A novel human in vitro papillomavirus type 16 positive tonsil cancer cell line with high sensitivity to radiation and cisplatin

Abstract

BackgroundHuman papillomavirus (HPV) is an established risk factor for oropharyngeal squamous cell carcinoma (OSCC). The aim was to establish cell lines from HPV-positive tonsil carcinomas to be used for treatment development.MethodsFresh samples from 23 HPV-positive tonsil carcinomas were cultivated in vitro. The established cell line was analyzed for viral characteristics, cell karyotype, TP53 status, and growth capabilities in nude mice. In vitro studies of sensitivities to radiation, cisplatin and cetuximab were performed.ResultsAfter 19 months (eight passages), one cell line, LU-HNSCC-26, was established in vitro and also grew as xenografts. The tumor was from a 48 year old non-smoking man with non-keratinizing, p16 positive tonsil OSCC, stage T2N0M0 with HPV16. It contained 19.5 (CV% 3.7) HPV16 copies/cell (passage 8). The complete HPV16 genome sequence was obtained. Episomal HPV16 was present with an E2/E7 ratio of 1.1 (CV% 2.6). In addition, HPV16 mRNA specific for the intact E2 gene was detected. The viral expression manifested 1.0 (CV% 0.1) E7 mRNA copies per HPV16 genome. The karyotype was determined and the cell line demonstrated wild type TP53. The ID50 for radiation was 0.90 Gy and the IC50 for cisplatin was 0.99 μmol/L. The cell line was inhibited to a maximum of 18% by cetuximab.ConclusionsWe established an in vitro tonsil carcinoma cell line containing episomal HPV16. This is an important step towards efficient treatment development.Electronic supplementary materialThe online version of this article (10.1186/s12885-019-5469-8) contains supplementary material, which is available to authorized users.

Short half‐life of HPV16 E6 and E7 mRNAs sensitizes HPV16‐positive tonsillar cancer cell line HN26 to DNA‐damaging drugs

“…We wished to investigate how the HPV16‐positive tonsillar cancer cell line LU‐HNSCC‐26 (herein called HN26) responded to a series of cancer drugs, and to compare the effect of these drugs on HN26 cells with the effect of cisplatin on these cells. The HN26 tonsillar cancer cells have been shown previously to contain episomal HPV16 DNA and to produce HPV16 early mRNAs . The HN26 cells were incubated with 100uM each of the indicated cancer drugs from the “approved oncology drugs set IV” library obtained from the National Cancer Institute (NCI), USA (https://wiki.nci.nih.gov/display/NCIDTPdata/Compound+Sets)) for 24 h followed by MTT assay to determine the number of viable cells.…”

“…The HN26 cell line (full name LU-HNSCC-26) was isolated from a 48-year-old nonsmoking man diagnosed with poorly differentiated, p16-positive squamous cell tonsillar carcinoma stage T2N0M0 with HPV16 and has been previously described ( Table 1). 31 The C33A2 cell line is derived from the HPV-negative and p53-negative cervical cancer cell line C33A, 32 whereas 3,310 cells were generated by transfection and immortalization of normal human primary keratinocytes with full-length HPV16 genome plasmid pHPV16AN. 32,33 Application of inhibitors and cancer drugs including melphalan to cultured cells Cell culture medium was replaced with medium containing indicated concentrations of drugs for indicated time periods.…”

“…The HN26 tonsillar cancer cells have been shown previously to contain episomal HPV16 DNA and to produce HPV16 early mRNAs. 31 The HN26 cells were incubated with 100uM each of the indicated cancer drugs from the "approved oncology drugs set IV" library obtained from the National Cancer Institute (NCI), USA (https://wiki. nci.nih.gov/display/NCIDTPdata/Compound+Sets) for 24 h followed by MTT assay to determine the number of viable cells.…”

Proteasome Inhibitors Counteract the Effect of Cisplatin in HPV-Positive Squamous Cell Carcinoma in Vitro

“…The cell line LU-HNSCC-26 (HN26) was previously isolated from a low-grade, oropharyngeal, p16 positive, squamous cell tonsillar stage II (T2N0M0 [17]) carcinoma [14]. The cells were cultured in R10 medium (RPMI 1640 with stable glutamine supplemented with 1 mmol/L sodium pyruvate, 1 × MEM non-essential amino acids, 20 µg/mL gentamicin and 10% fetal bovine serum (FBS), all from GE Healthcare (Piscataway, NJ, USA)) in a humidi ed 37 °C incubator with 5% CO 2 .…”

“…We have previously established a HPV16-positive tonsil cell line with wild-type p53 named LU-HNSCC-26 (HN26) [14]. Except for this cell line, there are only eight reported HPV-positive HNSCC cell lines to our knowledge, and only one of those is of oropharyngeal origin but not from tonsil [15,16].…”