Chemotherapy-Induced Tunneling Nanotubes Mediate Intercellular Drug Efflux in Pancreatic Cancer.

Intercellular communication plays a critical role in the ever-evolving landscape of invasive cancers. Recent studies have elucidated the potential role of tunneling nanotubes (TNTs) in this function. TNTs are long, filamentous, actin-based cell protrusions that mediate direct cell-to-cell communication between malignant cells. In this study, we investigated the formation of TNTs in response to variable concentrations of the chemotherapeutic drug doxorubicin, which is used extensively in the treatment of cancer patients. Doxorubicin stimulated an increased formation of TNTs in pancreatic cancer cells, and this occurred in a dose-dependent fashion. Furthermore, TNTs facilitated the intercellular redistribution of this drug between connected cells in both pancreatic and ovarian cancer systems in vitro. To provide supportive evidence for the relevance of TNTs in pancreatic cancer in vivo, we performed multiphoton fluorescence microscopy and imaged TNTs in tumor specimens resected from three human patients with pancreatic adenocarcinoma, and one with neuroendocrine carcinoma. In sum, TNT formation was upregulated in aggressive forms of pancreatic carcinoma, was further stimulated after chemotherapy exposure, and acted as a novel method for drug efflux. These findings implicate TNTs as a potential novel mechanism of drug resistance in chemorefractory forms of cancer.

A transwell assay that excludes exosomes for assessment of tunneling nanotube-mediated intercellular communication.

BACKGROUND: Tunneling nanotubes (TNTs) are naturally-occurring filamentous actin-based membranous extensions that form across a wide spectrum of mammalian cell types to facilitate long-range intercellular communication. Valid assays are needed to accurately assess the downstream effects of TNT-mediated transfer of cellular signals in vitro. We recently reported a modified transwell assay system designed to test the effects of intercellular transfer of a therapeutic oncolytic virus, and viral-activated drugs, between cells via TNTs. The objective of the current study was to demonstrate validation of this in vitro approach as a new method for effectively excluding diffusible forms of long- and close-range intercellular transfer of intracytoplasmic cargo, including exosomes/microvesicles and gap junctions in order to isolate TNT-selective cell communication. METHODS: We designed several steps to effectively reduce or eliminate diffusion and long-range transfer via these extracellular vesicles, and used Nanoparticle Tracking Analysis to quantify exosomes following implementation of these steps. RESULTS: The experimental approach outlined here effectively reduced exosome trafficking by >95%; further use of heparin to block exosome uptake by putative recipient cells further impeded transfer of these extracellular vesicles. CONCLUSIONS: This validated assay incorporates several steps that can be taken to quantifiably control for extracellular vesicles in order to perform studies focused on TNT-selective communication.

Lost in translation: applying 2D intercellular communication via tunneling nanotubes in cell culture to physiologically relevant 3D microenvironments.

Tunneling nanotubes (TNTs) are membranous conduits for direct cell-to-cell communication. Until the past decade, little had been known about their composite structure, function, and mechanisms of action in both normal physiologic conditions as well as in disease states. Now TNTs are attracting increasing interest for their key role(s) in the pathogenesis of disease, including neurodegenerative disorders, inflammatory and infectious diseases, and cancer. The field of TNT biology is still in its infancy, but inroads have been made in determining potential mechanisms and function of these remarkable structures. For example, TNTs function as critical conduits for cellular exchange of information; thus, in cancer, they may play an important role in critical pathophysiologic features of the disease, including cellular invasion, metastasis, and emergence of chemotherapy drug resistance. Although the TNT field is still in a nascent stage, we propose that TNTs can be investigated as novel targets for drug-based treatment of cancer and other diseases.

Pay for performance: will your hospital be ready?

With increased payment-or, in some cases, even the ability to contract with a payer-likely to be tied to the achievement of quality measures, providers need to: work with physicians in developing associated care delivery protocols, establish compliance mechanisms that are acceptable to clinicians and meet appropriate regulatory and legal standards.

Beyond saber rattling.

New legislative proposals are threatening aggressive regulation of tax-exempt healthcare providers. The proposals pertain to operations, governance, additional tax filing requirements, and enforcement. The proposals signal the determination of Congress to eliminate any perceived abuses among exempt healthcare organizations through increased scrutiny and regulation. Healthcare financial managers should monitor any developments related to the proposed legislation and participate in hearings and lobbying regarding these issues.

The new Medicare Advantage a disadvantage for providers?

The Medicare Advantage program, a provision of the Medicare Prescription Drug, Improvement, and Modernization Act of 2003, encourages providers to think about dealing with Medicare Advantage plans the way they now deal with commercial payers. Consequently, the Medicare Advantage program could either benefit or harm providers.

OIG targets contractual joint ventures.

A recent OIG Special Advisory Bulletin raises questions for providers involved in joint ventures. The Bulletin describes several characteristics that the OIG views as potentially suspect, including a referral stream controlled by the provider initiating the joint venture and the use of a wholly owned subsidiary of the provider to bill and collect for services. According to the OIG, profits paid by the subsidiary to the provider owner in such "suspect contractual joint ventures" could constitute illegal remuneration for referrals.

Sarbanes-Oxley raises red flag for not-for-profits.

In the near future, not-for-profits likely will feel the effects of these Sarbanes-Oxley directives: Establish audit committees with independent membership (precluding senior managers from being members) and make corresponding changes to corporate bylaws; Meet a higher standard for financial reporting, including increased disclosure and system certification representations as part of annual audits; Adopt a code of ethics for senior financial officers; Avoid senior executive compensation packages involving personal loans from the corporation; Ensure that board members are appropriately qualified and free of conflicts of interest.l