Blood vessel abnormalities in glioblastoma: characterization at baseline, at recurrence and following radiotherapy suggest a therapy induced phenotype


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Anastasia Widyadari1,Anastasia Widyadari2,Michael Grant2,Aruna Chakrabarty3,Valerie Speir4,Susan Short2,Georgia Mavria2
1Leeds Institue of Medical Sciences,2Leeds Institute of Medical Sciences,3LEEDS TEACHING HOSPITALS NHS TRUST,4University of Aberdeen

Abstract

Background

Glioblastoma multiforme (GBM) is the most lethal form of brain tumour with an overall median survival of 12-15 months with current standard therapy. Histologically GBM is characterised by high cellularity, nuclear anaplasia, microvascular proliferation and necrosis. Radiotherapy and temozolomide chemotherapy are standard therapies following surgery. Abnormalities in the vascular niche are associated with defective vessel function and development of hypoxia, which promotes disease progression and resistance to therapy. However, the precise nature of the vascular abnormalities and how they are affected by radiotherapy is unknown.

Method

To determine the nature and prevalence of abnormal blood vessels in glioblastoma we characterised the vasculature of 25 primary and 15 paired primary and recurrent GBM patient samples by immunostaining for CD31, nestin, OLIG2, SOX2, GFAP, aSMA and PDGFRb. To assess potential effects of radiotherapy on blood vessels we also treated orthotopic CT2A tumours grown in syngeneic mice with clinically relevant doses of radiation (5Gy2x) using SARRP. Samples were collected at early and late timepoints after treatment and stained.

Results

We show that vessels with high nestin positivity, larger size and glomeruloid structures are common in GBM, and high prevalence is associated with poor prognosis. Nestin positive perivascular cells in these samples were PDGFRband a-SMA positive, but negative for OLIG2 and SOX2, or GFAP. Coverage by nestin positive/SMA positive cells and vessel size significantly increased in recurrent tumours compared to primary tumours prior to therapy. Analyses of experimental CT2A tumours suggested that irradiation reduced vessel size early after treatment but vascular abnormalities significantly increased at later stages of tumour re-growth.

Conclusion

Patient samples show that blood vessel abnormalities increase with GBM progression. Experimental in vivo data suggest that radiotherapy contributes to vascular changes post-treatment. This abnormal vascular phenotype may contribute to resistance to subsequent therapy, including anti-angiogenic therapy, and may be a valuable prognostic marker.