6082 Background: Oral cavity squamous cell carcinoma (SCC) remains a common malignancy in the head and neck region, with challenges in tumor resection and recurrence prevention. Traditional methods like frozen-section analysis are limited by time delays, sampling errors, and tissue distortion. Reflectance Confocal Microscopy (RCM) provides a noninvasive alternative for real-time, high-resolution imaging, but interpreting RCM images accurately requires expert knowledge. Integrating artificial intelligence (AI) could improve the accuracy and reliability of RCM image interpretation for diagnosing SCC and assessing surgical margins. The integration of machine learning and artificial intelligence (AI) has the potential to enhance the accuracy and reliability of RCM image interpretation, providing a more efficient tool for diagnosing oral cavity SCC and assessing surgical margins in real-time during surgery. Methods: We developed an AI model using Google Cloud’s AutoML platform to classify RCM images for diagnosing oral cavity SCC and evaluating tumor margins. The dataset comprised 4,090 RCM images from 83 patients, including 1,998 images of benign tissue and 2,092 images of malignant tissue. The dataset was divided into training (80%), validation (10%), and test (10%) sets. A single-label classification approach was employed to differentiate benign and malignant tissue. Model performance was evaluated using sensitivity, specificity, accuracy, F1 score, and negative predictive value. Results: The AI model achieved an area under the curve (AUC) of 0.99, sensitivity of 98.09%, specificity of 95.00%, accuracy of 96.58%, and an F1 score of 96.70%. In comparison, expert human readers in our prior study achieved accuracies of 90.91% for normal tissue and 81.7% for tumor detection, highlighting the accuracy of the AI model's diagnostic performance. Conclusions: The combination of RCM imaging with AI-powered analysis provides an accurate, noninvasive method for real-time diagnosis and surgical margin assessment in oral cavity SCC. The AI-driven model has excellent sensitivity, specificity, and overall accuracy, offering a potentially efficient and reliable modality for the real-time evaluation of digital RCM images. This approach can reduce the time required for intraoperative margin assessment, minimize patient anesthesia time, and overcome challenges related to conventional histopathology, ultimately improving surgical outcomes in patients with oral cavity SCC.

Pritumumab is a human IgG1 kappa antibody that targets ecto-domain vimentin (EDV) which is overexpressed in several malignant tumors including glioblastomas. To understand preclinical biological activity and safety of pritumumab derived from Chinese hamster ovary (CHO) cells, we evaluated tumor targeting ability, brain-tumor barrier permeability, growth inhibition, and primate safety studies. In-vivo and ex-vivo imaging studies demonstrate pritumumab to cross the blood brain/blood tumor barrier and an 89Zr-labeled pritumumab immunoconjugate showed the antibody specifically targeted tumor cells. In mouse xenograft models, pritumumab inhibited the growth of U251 glioblastoma and PANC-1 pancreatic cancer cells. A 29-day intravenous toxicology study in cynomolgus monkeys was carried out to analyze the safety and toxicity of pritumumab, and no toxic effects were observed. Overall, these data together suggest pritumumab is biologically active and animal models can be used to further understand the various functions of the antibody. Clinical trials in brain cancer patients assessing safety and efficacy of pritumumab as a therapeutic for brain cancer are in process.

Supplementary Figure S2 displays in vivo Reflectance Confocal Microscopy (RCM) images of a tumor area in a 58-year-old male. This patient was a never smoker but had a history of 35 years of daily alcohol exposure. The tumor was identified as a 3.7 cm ulcerated moderately differentiated oral squamous cell carcinoma located on the left lateral tongue.

Kirsten rat sarcoma (KRAS) is a frequently mutated oncogene responsible for several oncogenic KRAS variants and for driving tumor proliferation. Some nonsmall cell lung cancer (NSCLC) tumors exhibit KRAS G12C mutations, which can be targeted for inhibition using covalent and more recently noncovalent inhibitors. Sotorasib was the first FDA-approved G12C inhibitor that has shown efficacy in lung cancer patients, but with mixed responses. The lack of efficacy can be attributed to tumor heterogeneity (lack of G12C mutations) and/or inefficient delivery. Targeted KRAS G12C imaging has potential to identify NSCLC lesions with the targeted mutation and elucidate the oncogene's role in driving tumor growth and correlating responses to treatment. Toward this goal, we have developed a sotorasib-based molecular agent for PET imaging and tested its efficacy in targeting tumor lesions with KRAS G12C mutations. Here, we describe the synthesis, in vitro and in vivo evaluation of an [124I]I-Sotorasib analog in targeting G12C mutant tumor lesions using PET imaging.

