Introduction 1 2 4 5 6 10 11 12 99m 13 99m 14 15 16 17 In the present study, we therefore aimed at assessing the feasibility of TAVS as a non-invasive technique to demonstrate treatment-induced apoptosis in vivo, in patients with HNSCC, early during treatment with concurrent chemoradiation. The purpose was to determine the degree of uptake on TAVS in normal tissue, primary tumour and lymph node metastases and to evaluate the treatment-induced Annexin uptake in relation to radiation dose. Furthermore, we questioned whether the differences in uptake would correlate with treatment response. Materials and methods 2 2 18 19 99m 99m Baseline diagnostic imaging with MRI (1.5-T system; Somatom; Siemens Medical Systems, Erlangen, Germany) or spiral CT (Tomoscan AVE1, Philips, Best, The Netherlands or HiSpeed CT, GE Medical Systems, USA) was performed within 3 weeks before the start of treatment and repeated 6–8 weeks after the end of the treatment for the evaluation of treatment response. Radiation treatment planning was done with our clinical treatment planning system (TPS; U-MPlan, University of Michigan, Ann Arbor, USA). Six patients were treated by IMRT, and the treatment plans were recalculated for the dose at the time of first post-treatment TAVS. The regions of interest (ROI) were delineated manually for each patient on the CT scan. These included the gross tumor volume (GTV) of the primary tumour and/or lymph nodes, the parotid glands and the submandibular glands. In the other seven patients, a standard three-field technique was used by virtual simulation with a CT scan. In these latter patients, the CT scan was imported into the TPS in which the delineations were done. The treatment fields were reconstructed, and the clinically applied dose distribution at the time of TAVS was recalculated in the TPS. The primary tumour and lymph node volumes were calculated by 3-D reconstruction of the delineated GTV. 20 U U U U U r p r p r p r p Evaluation of treatment Six to eight weeks after the end of treatment, the results of therapy were evaluated by means of radiological investigations (by MRI or CT scan and/or ultrasound) and examination under general anaesthesia, with biopsies taken in case of suspicious findings. For residual disease in the neck at the time of evaluation, salvage neck dissection was performed if the patient was judged operable. Follow-up visits were planned every 3 months in the first year after therapy, every 4 months in the second year and less frequent thereafter. A follow-up chest X-ray was performed annually. Statistical analysis t r P Results 1 99m 1 Fig. 1 solid line a b b a b  Table 1 Patient, tumour and treatment characteristics Patient number Gender Age (years) Tumor site TNM stage 3 U Interval cisplatin infusion and Annexin scan (h) RT technique Mode of cisplatin administration 1 M 64 Oral cavity III 21 62 52.2 Conventional 3 field Intra-arterially 2 M 58 Oropharynx III 47 203 50.3 IMRT Intra-arterially 3 F 65 Oropharynx IV 25 112 53.1 IMRT Intra-arterially 4 M 55 Oropharynx III 26 36 53.5 IMRT Intra-arterially 5 M 49 Hypopharynx II 13 43 53.3 IMRT Intra-arterially 6 F 66 Oral cavity IV 32 135 53.6 Conventional 3 field Intra-arterially 7 M 60 Oropharynx IV 147 191 54.3 Conventional 3 field Intra-arterially 8 M 46 Oropharynx IV 28 170 52.7 Conventional 3 field Intra-arterially 9 M 55 Oropharynx IV 41 42 49.9 IMRT Intravenously 10 M 57 Oropharynx IV 25 51 51.7 IMRT Intravenously 11 M 48 Oropharynx IV 91 117 51.5 Conventional 3 field Intravenously 12 M 50 Hypopharynx IV 100 132 50.3 Conventional 3 field Intravenously 13 M 26 Oropharynx IV 105 52 50.1 Conventional 3 field Intravenously M F IMRT Annexin uptake in normal tissue and tumour 7 14 21 2 p 3 U n n U r p 4 p 22 Fig. 2 diamonds squares a b c  Fig. 3 a c b d b d d  Fig. 4 U a Circles triangles b U  2 p U 4 2 p U r p 3 U r p 5 U r p Fig. 5 U colour y x  Response to treatment, locoregional control and survival U Discussion 4 23 24 23 26 27 28 2 U U 5 U 1 U 15 29 8 15 30 14 15 In conclusion, co-registration of Annexin V scintigraphy with radiotherapy-planning CT scan showed a radiation-dose-dependent uptake in parotid glands, indicative of early apoptosis during treatment. The inter-individual spread in Annexin uptake in primary tumours could not be related to differences in treatment schedule or tumour volume, but the Annexin uptake in tumour and lymph nodes were closely correlated. This effect might represent a tumour-specific apoptotic response.