Immunohistochemical validation and efficacy of 68Ga-Trivehexin PET CT as molecular imaging agent for αvβ6 integrin expression in patients of Head and Neck Squamous Cell Carcinoma and Pancreatic Ductal Adenocarcinoma.

Introduction: αvβ6-integrin is exclusively expressed in epithelial cells and is upregulated in many carcinomas, such as Pancreatic Ductal Adenocarcinoma (PDAC) and head and neck squamous cell carcinoma (H&NSCC). Expression of αvβ6 integrin is almost always tumour specific with surrounding normal stromal tissue not expressing αvβ6. Trivehexin is a recently synthesized improved trimerized αvβ6-integrin selective nanopeptide which can be used as a diagnostic agent after being labelled with a positron emitter like 68Gallium. This is a pilot study to assess the potential role of 68Ga-Trivehexin positron emission tomography/computed tomography (PET/CT) in patients of head and neck squamous cell carcinoma and pancreatic ductal adenocarcinoma and their correlation with level of αvβ6-integrin expression by the tumour tissue on IHC.

Methods: 32 patients (19-76 years; 20 males and 12 females) with suspected Head and Neck Squamous Cell Carcinoma (n=19), or Pancreatic Ductal Adenocarcinomas (n=13) or metastasis underwent whole-body 68Ga-Trivehexin PET/CT scans and 18F-FDG PET/CT scan on two separate days. All 32 patients, subsequently, underwent tissue biopsy from the suspected primary/ metastatic tumor site for histopathological diagnosis and IHC for integrin expression. The PET/CT scans were analyzed visually and semi-quantitatively by two experienced Nuclear Medicine Physicians blinded to the results of the biopsy. An experienced pathologist categorized the degree of αvβ6-integrin expression on IHC using the immunoreactive score (IRS) and modified 4-point IRS classification.

Results: Biopsy from the lesions (primary and/or metastasis) demonstrated 9 cases of PDACs and 18 cases of various H&NSCCs (Buccal carcinoma, n= 4; carcinoma tongue, n= 8; carcinoma nasopharynx, n= 1; Carcinoma tonsil, n= 2; Carcinoma pharynx, n= 1; Tracheal carcinoma, n= 1; alveolar carcinoma, n= 1). 2 treated patients of H&NSCC (one carcinoma tonsil and one buccal carcinoma) with suspected disease recurrence showed benign inflammatory changes on histopathology.The 68Ga-Trivehexin PET images demonstrated increased tracer uptake in primary and metastatic lesions (mean SUVmax 5.9 ± 3.3) with good lesion delineation in 8 out of the 9 cases of PDACs. On the other hand, FDG PET showed increased tracer uptake (mean SUVmax 6.2 ± 2.6) in the lesions in 7 out of the 9 cases of PDACs Among various cases of H&NSCC, increased uptake of 68Ga-Trivehexin (mean SUVmax 6.6 ± 4.5) and 18F-FDG (mean SUVmax 12.7 ± 6.7) were seen in 17 out of the 18 patients (one case of tracheal carcinoma showed no significant uptake of either tracer). The two treated cases of H&NSCC with suspected recurrence, and biopsy result of inflammatory changes, showed increased tracer uptake in 18F-FDG PET (mean SUVmax 7.98 ± 3.1) and no uptake in 68Ga-Trivehexin PET (2.2 ± 0.34). The lesions demonstrated more intense 68Ga-Trivehexin uptake along the periphery of the lesions consistent with known higher expression of αvβ6 integrins along the tumour margins. IHC was performed in 38 tissue samples (from primary and metastatic sites) from the 27 patients of PDAC and H&NSCC for αvβ6 expression (primary lesion, n= 27; lymph node, n= 7; liver lesion, n= 3; deposit, n= 1).68Ga-Trivehexin PET showed positive uptake in 36 out of the 38 lesions positive for αvβ6 integrin expression on IHC with mean higher SUVmax seen in lesions with higher IRS score. 68Ga-Trivehexin SUV Max tumor showed a positive association with αvβ6 expression (correlation coefficient 0.6830) with significant p-value (p <0.0001).The 68Ga-Trivehexin PET images demonstrated better lesion delineation than FDG PET in the cases of PDACs expressing αvβ6 integrin due to the absence of any non specific uptake in the background.The overall sensitivity and specificity of 68Ga-Trivehexin PET CT scan in detecting a lesion was 97.7% and 100% respectively. Meanwhile, 18F-FDG PET CT scan demonstrated a sensitivity of 100%. Accuracy of 68Ga-Trivehexin PET CT scan was slightly higher at 97.8% than 18F-FDG scan at 96%

Conclusions: 68Ga-Trivehexin is a promising non-invasive molecular imaging agent for tumors expressing αvβ6 integrin, especially in cases where 18F-FDG PET/CT scan may be suboptimal due to its low uptake owing to low GLUT receptors, or due to its nonspecific uptake around tumor sites. The information obtained with 68Ga-Trivehexin PET could help surgeons and oncologists to optimize their treatment, facilitating complete surgical removal, or radiation treatment planning. It can itself emerge as a potent theranostic agent.