INTRODUCTION:Zirconium-89 (89Zr) is a positron emitter with several advantages over other shorter-lived positron emission tomography (PET) compatible radiometals such as gallium-68 or copper-64. These include practically unlimited availability, extremely low cost, greatly facilitated distribution logistics, positron energy fit for medical PET imaging, and sufficiently long physical half-life to enable PET imaging at later time points for patient-specific dosimetry estimations. Despite these apparent benefits, the reception of 89Zr in the nuclear medicine community has been tepid. The driving factor for the absence of broader adaptation is mostly routed in its final formulation - [89Zr]zirconium oxalate. While serving as a suitable precursor solution for the gold standard chelator deferoxamine (DFO), [89Zr]Zr-oxalate is inaccessible for the most commonly used chelators, such as the macrocyclic DOTA, due to its pre-chelated state. Consequently, pioneering work has been conducted by multiple research groups to create oxalate-free forms of [89Zr]Zr4+, either via chemical conversion of oxalate into other counterion forms or via direct radiochemical isolation of [89Zr]ZrCl4, showing that [89Zr]Zr-DOTA complexes are possible and stable. However, this success was accompanied by challenges, including complex and labor-intensive radiochemical processing and radiolabeling procedures as well as the relatively minuscule conversion rates. Here, we report on the direct production of [89Zr]ZrCl4 avoiding oxalate and metal contaminants to enable efficient radiolabeling of DOTA constructs. METHODS:We based our direct production of [89Zr]ZrCl4 on previously reported methods and further optimized its quality by including an additional iron-removing step using the TK400 Resin. Here, we avoided using oxalic acid and effectively minimized the content of trace metal contaminants. Our two-step purification procedure was automated, and we confirmed excellent radionuclide purity, minimal trace metals content, great reactivity over time, and high specific molar activity. In addition, DOTA-based PSMA-617 and DOTAGA-based PSMA-I&T were radiolabeled to demonstrate the feasibility of direct radiolabeling and to estimate the maximum apparent specific activities. Lastly, the biodistribution of [89Zr]Zr-PSMA-617 was assessed in mice bearing PC3-PIP xenografts, and the results were compared to the previously published data. RESULTS:A total of 18 batches, ranging from 6.9 to 20 GBq (186 to 541 mCi), were produced. The specific molar activity for [89Zr]ZrCl4 exceeded 0.96 GBq (26 mCi) per nanomole of zirconium. The radionuclidic purity was >99 %, and the trace metals content was in the <1 ppm range. The [89Zr]ZrCl4 remained in its reactive chemical form for at least five days when stored in cyclic olefin polymer (COP) vials. Batches of 11.1 GBq (300 mCi) of [89Zr]Zr-PSMA-617 and 14.4 GBq (390 mCi) of [89Zr]Zr-PSMA-I&T, corresponding to specific activities of 11.1 MBq/μg (0.3 mCi/μg), and 14.4 MBq/μg (0.39 mCi/μg), respectively, were produced. [89Zr]Zr-PSMA-617 animal PET imaging results were in agreement with the previously published data. CONCLUSION:In this work, we report on a suitable application of TK400 Resin to remove iron during [89Zr]ZrCl4 radiochemical isolation. The breakthrough allows for direct radiolabeling of DOTA-based constructs with [89Zr]ZrCl4, leading to high apparent molar activities and excellent conversion rates.

