Lindsay S. Moore

In Vivo Fluorescence Immunohistochemistry: Localization of Fluorescently Labeled Cetuximab in Squamous Cell Carcinomas

Anti-EGFR (epidermal growth factor receptor) antibody based treatment strategies have been successfully implemented in head and neck squamous cell carcinoma (HNSCC). Unfortunately, predicting an accurate and reliable therapeutic response remains a challenge on a per-patient basis. Although significant efforts have been invested in understanding EGFR-mediated changes in cell signaling related to treatment efficacy, the delivery and histological localization in (peri-)tumoral compartments of antibody-based therapeutics in human tumors is poorly understood nor ever made visible. In this first in-human study of a systemically administered near-infrared (NIR) fluorescently labeled therapeutic antibody, cetuximab-IRDye800CW (2.5 mg/m2, 25 mg/m2, and 62.5 mg/m2), we show that by optical molecular imaging (i.e. denominated as In vivo Fluorescence Immunohistochemistry) we were able to evaluate localization of fluorescently labeled cetuximab. Clearly, optical molecular imaging with fluorescently labeled antibodies correlating morphological (peri-)tumoral characteristics to levels of antibody delivery, may improve treatment paradigms based on understanding true tumoral antibody delivery.

Oncologic Procedures Amenable to Fluorescence-guided Surgery

Objective: Although fluorescence imaging is being applied to a wide range of cancers, it remains unclear which disease populations will benefit greatest. Therefore, we review the potential of this technology to improve outcomes in surgical oncology with attention to the various surgical procedures while exploring trial endpoints that may be optimal for each tumor type. Background: For many tumors, primary treatment is surgical resection with negative margins, which corresponds to improved survival and a reduction in subsequent adjuvant therapies. Despite unfavorable effect on patient outcomes, margin positivity rate has not changed significantly over the years. Thus, patients often experience high rates of re-excision, radical resections, and overtreatment. However, fluorescence-guided surgery (FGS) has brought forth new light by allowing detection of subclinical disease not readily visible with the naked eye. Methods: We performed a systematic review of clinicatrials.gov using search terms “fluorescence,” “image-guided surgery,” and “near-infrared imaging” to identify trials utilizing FGS for those received on or before May 2016. Inclusion criteria: fluorescence surgery for tumor debulking, wide local excision, whole-organ resection, and peritoneal metastases. Exclusion criteria: fluorescence in situ hybridization, fluorescence imaging for lymph node mapping, nonmalignant lesions, nonsurgical purposes, or image guidance without fluorescence. Results: Initial search produced 844 entries, which was narrowed down to 68 trials. Review of literature and clinical trials identified 3 primary resection methods for utilizing FGS: (1) debulking, (2) wide local excision, and (3) whole organ excision. Conclusions: The use of FGS as a surgical guide enhancement has the potential to improve survival and quality of life outcomes for patients. And, as the number of clinical trials rise each year, it is apparent that FGS has great potential for a broad range of clinical applications.

Sensitivity and Specificity of Cetuximab-IRDye800CW to Identify Regional Metastatic Disease in Head and Neck Cancer

Purpose: Comprehensive cervical lymphadenectomy can be associated with significant morbidity and poor quality of life. This study evaluated the sensitivity and specificity of cetuximab-IRDye800CW to identify metastatic disease in patients with head and neck cancer. Experimental Design: Consenting patients scheduled for curative resection were enrolled in a clinical trial to evaluate the safety and specificity of cetuximab-IRDye800CW. Patients (n = 12) received escalating doses of the study drug. Where indicated, cervical lymphadenectomy accompanied primary tumor resection, which occurred 3 to 7 days following intravenous infusion of cetuximab-IRDye800CW. All 471 dissected lymph nodes were imaged with a closed-field, near-infrared imaging device during gross processing of the fresh specimens. Intraoperative imaging of exposed neck levels was performed with an open-field fluorescence imaging device. Blinded assessments of the fluorescence data were compared to histopathology to calculate sensitivity, specificity, negative predictive value (NPV), and positive predictive value (PPV). Results: Of the 35 nodes diagnosed pathologically positive, 34 were correctly identified with fluorescence imaging, yielding a sensitivity of 97.2%. Of the 435 pathologically negative nodes, 401 were correctly assessed using fluorescence imaging, yielding a specificity of 92.7%. The NPV was determined to be 99.7%, and the PPV was 50.7%. When 37 fluorescently false-positive nodes were sectioned deeper (1 mm) into their respective blocks, metastatic cancer was found in 8.1% of the recut nodal specimens, which altered staging in two of those cases. Conclusions: Fluorescence imaging of lymph nodes after systemic cetuximab-IRDye800CW administration demonstrated high sensitivity and was capable of identifying additional positive nodes on deep sectioning. Clin Cancer Res; 23(16); 4744–52. ©2017 AACR.

