2): 143-153. https://doi.org/10.1615/CritRevEukarGeneExpr.v21.i2.40 PMID:22077153
Boxberg, M., Götz, C., Haidari, S.,
Dorfner, C., Jesinghaus, M., Drecoll, E., Boskov, M., Wolff, K.D., Weichert,
W., Haller, B. & Kolk, A. 2018. Immunohistochemical expression of CD44 in
oral squamous cell carcinoma in relation to histomorphological parameters and
clinicopathological factors. Histopathology 73(4): 559-572. https://doi.org/10.1111/his.13496
Breier, A., Gibalova, L., Seres, M.,
Barancik, M. & Sulova, Z. 2013. New insight into P-glycoprotein as a drug
target. Anti-Cancer Agents in Medicinal
Chemistry- Anti-Cancer Agents 13(1): 159-170. https://doi.org/10.2174/187152013804487380
Brock, I., Hipfner, D.R., Nielsen,
B.S., Jensen, P.B., Deeley, R.G., Cole, S.P. & Sehested, M. 1995.
Sequential co-expression of the multidrug resistance genes MRP and Mdr1 and
their products in VP-16 (Etoposide)-Selected H69 small cell lung cancer cells. Cancer Research 55(3): 459-462. https://pubmed.ncbi.nlm.nih.gov/7834606/
Chaudhary, P.M. & Roninson, I.B.
1993. Induction of multidrug resistance in human cells by transient exposure to
different chemotherapeutic drugs. JNCI
Journal of the National Cancer Institute 85(8): 632-639. https://doi.org/10.1093/jnci/85.8.632
Chen, D. & Wang, C.Y. 2019.
Targeting cancer stem cells in squamous cell carcinoma. Precision Clinical Medicine 2(3): 152-165. https://doi.org/10.1093/pcmedi/pbz016
Chen, J., Zhou, J., Lu, J., Xiong,
H., Shi, X. & Gong, L. 2014. Significance of CD44 expression in head and
neck cancer: A systemic review and meta-analysis. BMC Cancer 14: 15. https://doi.org/10.1186/1471-2407-14-15
Garcia, A.S., Assao, A., Carvalho,
A.L., Soares, F.A., Kowalski, L.P. & Oliveira, D.T. 2019. The stem cell
markers expression CD44v6 and podoplanin in lip cancer: Clinical significance. Virchows Archiv: An International Journal of
Pathology 474(6): 745-754. https://doi.org/10.1007/s00428-019-02539-3
Ghosh, S.C., Alpay, S.N. &
Klostergaard, K. 2012. CD44: A validated target for improved delivery of cancer
therapeutics. Expert Opinion on
Therapeutic Targets 16(7): 635-650. https://doi.org/10.1517/14728222.2012.687374
Govindan, S.V., Kulsum, S., Pandian,
R.S., Das, D., Seshadri, M., Hicks Jr., W., Kuriakose, M. A. & Suresh, A.
2015. Establishment and characterization of triple drug resistant head and neck
squamous cell carcinoma cell lines. Molecular
Medicine Reports 12(2): 3025-3032. https://doi.org/10.3892/mmr.2015.3768
Gupta, B., Johnson, N.W. &
Kumar, N. 2016. Global epidemiology of head and neck cancers: A continuing
challenge. Oncology 91(1): 13-23. https://doi.org/10.1159/000446117
Hedley, D.W., Friedlander, M.L., Taylor, I.W., Rugg,
C.A. & Musgrove, E.A. 1983. Method for analysis of cellular DNA content of
paraffn-embedded pathological material using flow cytometry. J. Histoch. Cytochem. 31:
1333-1335.
Johnson, D.E., Burtness, B.,
Leemans, C.R., Lui, V.W.Y., Bauman, J.E. & Grandis, J.R. 2020. Head and
neck squamous cell carcinoma. Nature
Reviews Disease Primers 6(1): 1-22.
