Targeting DNA damage response in head and neck cancers through abrogation of cell cycle checkpoints

Abstract

Purpose

Head and neck cancers (HNSCC) are routinely treated with radiotherapy; however, normal tissue toxicity remains a concern. Therefore, it is important to validate treatment modalities combining molecularly targeted agents with radiotherapy to improve the therapeutic ratio. The aim of this study was to assess the ability of the PARP inhibitor niraparib (MK-4827) alone, or in combination with cell cycle checkpoint abrogating drugs targeting Chk1 (MK-8776) or Wee1 (MK-1775), to radiosensitize HNSCCs in the context of HPV status.

Materials and methods

PARP1, PARP2, Chk1 or Wee1 shRNA constructs were analyzed from an in vivo shRNA screen of HNSCC xenografts comparing radiosensitization differences between HPV(+) and HPV(−) tumors. Radiosensitization by niraparib alone or in combination with MK-8776 or MK-1775 was assessed by clonogenic survival in HPV(−) and HPV(+) cells; and the role of p16 in determining response was explored. Relative expressions of DNA repair genes were compared by PCR array in HPV(+) and HPV(−) cells, and following siRNA-mediated knockdown of TRIP12 in HPV(−) cells.

Results

In vivo shRNA screening showed a modest preferential radiosensitization by Wee1 and PARP2 in HPV(−) and Chk1 in HPV(+) tumor models. Niraparib alone enhanced the radiosensitivity of all HNSCC cell lines tested. However, HPV(−) cells were sensitized to a greater degree, as suggested by the shRNA screen. When combined with MK-8776 or MK-1775, radiosensitization was further enhanced in an HPV dependent manner with HPV(+) cells enhanced by MK-8776 and HPV(−) cells enhanced by MK-1775. A PCR array for DNA repair genes showed PARP and HR proteins BRCA1 and RAD51 were much lower in HPV(+) cells than in HPV(−). Similarly, directly knocking down p16-dependent TRIP12 decreased expression of these same genes. Overexpressing p16 decreased TRIP12 expression and increased radiosensitivity in HPV(−) HN5. However, while PARP inhibition led to significant radiosensitization in the control, it led to no further significant radiosensitization in p16 overexpressing cells. Forced p16 expression in HPV(−) HN5 increased accumulation in G1 and subG1 and limited progression to S phase, thus reducing effectiveness of PARP inhibition.

Conclusions

Niraparib effectively radiosensitizes HNSCCs with a greater benefit seen in HPV(−). HPV status also plays a role in response to MK-8776 or MK-1775 when combined with niraparib due to differences in DNA repair mechanisms. This study suggests that using cell cycle abrogators in combination with PARP inhibitors may be a beneficial treatment option in HNSCC, but also emphasizes the importance of HPV status when considering effective treatment strategies.

Keywords:

• HPV
• PARP
• Chk1
• Wee1
• HNSCC

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

This work was supported by the National Cancer Institute under Grant [R01 CA168485 (HS)]; the National Institute of Dental and Craniofacial Research under Grants [R01 DE028105 (HS)], [R01 DE028061 (CP and HS)], [U01DE025181 (JM and MF)]; and the Cancer Research Institute of Texas (CPRIT) by [RP150293]. The Characterized Cell Line Core Facility was supported by the Cancer Center Support Grant [P30 NIH CA016672].

Notes on contributors

Jessica M. Molkentine

Jessica M. Molkentine, BS, is a Research Scientist and Laboratory Manager in the Department of Radiation Oncology at the University of Pittsburgh/UPMC Hillman Cancer Center.

David P. Molkentine

David P. Molkentine, BS, is a Research Scientist in the Department of Radiation Oncology at the University of Pittsburgh/UPMC Hillman Cancer Center.

Kathleen A. Bridges

Kathleen A. Bridges, MS, is a Research Investigator in the Department of Experimental Radiation Oncology at The University of Texas MD Anderson Cancer Center.

Tongxin Xie

Tongxin Xie, MD, PhD, is an Instructor in the Department of Head and Neck Surgery at The University of Texas MD Anderson Cancer Center.

Liangpeng Yang

Liangpeng Yang, PhD, is a Research Scientist in the Department of Radiation Oncology at The University of Texas MD Anderson Cancer Center.

Aakash Sheth

Aakash Sheth, BS, is a Medical Student at Rutgers Robert Wood Johnson Medical School.

Timothy P. Heffernan

Timothy P. Heffernan, PhD, is Executive Director and Head of Oncology Research for the Institute for Applied Cancer Science at The University of Texas MD Anderson Cancer Center.

David A. Clump

David A. Clump II, MD, PhD, is an Assistant Professor in the Department of Radiation Oncology at the University of Pittsburgh/UPMC Hillman Cancer.

Alma Z. Faust

Alma Z. Faust, PhD, is Director of Faculty Research Activities at the University of Illinois at Chicago.

Robert L. Ferris

Robert L. Ferris, MD, PhD, is Director of the UPMC Hillman Cancer Center, Hillman Professor of Oncology, Associate Vice Chancellor for Cancer Research, and Professor of Otolaryngology, of Immunology, and of Radiation Oncology.

Jeffrey N. Myers

Jeffrey N. Myers, MD, PhD, is Chair of the Department of Head and Neck Surgery at The University of Texas MD Anderson Cancer Center.

Mitchell J. Frederick

Mitchell J. Frederick, PhD, is an Associate Professor in the Department of Otolaryngology - Head and Neck Surgery at Baylor College of Medicine.

Kathryn A. Mason

Kathryn A. Mason, MS, is a retired Research Professor in the Department of Experimental Radiation Oncology at The University of Texas MD Anderson Cancer Center.

Raymond E. Meyn

Raymond E. Meyn, PhD, was a Professor of Experimental Radiation Oncology at the University of Texas MD Anderson Cancer Center. Over the course of his career, Dr. Meyn contributed an amazing amount to our understanding of radiobiology and radiosensitization. This work is a small tribute to his lasting contribution to the field.

Curtis R. Pickering

Curtis R. Pickering, PhD, is an Assistant Professor in the Department of Head and Neck Surgery at The University of Texas MD Anderson Cancer Center.

Heath D. Skinner

Heath D. Skinner, MD, PhD, is an Associate Professor and Hillman Fellow for Innovative Cancer Research at the University of Pittsburgh/UPMC Hillman Cancer Center.