A ratiometric threshold for determining presence of cancer during fluorescence-guided surgery.

Fluorescence‐guided imaging to assist in identification of malignant margins has the potential to dramatically improve oncologic surgery. However, a standardized method for quantitative assessment of disease‐specific fluorescence has not been investigated. Introduced here is a ratiometric threshold derived from mean fluorescent tissue intensity that can be used to semi‐quantitatively delineate tumor from normal tissue.

Fluorescence imaging to localize head and neck squamous cell carcinoma for enhanced pathological assessment

Accurately identifying close or positive margins in real‐time permits re‐excision during surgical procedures. Intraoperative assessment of margins via gross examination and frozen section is a widely used tool to assist the surgeon in achieving complete resection. While this methodology permits diagnosis of freshly resected tissue, the process is fraught with misinterpretation and sampling errors. During fluorescence‐guided surgery, an exogenous fluorescent agent specific for the target disease is imaged in order to navigate the surgical excision. As this technique quickly advances into the clinic, we hypothesize that the disease‐specific fluorescence inherently contained within the resected tissues can be used to guide histopathological assessment. To evaluate the feasibility of fluorescence‐guided pathology, we evaluated head and neck squamous cell carcinoma tumour specimens and margins resected from animals and patients after systemic injection of cetuximab‐IRDye800CW. In a preclinical model of luciferase‐positive tumour resection using bioluminescence as the gold standard, fluorescence assessment determined by closed‐field fluorescence imaging of fresh resected margins accurately predicted the presence of disease in 33/39 positive margins yielding an overall sensitivity of 85%, specificity of 95%, positive predictive value (PPV) of 94%, and a negative predictive value (NPV) of 87%, which was superior to both surgical assessment (54%, 61%, 57%, and 58%) and pathological assessment (49%, 95%, 91%, and 66%), respectively. When the power of the technique was evaluated using human‐derived tumour tissues, as little as 0.5mg (1mm3) of tumour tissue was identified (tumour‐to‐background‐ratio:5.2). When the sensitivity/specificity of fluorescence‐guided pathology was determined using traditional histological assessment as the gold standard in human tissues obtained during fluorescence‐guided surgery, the technique was highly accurate with a sensitivity of 91%, specificity of 85%, PPV of 81%, and NPV of 93% for 90 human‐derived samples. This approach can be used as a companion to the pathologist, eliminating confounding factors while impacting surgical intervention and patient management.

Characterizing the Utility and Limitations of Repurposing an Open-Field Optical Imaging Device for Fluorescence-Guided Surgery in Head and Neck Cancer Patients

The purpose of this study was to assess the potential of U.S. Food and Drug Administration–cleared devices designed for indocyanine green–based perfusion imaging to identify cancer-specific bioconjugates with overlapping excitation and emission wavelengths. Recent clinical trials have demonstrated potential for fluorescence-guided surgery, but the time and cost of the approval process may impede clinical translation. To expedite this translation, we explored the feasibility of repurposing existing optical imaging devices for fluorescence-guided surgery. Methods: Consenting patients (n = 15) scheduled for curative resection were enrolled in a clinical trial evaluating the safety and specificity of cetuximab-IRDye800 (NCT01987375). Open-field fluorescence imaging was performed preoperatively and during the surgical resection. Fluorescence intensity was quantified using integrated instrument software, and the tumor-to-background ratio characterized fluorescence contrast. Results: In the preoperative clinic, the open-field device demonstrated potential to guide preoperative mapping of tumor borders, optimize the day of surgery, and identify occult lesions. Intraoperatively, the device demonstrated robust potential to guide surgical resections, as all peak tumor-to-background ratios were greater than 2 (range, 2.2–14.1). Postresection wound bed fluorescence was significantly less than preresection tumor fluorescence (P < 0.001). The repurposed device also successfully identified positive margins. Conclusion: The open-field imaging device was successfully repurposed to distinguish cancer from normal tissue in the preoperative clinic and throughout surgical resection. This study illuminated the potential for existing open-field optical imaging devices with overlapping excitation and emission spectra to be used for fluorescence-guided surgery.

