Role of telomeric RAP1 in radiation sensitivity modulation and its interaction with CSC marker KLF4 in colorectal cancer

Abstract

Aims: Radiotherapy is predominantly used as one of the treatment modalities to treat local tumor in colorectal cancer (CRC). Hindrance in disease treatment can be attributed to radio-tolerance of cancer stem cells (CSCs) subsistence in the tumor. Understanding the radio-resistant property of CSCs might help in the accomplishment of targeted radiotherapy treatment and increased disease-free survival. Telomeric RAP1 contributes in modulation of various transcription factors leading to aberrant cell proliferation and tumor cell migration. Therefore, we investigated the role of RAP1 in maintaining resistance phenotype and acquired stemness in radio-resistant cells.

Main methods: Characterization of HCT116 derived radio-resistant cell (HCT116RR) was performed by cell survival and DNA damage profiling. RAP1 silenced cells were investigated for DNA damage and expression of CSC markers through western blotting and Real-time PCR post-irradiation. Molecular docking and co-immunoprecipitation study were performed to investigate RAP1 and KLF4 interaction followed by RAP1 protein status profiling in CRC patient.

Key findings: We established radio-resistant cells, which showed tolerance to radiotherapy and elevated expression of CSC markers along with RAP1. RAP1 silencing showed enhanced DNA damage and reduced expression of CSC markers post-irradiation. We observed strong physical interaction between RAP1 and KLF4 protein. Furthermore, higher RAP1 expression was observed in the tumor of CRC patients. Dataset analysis also revealed that high expression of RAP1 expression is associated with poor prognosis.

Significance: We conclude that higher expression of RAP1 implicates its possible role in promoting radio-resistance in CRC cells by modulating DNA damage and CSC phenotype.

Keywords:

• Radiotherapy
• cancer stem cells
• genomic instability
• shelterin component
• radio-resistance

Acknowledgments

We thank Mr. Kshama Sagar Jena, Hemalata Hospital, Bhubaneswar for providing us the LINAC facility, Dr. Nachiketa Mahapatra, Apollo Hospital, Bhubaneswar for helping us in histopathological studies of patient tissues and members of MSSB group, KIIT School of biotechnology, Bhubaneswar for technical support.

Disclosure statement

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

Additional information

Funding

This work was supported by the Board of Research in Nuclear Science, Department of Atomic Energy, Government of India (Grant No.: 35/14/06/2015-BRNS/35081). Dr. Yusuf Akhter lab is funded by Indian Council of Medical Research (Government of India) and Department of Biotechnology (Government of India).

Notes on contributors

Kumari Anuja

Kumari Anuja, Ph.D. in Cancer Biology from School of Biotechnology, KIIT University, Bhubaneswar, India.

Madhabananda Kar

Madhabananda Kar, MBBS, MS in Surgical Oncology. He is head of Surgical Oncology Department in All India Institute of Medical Sciences (AIIMS), Bhubaneswar, India.

Amit Roy Chowdhury

Amit Roy Chowdhury, M.Sc. in Human Genetics and currently pursuing Ph.D. in School of Biotechnology, KIIT University, Bhubaneswar, India.

Gauri Shankar

Gauri Shankar, MPhil in Plant Biotechnology and currently pursuing as JRF in computational biology lab at Babasaheb Bhimrao Ambedkar University, Lucknow, India.

Swatishree Padhi

Swatishree Padhi, Ph.D. in Cancer Biology. She is currently working as Chief Scientific Officer in inDNA Lifesciences Pvt Ltd, Bhubaneswar, India.

Souvick Roy

Souvick Roy has completed M.Sc. in Biotechnology and currently pursuing Ph.D. in School of Biotechnology, KIIT University, Bhubaneswar, India.

Yusuf Akhter

Yusuf Akhter, Ph.D. in Biochemistry, He is an Assistant Professor in Babasaheb Bhimrao Ambedkar University, Lucknow, India.

Arabinda Kumar Rath

Arabinda Kumar Rath, Ph.D., MBA, Chairman and Managing Director of Hemalata Hospitals & Research Centre with around more than 20 years of experience as a specialist in Radiation Physics in Cancer Care and Teaching.

Birendranath Banerjee

Birendranath Banerjee, Ph.D. in Cancer Biology. He is Associate Professor and research scientist in School of Biotechnology, KIIT University, Bhubaneswar, India.