Joshua, B., Kaplan, M.J., Doweck,
I., Pai, R., Weissman, I.L., Prince, M.E. & Ailles, L.E. 2012. Frequency of
cells expressing CD44, a head and neck cancer stem cell marker: Correlation
with tumor aggressiveness. Head &
Neck 34(1): 42-49. https://doi.org/10.1002/hed.21699
Kashif, M., Ishfaq, M. & Nagi,
A.H. 2015. Expression of prostanoid EP3 receptors in oral squamous epithelium
and oral squamous cell carcinoma. Pathology
Research International 2015 (February): 1-7. https://doi.org/10.1155/2015/602929
Mărgăritescu, C.I.,
Pirici, D., Simionescu, C. & Stepan, A. 2011. The utility of CD44, CD117
and CD133 in identification of cancer stem cells (CSC) in oral squamous cell
carcinomas (OSCC). Romanian Journal of
Morphology and Embryology = Revue Roumaine de Morphologie et Embryologie 52(3 Suppl): 985-993. https://pubmed.ncbi.nlm.nih.gov/22119814/
Minhas, S., Kashif, M., Altaf, W.
& Nagi, A.H. 2016. Oral squamous cell carcinoma: Epidemiological, clinical
and histological features. Rawal Medical
Journal 41(1): 81-85. http://www.rmj.org.pk/?mno=199313
Prince, M.E., Sivanandan, R.,
Kaczorowski, A., Wolf, G.T., Kaplan, M.J., Dalerba, P., Weissman, I.L., Clarke,
M.F. & Ailles, L.E. 2007. Identification of a subpopulation of cells with
cancer stem cell properties in head and neck squamous cell carcinoma. Proceedings of the National Academy of
Sciences of the United States of America 104(3): 973-978. https://doi.org/10.1073/pnas
Scully, C. & Bagan, J. 2009.
Oral squamous cell carcinoma overview. Oral
Oncology 45: 301-308.
Shen, C., Xiang, M., Nie, C., Hu,
H., Ma, Y. & Wu, H. 2013. CD44 as a molecular marker to screen cancer stem
cells in hypopharyngeal cancer. Acta
Oto-Laryngologica 133(11): 1219-1226. https://doi.org/10.3109/00016489.2013.811750
Sterz, C.M., Kulle, C., Dakic, B.,
Makarova, G., Böttcher, M.C., Bette, M., Werner, J.A. & Mandic, R. 2010. A
basal-cell-like compartment in head and neck squamous cell carcinomas
represents the invasive front of the tumor and is expressing MMP-9. Oral Oncology 46(2): 116-122. https://doi.org/10.1016/j.oraloncology.2009.11.011
Su, J., Xu, X.H., Huang, Q., Lu,
M.Q., Li, D.J., Xue, F., Yi, F., Ren, J.H. & Wu, Y.P. 2011. Identification
of cancer stem-like CD44+ cells in human nasopharyngeal carcinoma cell line. Archives of Medical Research 42(1):
15-21. https://doi.org/10.1016/j.arcmed.2011.01.007
Vikram, R., Chou, W.C., Hung, S.C.
& Shen, C.Y. 2020. Tumorigenic and metastatic role of
CD44−/Low/CD24−/Low cells in luminal breast cancer. Cancers 12(5): 1239. https://doi.org/10.3390/cancers12051239
Yang, X., lyer, A.K., Singh, A.,
Choy, E., Hornicek, F.J., Amiji, M.M. & Duan, Z. 2015. MDR1 SiRNA loaded
hyaluronic acid-based CD44 targeted nanoparticle systems circumvent paclitaxel
resistance in ovarian cancer. Scientific Reports 5: 8509. https://doi.org/10.1038/srep08509
Zou, F., Seike, M., Noro, R.,
Kunugi, S., Kubota, K. & Gemma, A. 2017. Prognostic significance of ABCB1
in Stage I lung adenocarcinoma. Oncology
Letters 14(1): 313-321. https://doi.org/10.3892/ol.2017.6145
*Pengarang untuk surat-menyurat;
email: drkashifazam@gmail.com
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