On the horizon: Optical imaging for cutaneous squamous cell carcinoma

Surgical resection with negative margins remains the standard of care for high‐risk cutaneous squamous cell carcinoma (SCC). However, surgical management is often limited by poor intraoperative tumor visualization and inability to detect occult nodal metastasis. The inability to intraoperatively detect microscopic disease can lead to additional surgery, tumor recurrence, and decreased survival.

Fluorescence Imaging for Cancer Screening and Surveillance

The advent of fluorescence imaging (FI) for cancer cell detection in the field of oncology is promising for both cancer screening and surgical resection. Particularly, FI in cancer screening and surveillance is actively being evaluated in many new clinical trials with over 30 listed on Clinical Trials.gov. While surgical resection forms the foundation of many oncologic treatments, early detection is the cornerstone for improving outcomes and reducing cancer-related morbidity and mortality. The applications of FI are twofold as it can be applied to high-risk patients in addition to those undergoing active surveillance. This technology has the promise of highlighting lesions not readily detected by conventional imaging or physical examination, allowing disease detection at an earlier stage of development. Additionally, there is a persistent need for innovative, cost-effective imaging modalities to ameliorate healthcare disparities and the global burden of cancer worldwide. In this review, we outline the current utility of FI for screening and detection in a range of cancer types.

Antiangiogenic antibody improves melanoma detection by fluorescently labeled therapeutic antibodies

Evaluate if vascular normalization with an antiangiogenic monoclonal antibody improves detection of melanoma using fluorescently labeled antibody‐based imaging.

Laser-Assisted Indocyanine Green Dye Angiography for Postoperative Fistulas After Salvage Laryngectomy

Importance Pharyngocutaneous fistula formation is an unfortunate complication after salvage laryngectomy for head and neck cancer that is difficult to anticipate and related to a variety of factors, including the viability of native pharyngeal mucosa. Objective To examine whether noninvasive angiography with indocyanine green (ICG) dye can be used to evaluate native pharyngeal vascularity to anticipate pharyngocutaneous fistula development. Design, Setting, and Participants This cohort study included 37 patients enrolled from June 1, 2013, to June 1, 2016, and follow-up was for at least 1 month postoperatively. The study was performed at the University of Alabama at Birmingham, a tertiary care center. Included patients were those undergoing salvage total laryngectomy who were previously treated with chemoradiotherapy or radiotherapy alone. Exposures The ICG dye was injected intraoperatively, and laser-assisted vascular imaging was used to evaluate the native pharyngeal mucosa after the ablative procedure. The center of the native pharyngeal mucosa was used as the reference to compare with the peripheral mucosa, and the lowest mean ICG dye percentage of mucosal perfusion was recorded for each patient. Main Outcomes and Measures The primary outcome was the formation of a postoperative fistula, which was assessed by clinical and radiographic assessment to test the hypothesis formulated before data collection. Results A total of 37 patients were included (mean [SD] age, 62.3 [8.5] years; 32 [87%] male and 5 [14%] female); 20 had a history of chemoradiotherapy, and 17 had history of radiotherapy alone. Thirty-four patients (92%) had free flap reconstruction, and 3 had primary closure (8%). Ten patients (27%) developed a postoperative fistula. No significant difference was found in fistula rate between patients who underwent neck dissection and those who did not and patients previously treated with chemoradiotherapy and those treated with radiotherapy alone. A receiver operator characteristic curve was generated to determine the diagnostic performance of the lowest mean ICG dye percentage of mucosal perfusion determined by fluorescence imaging, which was found to be a threshold value of 26%. The area under the curve was 0.85 (95% CI, 0.73-0.97), which was significantly greater than the chance diagonal. The overall mean lowest ICG dye percentage of mucosal perfusion was 31.3%. The mean lowest ICG dye percentage of mucosal perfusion was 22.0% in the fistula group vs 34.9% in the nonfistula group (absolute difference, 12.9%; 95% CI, 5.1%-21.7%). Conclusions and Relevance Patients who developed postoperative fistulas had lower mucosal perfusion as detected by ICG dye angiography when compared with patients who did not develop fistulas.