Intratumoral injection of herpes simplex virus HF10 in recurrent breast cancer

A. Nakao*,1, H. Kimata1, T. Imai1, T. Kikumori1, O. Teshigahara1, T. Nagasaka2, F. Goshima3 and Y. Nishiyama3

Departments of 1 Surgery II, 2 Laboratory Medicine and 3 Virology, Graduate School of Medicine, Nagoya University, Nagoya, Japan

*E-mail: nakaoaki@med.nagoya-u.ac.jp

Breast cancer is one of the most common and feared cancers. Although there have been many advances in clinical studies, breast cancer remains a major contributor to the number of cancer deaths [1]. Advances in genetic engineering techniques have enabled investigators to develop more effective viruses for oncolytic viral therapy. We previously reported that a replication-competent, spontaneous herpes simplex virus-1 (HSV-1) variant, HF10 [2], was effective in treating disseminated peritoneal colon carcinoma and breast cancer in an immunocompetent mouse model [35]. To assess the therapeutic potential of HF10 in human disease, we did a preliminary study of toxicity and possible efficacy in six patients with recurrent metastatic breast cancer. All patients gave written informed consent, and the study was approved by the local ethics committee and institutional review board of our hospital. All six patients had pathologically proven recurrent breast cancer with cutaneous or subcutaneous metastatic nodules accessible to direct intratumoral injection. For each patient, a test nodule was injected with an HF10 diluent of various content [patient 1, 104 plaque-forming units (p.f.u.)/0.5 ml; patient 2, 105 p.f.u./0.5 ml; patient 3, 105 p.f.u./0.5 ml x 3 days; patient 4, 5 x 105 p.f.u./0.5 ml; patient 5, 5 x 105 p.f.u./0.5 ml x 3 days; patient 6, 5 x 105 p.f.u./ 0.5 ml x 3 days]. A second nodule was injected with 0.5 ml sterile saline. All patients were monitored for systemic adverse effects, and the injected nodules were examined for sign and size of inflammation. The injected nodules were excised in all patients 14 days after injection for examination by conventional haematoxylin and eosin staining and immunofluorescence examination using anti-herpes simplex virus type 1 (DAKO, Glostrup, Denmark) antibody. All patients tolerated HF10 well, were seropositive for HSV before virus injection, and their IgG and IgM titres did not substantially alter during the trial. There was no virus shedding or reactivation of endogenous latent HSV in any patient. No clinical change was seen in the HF10-injected nodules in any patient.

No patient showed evidence of regression of tumour mass at any other adjacent or distant tumour site. Pathological examination of excised nodules showed no cell death in the saline-injected nodules (Figure 1A). However, tumour cell death and nuclear viral inclusion bodies in breast cancer cells were observed in HF10-injected nodules (Figure 1B). Death was observed among 30–100% of cancer cells overall.



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Figure 1. Histopathological findings from nodules of recurrent breast cancer excised from patient 3 (stained with haematoxylin and eosin). (A) Control nodule. (B) Nodule excised 14 days after three injections each of 105 p.f.u./0.5 ml HF10 on days 1, 2 and 3. (C) Immunofluorescence findings of HF10-injected nodule excised from patient 3.

 
Immunofluorescence examination of excised HF10-injected nodules showed evidence of viral infection confined to breast cancer cells in all patients (Figure 1C). HF10 was seen in the breast cancer cells from the virus-treated nodules, with no antigen staining in the adjacent normal connective tissue. The fact that each of the patients was seropositive for HSV before HF10 injection indicates that the ability of the virus to replicate within tumour cells is not blocked by previous exposure to HSV.

This preliminary study shows that HF10 replicates in breast cancer cells to cause tumour cell death, and is not toxic. The results are sufficiently encouraging to continue HF10 in patients with metastatic breast cancer, as well as in other types of cancer.

REFERENCES

1. Weir HK, Thun MJ, Hankey BF et al. Annual report to the nation on the status of cancer, 1975–2000, featuring the uses of surveillance data for cancer prevention and control. J Natl Cancer Inst 2003; 95: 1276–1299.[Abstract/Free Full Text]

2. Nishiyama Y, Kimura H, Daikoku T. Complementary lethal invasion of the central nervous system by nonneuroinvasive herpes simplex virus type 1 and 2. J Virol 1991; 65: 4520–4524.[ISI][Medline]

3. Takakuwa H, Goshima F, Nozawa N et al. Oncolytic viral therapy using a spontaneously generated herpes simplex virus type 1 variant for disseminated peritoneal tumor in immunocompetent mice. Arch Virol 2003; 48: 813–825.[CrossRef]

4. Kimata H, Takakuwa H, Goshima F et al. Effective treatment of disseminated peritoneal colon cancer with new replication-competent herpes simplex viruses. Hepatogastroenterology 2003; 50: 961–966.[ISI][Medline]

5. Teshigahara O, Goshima F, Takao K et al. Oncolytic viral therapy for breast cancer with herpes simplex virus type 1 mutant HF10. J Surg Oncol 2004; 85: 42–47.[CrossRef][ISI][Medline]





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