A successful, and novel, technique to kill metastatic breast cancer cells by circumventing their chemo- and radioresistant mechanisms was by presented by Dr John Giannios, Head of Radiotherapeutic Cancer Research at the IASO Hospital, Athens, Greece at the 18th Meeting of the European Association for Cancer Research on Tuesday 6 July 2004.
Advanced breast cancer, with metastases to lung and bone, has a very poor prognosis and current treatment protocols for this stage of disease generally result in survival periods of less than two years. One of the reasons for this poor prognosis is that metastatic cancer cells are less responsive to treatment than primary tumour cells. This is partly caused by the fact that the normal cell death process (apoptosis) is repressed by the overexpression of oncogenes such as bcl-2, HER-2, Raf-1 and cdc25c (these oncogenes are expressed more strongly in metastatic tumour cells), which means that the cells fail to die following treatment with chemotherapy drugs and radiation therapy.
Using metastatic tumour tissue taken from a patient with advanced breast cancer, Dr Giannios’s team analysed the cells to determine if known oncogenes were being overexpressed. In addition to finding overexpression of the oncogenes bcl-2, HER-2, Raf-1 and cdc25c they also detected overexpression of DNMT1 (a DNA methyltransferase, involved in DNA replication during cell division, and implicated in cancer development) and they also detected methylation of BRCA1 promoter (a process implicated specifically in the development of breast cancer tumours).
The experimental treatment, termed ‘chemoradioimmunotherapy’, combined chemotherapy, radiation therapy and immunotherapy in one. The chemotherapy component consisted of vinorelbine-tartrate (a cytotoxic drug used in the treatment of breast (and other) cancers), the radiotherapy component was provided through the addition of high energy radioisotopes, whilst the immunotherapy aspect was achieved by attaching an antibody specific to HER-2 to those radioisotopes, as well as through the inclusion of a separate 21-nucleotide double stranded siRNA (‘small interfering RNA’) generated against DNMT1.
It was hoped that the novel treatment regime would effectively target the tumour cells by blocking the genetic mechanisms that protect the cells from conventional treatment thereby allowing the chemotherapy and radiation therapy components to exert their cytotoxic effects.
By 24 hours post-treatment there was clear evidence that the treated tumour cells were undergoing significantly greater apoptosis than the untreated controls. Apoptosis was confirmed by the detection of activation of caspase-3-9 (an enzyme involved in apoptosis), inhibition of DNA synthesis and metabolic activity in the tumour cells and the formation of apoptotic bodies. These apoptotic bodies were seen to be phagocytosed (absorbed) by adjacent tumour cells, which resulted in the subsequent apoptosis of the tumour cells through a ‘bystander’ killing effect.
Several diagnostic tests were employed to determine the molecular basis for the observed success of the chemoradioimmunotherapy treatment. The tests proved that the novel regimen had specifically impacted on the identified oncogenes that are essential to the propagation and perpetuation of the tumour cells. Evidence was found to show 1) there was clear downregulation of HER-2 as a consequence of the action of antiHER-2 scFv antibody; 2) there was re-expression of the tumour suppressor gene BRCA1 as a consequence of the inhibition of the DNMT1 mRNA and; 3) the radioisotopes had induced DNA double strand breaks in the tumour cells. The combination of these molecular actions was responsible for circumvention of chemo- and radioresistant mechanisms in the tumour cells, allowing them to be effectively targeted and damaged by the chemotherapy and radiation therapy components leading to induction of apoptosis.
According to Dr Giannios, “This technique will be very applicable in a clinical setting where treatment difficulties will be limited because, as a tailored and targeted anti-cancer treatment, the treatment will reduce systemic toxicity whilst enhancing the therapeutic index.” “Introducing the radiation by linking the radioisotopes to the anti-HER-2 antibody is more efficient than conventional external beam radiotherapy because the radiation is targeted specifically to those breast cancer cells that over-express HER-2/neu, leaving normal cells unaffected and thereby reducing system toxicity”, he added.
“These results open the possibility of combining targeted immunotherapy with chemotherapy and radiation therapy to successfully kill metastatic tumour cells”, said Dr Giannios. “Theoretically this novel technique should be as effective in other types of cancer that are characterised by hypermethylation of tumour suppressor genes and the overexpression of oncogenes such as HER-2 and bcl-2”. “Our next step will be to develop the treatment in patients, and on a bigger scale, in a Phase I clinical trial”.