Purpose In robotic-assisted partial nephrectomy (RAPN), the use of intraoperative ultrasound (IOUS) helps to localise and outline the tumours as well as the blood vessels within the kidney. The aim of this work is to evaluate the use of the pneumatically attachable flexible (PAF) rail system for US 3D reconstruction of malignant masses in RAPN. The PAF rail system is a novel device developed and previously presented by the authors to enable track-guided US scanning. Methods We present a comparison study between US 3D reconstruction of masses based on: the da Vinci Surgical System kinematics, single-and stereo-camera tracking of visual markers embedded on the probe. An US-realistic kidney phantom embedding a mass is used for testing. A new design for the US probe attachment to enhance the performance of the kinematic approach is presented. A feature extraction algorithm is proposed to detect the margins of the targeted mass in US images. Results To evaluate the performance of the investigated approaches the resulting 3D reconstructions have been compared to a CT scan of the phantom. The data collected indicates that single camera reconstruction outperformed the other approaches, reconstructing with a sub-millimetre accuracy the targeted mass. Conclusions This work demonstrates that the PAF rail system provides a reliable platform to enable accurate US 3D reconstruction of masses in RAPN procedures. The proposed system has also the potential to be employed in other surgical procedures such as hepatectomy or laparoscopic liver resection.

Development of metastases to central nervous system (CNS) is an increasing clinical issue following the diagnosis of advanced breast cancer. The propensity to metastasize to CNS varies by breast cancer subtype. Of the four breast cancer subtypes, triple-negative breast cancers (TNBC) have the highest rates of both parenchymal brain metastasis and leptomeningeal metastasis (LM). LM is rapidly fatal due to poor detection and limited therapeutic options. Therapy of TNBC brain metastasis and LM is challenged by multifocal brain metastasis and diffuse spread of LM, and must balance brain penetration, tumor cytotoxicity, and the avoidance of neurotoxicity. Thus, there is an urgent need for novel therapeutic options in TNBCs CNS metastasis. QBS10072S is a novel chemotherapeutic that leverages TNBC-specific defects in DNA repair and LAT1 (Lamino acid transporter type 1)-dependent transport into the brain. In our study, activity of QBS10072S was investigated in vitro with various cell lines including the human TNBC cell line MDA-MB-231 and its brain-tropic derivative MDA-MB-231-BR3. QBS10072S was preferentially toxic to TNBC cells. The efficacy of QBS10072S against brain metastasis and LM was tested using a model of brain metastasis based on the internal carotid injection of luciferaseexpressing tumor cells into NuNu mice. The compound was well tolerated, delayed tumor growth and reduced leptomeningeal dissemination, resulting in significant extension of survival. Given that current treatments for LM are palliative with only few studies reporting a survival benefit, QBS10072S is planned to be investigated in clinical trials as a therapeutic for TNBC LM.Significance: TNBC brain metastasis often involves dissemination into leptomeninges. Treatment options for TNBC leptomeningeal metastasis are limited and are mostly palliative. Our study demonstrates significant efficacy of the brain-penetrating agent QBS10072S against TNBC brain metastasis and leptomeningeal spread.

Non-small cell lung cancer (NSCLC) metastatic to the brain leptomeninges is rapidly fatal, cannot be biopsied, and cancer cells in the cerebrospinal fluid (CSF) are few; therefore, available tissue samples to develop effective treatments are severely limited. This study aimed to converge single-cell RNA-seq and cell-free RNA (cfRNA) analyses to both diagnose NSCLC leptomeningeal metastases (LM), and to use gene expression profiles to understand progression mechanisms of NSCLC in the brain leptomeninges. NSCLC patients with suspected LM underwent withdrawal of CSF via lumbar puncture. Four cytology-positive CSF samples underwent single-cell capture (n = 197 cells) by microfluidic chip. Using robust principal component analyses, NSCLC LM cell gene expression was compared to immune cells. Massively parallel qPCR (9216 simultaneous reactions) on human CSF cfRNA samples compared the relative gene expression of patients with NSCLC LM (n = 14) to non-tumor controls (n = 7). The NSCLC-associated gene, CEACAM6, underwent in vitro validation in NSCLC cell lines for involvement in pathologic behaviors characteristic of LM. NSCLC LM gene expression revealed by single-cell RNA-seq was also reflected in CSF cfRNA of cytology-positive patients. Tumor-associated cfRNA (e.g., CEACAM6, MUC1) was present in NSCLC LM patients’ CSF, but not in controls (CEACAM6 detection sensitivity 88.24% and specificity 100%). Cell migration in NSCLC cell lines was directly proportional to CEACAM6 expression, suggesting a role in disease progression. NSCLC-associated cfRNA is detectable in the CSF of patients with LM, and corresponds to the gene expression profile of NSCLC LM cells. CEACAM6 contributes significantly to NSCLC migration, a hallmark of LM pathophysiology.

Leptomeningeal metastases (LM), a diffuse form of brain metastases is rare and fatal progression of non-small cell lung cancer (NSCLC). In LM, metastatic cancer cells spread and resign on the brain meninges, the cerebrospinal fluid (CSF), cranial and spinal nerves. Rapid disease progression and scarce tissue availability hinder the progress of scientific study of LM and its treatment. To overcome the critical lack of tissue and to determine the genetic profile of NSCLC LM, we have developed methods to extract tumor-associated cell-free RNA from CSF, and isolated and sequenced circulating single cells from CSF. Herein, we used high throughput qPCR to target lung and brain-associated genes and identified NSCLC LM metastases-related RNA. Brain-specific gene signature (GFAP, NRGN, SNCB, ZBTB18) was detected in all CSF sample (control and metastases), whereas lung-specific genes (MUC1, SFTPB, SFTPD, SLC34A2) were detected in CSF of brain metastases patients. Normal, healthy CSF lacks cellular component, but CSF of patients with LM metastases inhabited with very low amount of circulating tumor cells. Single cells from CSF of 4 patients with NSCLC LM metastases were captured with microfluidic chip. Cells (n = 197) were clustered by significantly differential expressed genes demonstrating two distinct populations of white blood and tumor cells. These data identified specific cfRNA and single cell transcriptome profiles compared to normal cells or patients without NSCLC LM metastases, and highlighted metastases-associated carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) as highly expressed in patients with NSCLC LM metastases. CEACAM6 mRNA was detected in CSF of 86% of patients with NSCLC LM but not in the CSF of control patients. In vitro inhibition of CEACAM6 protein lead to decreased invasion in NSCLC cells which was rescued by overexpression of the protein. We have developed sensitive and robust techniques to leverage human CSF to study NSCLC LM.