Non-contact Biopsies for Endoscopy
Endoscopy remains the gold standard technique for finding and assessing multiple types of cancers residing in internal organs and structures. This includes all gastrointestinal cancers, all head and neck cancers, most lung cancers, bladder cancers, cervix and vaginal cancers, as well as laproscopic and thorascopic evaluations for malignant disease. The real benefit of endoscopy is that it allows a minimally invasive approach to see inside cavities and obtain tissue specimens (biopsies) that allow histopathologic confirmation of cancers. The sensitivity, specificity, and ability to obtain tissue specimen makes endoscopy irreplaceable in its role in cancer care. Although advances in imaging have been suggested as an alternative to endoscopy, there are still significant short-comings that relegate non-invasive imaging to being used as a second-line approach or as an adjunct to endoscopy. In our lab, we are developing optical endoscopy techniques to provide functional and molecular marker imaging, which allows in situ diagnosis and characterization of cancer’s microenvironment (ex: oxygenation status), aggressiveness, and sensitivity to targeted therapies.
Each year gastrointestinal (GI) diseases impact 60 to 70 million people in the United States alone, more than any other common disease group. GI diseases represent a spectrum of conditions including common benign afflictions such as gastroesophageal reflux disease (GERD) with a prevalence of 20% of North Americans, and life-threatening problems such as colorectal cancer. In GI diseases, endoscopy is the most common diagnostic maneuver, with 15 million procedures annually for colonoscopies alone. Endoscopy provides a minimally invasive visual assessment of otherwise inaccessible tissues, identifying polyps, macroscopic lesions, bleeding and other tissue abnormalities. However, current endoscopy techniques are limited by the discordance between macroscopic superficial features and microscopic tissue morphology, and the pragmatic limits on the number of biopsies that can be safely obtained. The current workflow requires clinicians to perform endoscopic biopsies on areas of macroscopic interest, and the tissue fragments then undergo histopathological assessment. In the case of colorectal cancer diagnosis, a limited number of biopsies are taken for histological analysis.
Indeed, the number of samples collected is usually restricted to reduce the risks of bleeding, infection, and perforation. Additionally, collection of multiple samples can be complicated by bleeding from ulcerated tumors. While there are standard guidelines for biopsy site selection, the lack of real-time microscopic morphological feedback risks the underdiagnosis of malignant diseases. Ultimately, biopsy location is decided by the judgement of the clinician and the collected specimens may not be fully representative of the disease, leading to delays in definitive diagnosis and associated poor patient outcomes. These problems could be mitigated by gastroenterologists receiving real-time histological information through an endoscopic tool, allowing live in-situ “virtual biopsies”. Such a technique would improve the diagnostic accuracy of endoscopy biopsies by providing rapid and in-situ morphological tissue assessment to ensure that adequate and diagnostic tissue samples were obtained for definitive histopathological analysis.
An ideal virtual biopsy device would provide in-situ contrast akin to standard staining processes such as hematoxylin and eosin (H&E); this would permit rapid adoption without extensive user re-training. Providing H&E like contrast label-free in-situ requires imaging of endogenous nuclear contrast. Furthermore, imaging thick tissues in-situ requires a reflection mode architecture. While meeting these design criteria, the virtual biopsy system should conform to the stringent restrictions (<15 mm diameter) on device construction for viable endoscopy-based tools. The requirements for a label-free reflection-mode architecture rule out the use of many emerging histopathological imaging techniques. While Stimulated Raman Spectroscopy has presented H&E like images without requiring exogenous labeling, this system typically operates in transmission mode requiring thin sectioned tissue. Microscopy with ultraviolet surface excitation (MUSE) is a reflection mode system showing promising H&E-like images, but requires exogenous dyes to achieve contrast. Optical coherence tomography (OCT) meets most of the desired criteria and have been implemented as endoscopes. However, while OCT can image cellular scale structures in human tissues, the scattering contrast mechanism does not permit the biomolecule-specific contrast required to generate H&E-like images. To simulate H&E, OCT systems must use external processing techniques, such as artificial intelligence-based image processing.
Photoacoustic remote sensing microscopy (PARS®) is a recently reported imaging technique that provides virtual biopsy capabilities. PARS® captures the endogenous optical absorption contrast of biomolecules in an all optical, label-free non-interferometric reflection mode architecture. A picosecond to nanosecond scale pulsed excitation laser is used to deposit optical energy into the sample, inducing local refractive index modulations proportional to the absorbed optical energy. These modulations are observed by a secondary co-focused continuous wave detection laser. The use of optical absorption contrast provides high imaging specificity compared to alternative techniques. PARS® can recover real-time nuclear morphology in a miniaturized footprint reminiscent of H&E staining. Furthermore, the reflection-mode architecture of PARS® readily enables imaging of thick tissues. .
(a) Bright field H&E image of a normal colonic mucosa in sectioned formalin fixed paraffin embedded (FFPE) form. Scale bar: 100 μm (b) Wide field of view PARS® image of unstained sectioned FFPE normal colonic mucosa biopsy. Scale bar: 200 μm (c) Small field PARS® image of unstained sectioned FFPE normal colonic mucosa biopsy. Scale bar: 100 μm. Artificial H&E-like coloring has been applied to the PARS® images.