Abstract Purpose: We investigated reflectance confocal microscopy (RCM) as a possible noninvasive approach for the diagnosis of cancer and real-time assessment of surgical margins. Experimental Design: In a phase I study on 20 patients, we established the RCM imaging morphologic features that distinguish oral squamous cell carcinoma (OSCC) from normal tissue with a newly developed intraoral RCM probe. Our subsequent phase II prospective double-blinded study in 60 patients tested the diagnostic accuracy of RCM against histopathology. Five RCM videos from the tumor and five from normal surrounding mucosa were collected on each patient, followed by a 3-mm punch biopsy of the imaged area. An experienced RCM reader, who was blinded to biopsy location and histologic diagnosis, examined the videos from both regions and classified each as “tumor” or “not tumor” based on RCM features established in phase I. Hematoxylin and eosin slides from the biopsies were read by a pathologist who was blinded to RCM results. Using histology as the gold standard, we calculated the sensitivity and specificity of RCM. Results: We report a high agreement between the blinded readers (95% for normal tissue and 81.7% for tumors), high specificity (98.3%) and negative predictive values (96.6%) for normal tissue identification, and high sensitivity (90%) and positive predictive values (88.2%) for tumor detection. Conclusions: RCM imaging is a promising technology for noninvasive in vivo diagnosis of OSCC and for real-time intraoperative evaluation of mucosal surgical margins. Its inherent constraint, however, stems from the diminished capability to evaluate structures located at more substantial depths within the tissue.

Supplementary Video S4 demonstrates irregular tissue pattern. Carcinoma comprise cells arranged in a disorganized and irregular pattern.

Supplementary Figure S7 shows in vivo RCM images of a tumor area in a 75-year-old male, with a history of 30 packs/year of cigarette exposure and no history of alcohol use, harboring an exophytic 2 cm right lateral tongue moderately differentiated oral squamous cell carcinoma.

Introduction During radical prostatectomy (RP), preservation of the cavernosal nerves is paramount for erectile function and continence recovery. To date, nerve sparing is based on ambient light and is only improved by surgeon experience and understanding the anatomic relationship between the neurovascular bundle (NVB) and the prostate capsule. Tsp1a-Pra0-IR800 is a fluorescent-labeled peptide that targets the voltage-gated sodium channel Nav1.7 on peripheral nerves and fluoresces in the near-infrared (NIR) region. We have previously established that Tsp1a-Pra0-IR800 can visualize nerves in a mouse model following intravenous administration, highlighting its potential for use as an intraoperative agent to identify the NVB during RP. However, expression data of Nav1.7 in prostate cancer cells, which could interfere with nerve imaging, is not available. In this study, we sought to better characterize the expression of Nav1.7 for intra-prostatic nerves in patients who underwent RP for prostate cancer. Methods Following IRB approval, histopathologic slides from patients who underwent RP for prostate cancer at Memorial Sloan Kettering Cancer Center were evaluated by an experienced genitourinary pathologist. Nav1.7 immunohistochemistry (IHC) was performed using an anti-Nav1.7 rabbit monoclonal antibody. All slides were scanned to allow for digital analysis. For each slide, regions of interest (ROI) were delineated over 10 areas of suspected intra-prostatic nerves, 10 areas of prostate adenocarcinoma, and 10 areas of normal prostate tissue. Quantification of Nav1.7 expression for each ROI was performed using Fiji, an image processing package, and calculated as a percentage of total identified tissue area. Paired t-test was used to compare the average Nav1.7 expression between the three identified groups – nerve, adenocarcinoma, and normal prostate tissue. Results A total of 15 histopathologic slides from patients who underwent RP for prostate cancer were evaluated. The average expression positivity of Nav1.7 within the intra-prostatic nerves was 41.0 ± 21.5%. This was significantly greater than the expression in regions of adenocarcinoma (0.2 ± 0.5%, p < 0.001) and normal surrounding tissue (0.01 ± 0.02%, p < 0.001). Conclusions Our findings suggest that Nav1.7 expression is restricted to nerves within the prostate and not influenced by the presence of cancerous cells. Further research is planned to evaluate Nav1.7 as a potential target for intraoperative nerve identification during RP, including confirming Nav1.7 IHC expression within resected cavernosal nerves in men with higher risk prostate cancer.

Supplementary Figure S9 shows in vivo RCM images of a tumor area in a 47-year-old male, never smoker and with no history of alcohol exposure, harboring a 1.5 cm leukoplastic lesion on his left lateral tongue.

