Dennis Jones

The lymph node microenvironment and its role in the progression of metastatic cancer

Lymph nodes are initial sites for cancer metastasis in many solid tumors. However, their role in cancer progression is still not completely understood. Emerging evidence suggests that the lymph node microenvironment provides hospitable soil for the seeding and proliferation of cancer cells. Resident immune and stromal cells in the lymph node express and secrete molecules that may facilitate the survival of cancer cells in this organ. More comprehensive studies are warranted to fully understand the importance of the lymph node in tumor progression. Here, we will review the current knowledge of the role of the lymph node microenvironment in metastatic progression.

Solid stress and elastic energy as measures of tumour mechanopathology

Solid stress and tissue stiffness affect tumour growth, invasion, metastasis and treatment. Unlike stiffness, which can be precisely mapped in tumours, the measurement of solid stresses is challenging. Here, we show that two-dimensional spatial mappings of solid stress and the resulting elastic energy in excised or in situ tumours with arbitrary shapes and wide size ranges can be obtained via three distinct and quantitative techniques that rely on the measurement of tissue displacement after disruption of the confining structures. Application of these methods in models of primary tumours and metastasis revealed that: (i) solid stress depends on both cancer cells and their microenvironment; (ii) solid stress increases with tumour size; and (iii) mechanical confinement by the surrounding tissue significantly contributes to intratumoural solid stress. Further study of the genesis and consequences of solid stress, facilitated by the engineering principles presented here, may lead to significant discoveries and new therapies.

Investigation of the Lack of Angiogenesis in the Formation of Lymph Node Metastases

BACKGROUND To date, antiangiogenic therapy has failed to improve overall survival in cancer patients when used in the adjuvant setting (local-regional disease with no detectable systemic metastasis). The presence of lymph node metastases worsens prognosis, however their reliance on angiogenesis for growth has not been reported. METHODS Here, we introduce a novel chronic lymph node window (CLNW) model to facilitate new discoveries in the growth and spread of lymph node metastases. We use the CLNW in multiple models of spontaneous lymphatic metastases in mice to study the vasculature of metastatic lymph nodes (n = 9-12). We further test our results in patient samples (n = 20 colon cancer patients; n = 20 head and neck cancer patients). Finally, we test the ability of antiangiogenic therapy to inhibit metastatic growth in the CLNW. All statistical tests were two-sided. RESULTS Using the CLNW, we reveal the surprising lack of sprouting angiogenesis during metastatic growth, despite the presence of hypoxia in some lesions. Treatment with two different antiangiogenic therapies showed no effect on the growth or vascular density of lymph node metastases (day 10: untreated mean = 1.2%, 95% confidence interval [CI] = 0.7% to 1.7%; control mean = 0.7%, 95% CI = 0.1% to 1.3%; DC101 mean = 0.4%, 95% CI = 0.0% to 3.3%; sunitinib mean = 0.5%, 95% CI = 0.0% to 1.0%, analysis of variance P = .34). We confirmed these findings in clinical specimens, including the lack of reduction in blood vessel density in lymph node metastases in patients treated with bevacizumab (no bevacizumab group mean = 257 vessels/mm(2), 95% CI = 149 to 365 vessels/mm(2); bevacizumab group mean = 327 vessels/mm(2), 95% CI = 140 to 514 vessels/mm(2), P = .78). CONCLUSION We provide preclinical and clinical evidence that sprouting angiogenesis does not occur during the growth of lymph node metastases, and thus reveals a new mechanism of treatment resistance to antiangiogenic therapy in adjuvant settings. The targets of clinically approved angiogenesis inhibitors are not active during early cancer progression in the lymph node, suggesting that inhibitors of sprouting angiogenesis as a class will not be effective in treating lymph node metastases.

Lymph node metastases can invade local blood vessels, exit the node, and colonize distant organs in mice

An alternate route for metastatic cells Metastatic tumor cells are thought to reach distant organs by traveling through the blood circulation or the lymphatic system. Two studies of mouse models now suggest a hybrid route for tumor cell dissemination. Pereira et al. and Brown et al. used distinct methodologies to monitor the fate of tumor cells in lymph nodes. They found that tumor cells could invade local blood vessels within a node, exit the node by entering the blood circulation, then go on to colonize the lung. Whether this dissemination route occurs in cancer patients is unknown; the answer could potentially change the way that affected lymph nodes are treated in cancer. Science, this issue p. 1403, p. 1408 In mice, tumor cells can metastasize to distant organs by entering blood vessels within the local lymph node. Lymph node metastases in cancer patients are associated with tumor aggressiveness, poorer prognoses, and the recommendation for systemic therapy. Whether cancer cells in lymph nodes can seed distant metastases has been a subject of considerable debate. We studied mice implanted with cancer cells (mammary carcinoma, squamous cell carcinoma, or melanoma) expressing the photoconvertible protein Dendra2. This technology allowed us to selectively photoconvert metastatic cells in the lymph node and trace their fate. We found that a fraction of these cells invaded lymph node blood vessels, entered the blood circulation, and colonized the lung. Thus, in mouse models, lymph node metastases can be a source of cancer cells for distant metastases. Whether this mode of dissemination occurs in cancer patients remains to be determined.