Surgeries and trauma result in traumatic and iatrogenic nerve damage that can result in a debilitating condition that approximately affects 189 million individuals worldwide. The risk of nerve injury during oncologic surgery is increased due to tumors displacing normal nerve location, blood turbidity, and past surgical procedures, which complicate even an experienced surgeon's ability to precisely locate vital nerves. Unfortunately, there is a glaring absence of contrast agents to assist surgeons in safeguarding vital nerves. To address this unmet clinical need, we leveraged the abundant expression of the voltage-gated sodium channel 1.7 (NaV1.7) as an intraoperative marker to access peripheral nerves in vivo, and visualized nerves for surgical guidance using a fluorescently-tagged version of a potent NaV1.7-targeted peptide, Tsp1a, derived from a Peruvian tarantula. We characterized the expression of NaV1.7 in sensory and motor peripheral nerves across mouse, primate, and human specimens and demonstrated universal expression. We synthesized and characterized a total of 10 fluorescently labeled Tsp1a-peptide conjugates to delineate nerves. We tested the ability of these peptide-conjugates to specifically accumulate in mouse nerves with a high signal-to-noise ratio in vivo. Using the best-performing candidate, Tsp1a-IR800, we performed thyroidectomies in non-human primates and demonstrated successful demarcation of the recurrent laryngeal and vagus nerves, which are commonly subjected to irreversible damage. The ability of Tsp1a to enhance nerve contrast during surgery provides opportunities to minimize nerve damage and revolutionize standards of care across various surgical specialties.

Supplementary Video S1 shows a Keratin Pearl. Keratin pearls are often present in oral squamous cell carcinoma (OSCC). They are composed of concentric rings of squamous cells showing gradual keratinization.

Supplementary Figure S6 shows in vivo RCM images of a tumor area in a 61-year-old male, with a recurrent 1 cm moderately differentiated invasive oral squamous cell carcinoma of the left ventral tongue.

Supplementary Table S3 shows the measures of association and effect for RCM features by presence/absence of tumor by H&E.

Supplementary Figure S10 shows in vivo RCM images of a tumor area in a 73-year-old male, with a 12-year/packs exposure to tobacco and daily alcohol exposure, harboring a 2.5 cm moderately differentiated oral squamous cell carcinoma of the left lateral tongue.

Abstract Background Fluorine 18-labelled tetrafluoroborate ([18F]TFB) is a substrate for the sodium/iodide symporter. In thyroid cancer, [18F]TFB-PET/CT may be an alternative to iodine imaging to evaluate the extent of disease, eligibility for radioiodine treatment, and success of redifferentiation therapies. We report the results of a pilot study to determine tumor uptake of [18F]TFB and compare its properties to [124I]IodinePET/CT in patients with metastatic thyroid cancer. Methods Five patients were included in a prospective study. All patients received PET/CT 1 h after injection of 356 ± 12 MBq [18F]TFB and were given 230 ± 9 MBq [124I]Iodine orally on the same day, followed by PET/CT after 48 h. Before redifferentiation therapy, patients underwent an additional baseline [124I]Iodine PET/CT. Cases were analyzed by two board-certified specialists. Detection rates and Spearman correlation for [18F]TFB and [124I]Iodine were calculated. Results Three patients had poorly differentiated thyroid cancer and received trametinib in a redifferentiation trial. Two patients had papillary thyroid cancer and did not receive redifferentiation therapy. Of the 33 lesions seen before/without redifferentiation therapy, 19 (58%) were visible on [18F]TFB and 30 (91%) on [124I]Iodine imaging. In the patients who underwent redifferentiation therapy, 48 lesions were newly seen on [124I]Iodine PET/CT with a median SUVmax of 3.3 (range, 0.4–285.0). All of these lesions were [18F]TFB-negative. Conclusion [18F]TFB failed to predict radioactive iodine uptake in patients with poorly differentiated thyroid cancer who underwent redifferentiation therapy with trametinib. It is unclear whether such discrepancies may also occur in other redifferentiation therapies or may even be encountered in redifferentiation-naïve thyroid cancer. Trial registration number NCT03196518, registered on June 22, 2017.

Supplementary Figure S8 shows in vivo RCM images of a tumor area in a 71-year-old male, never smoker and occasional alcohol exposure, harboring an ulcerated 1.5 cm moderately differentiated oral squamous cell carcinoma of the right lateral tongue.

Supplementary Video S2 shows enlarged multinucleated cells. Their presence usually indicates cell-cell fusion or incomplete cytokinesis.

Supplementary Figure S3 shows in vivo RCM images of a tumor area in a 74-year-old male, with 35 packs/years of smoking history and daily alcohol exposure. The patient harbored a leukoplastic lesion on the upper gingiva.