Deletion of the endothelial Bmx tyrosine kinase decreases tumor angiogenesis and growth

Bmx, [corrected] also known as Etk, is a member of the Tec family of nonreceptor tyrosine kinases. Bmx is expressed mainly in arterial endothelia and in myeloid hematopoietic cells. Bmx regulates ischemia-mediated arteriogenesis and lymphangiogenesis, but its role in tumor angiogenesis is not known. In this study, we characterized the function of Bmx in tumor growth using both Bmx knockout and transgenic mice. Isogenic colon, lung, and melanoma tumor xenotransplants showed reductions in growth and tumor angiogenesis in Bmx gene-deleted ((-/-)) mice, whereas developmental angiogenesis was not affected. In addition, growth of transgenic pancreatic islet carcinomas and intestinal adenomas was also slower in Bmx(-/-) mice. Knockout mice showed high levels of Bmx expression in endothelial cells of tumor-associated and peritumoral arteries. Moreover, endothelial cells lacking Bmx showed impaired phosphorylation of extracellular signal-regulated kinase (Erk) upon VEGF stimulation, indicating that Bmx contributes to the transduction of vascular endothelial growth factor signals. In transgenic mice overexpressing Bmx in epidermal keratinocytes, tumors induced by a two-stage chemical skin carcinogenesis treatment showed increased growth and angiogenesis. Our findings therefore indicate that Bmx activity contributes to tumor angiogenesis and growth.

In vivo label-free measurement of lymph flow velocity and volumetric flow rates using Doppler optical coherence tomography.

Direct in vivo imaging of lymph flow is key to understanding lymphatic system function in normal and disease states. Optical microscopy techniques provide the resolution required for these measurements, but existing optical techniques for measuring lymph flow require complex protocols and provide limited temporal resolution. Here, we describe a Doppler optical coherence tomography platform that allows direct, label-free quantification of lymph velocity and volumetric flow rates. We overcome the challenge of very low scattering by employing a Doppler algorithm that operates on low signal-to-noise measurements. We show that this technique can measure lymph velocity at sufficiently high temporal resolution to resolve the dynamic pulsatile flow in collecting lymphatic vessels.

Grazing-incidence x-ray reflectivity: studies for the AXAF Observatory

The energy bandwidth and total throughput of a grazing incidence optics system is a strong function of the X-ray reflectivity of the surface coating. In support of the Advanced X-ray Astrophysics Facility (AXAF), studies are underway to evaluate and characterize the reflectivity of potential AXAF coatings. Here we report on results obtained for Au, Ir, and Ni coatings produced by electron-beam evaporation, evaporation with ion-assist, and sputtering. Effects of coating thickness and deposition angle have been evaluated at 6.4 and 8.1 keV; the highest reflectivities are those of the thinner, about 200 A vs about 700 A, coatings. While considerable variations exist, the best Ir samples have higher reflectivity than any of the Au coatings. Data results have been compared with models for theoretical reflectivity, particularly with regard to the effective density of the coatings. Independent measurements of the coating densities have been carried out for comparison with the reflectivity results.

Methicillin-resistant Staphylococcus aureus causes sustained collecting lymphatic vessel dysfunction

Impairment in lymph flow caused by methicillin-resistant Staphylococcus aureus toxins persists after skin and soft tissue infections clear. Lymphatics limp along after MRSA Lymphedema is associated with skin and soft tissue infections, and both can be recurring, causing continual suffering in affected patients. To better understand the relationship between bacterial infections and lymphedema, Jones et al. used intravital imaging to examine the lymphatics of mice infected with MRSA. They observed lymphatic muscle cell death, which led to prolonged dysfunction months after the bacteria had been cleared. In vitro experiments with human cells indicated that bacterial toxins were responsible for damaging the lymphatic muscle cells, suggesting that the toxins could be targeted in patients to interrupt this brutal cycle. Methicillin-resistant Staphylococcus aureus (MRSA) is a major cause of morbidity and mortality worldwide and is a frequent cause of skin and soft tissue infections (SSTIs). Lymphedema—fluid accumulation in tissue caused by impaired lymphatic vessel function—is a strong risk factor for SSTIs. SSTIs also frequently recur in patients and sometimes lead to acquired lymphedema. However, the mechanism of how SSTIs can be both the consequence and the cause of lymphatic vessel dysfunction is not known. Intravital imaging in mice revealed an acute reduction in both lymphatic vessel contractility and lymph flow after localized MRSA infection. Moreover, chronic lymphatic impairment is observed long after MRSA is cleared and inflammation is resolved. Associated with decreased collecting lymphatic vessel function was the loss and disorganization of lymphatic muscle cells (LMCs), which are critical for lymphatic contraction. In vitro, incubation with MRSA-conditioned supernatant led to LMC death. Proteomic analysis identified several accessory gene regulator (agr)–controlled MRSA exotoxins that contribute to LMC death. Infection with agr mutant MRSA resulted in sustained lymphatic function compared to animals infected with wild-type MRSA. Our findings suggest that agr is a promising target to preserve lymphatic vessel function and promote immunity during SSTIs.

Lymph node effective vascular permeability and chemotherapy uptake

Lymph node metastases are a poor prognostic factor. Additionally, responses of lymph node metastasis to therapy can be different from the primary tumor. Investigating the physiologic lymph node blood vasculature might give insight into the ability of systemic drugs to penetrate the lymph node, and thus into the differential effect of therapy between lymph node metastasis and primary tumors. Here, we measured effective vascular permeability of lymph node blood vessels and attempted to increase chemotherapy penetration by increasing effective vascular permeability.

Growth and Immune Evasion of Lymph Node Metastasis

Cancer patients with lymph node (LN) metastases have a worse prognosis than those without nodal disease. However, why LN metastases correlate with reduced patient survival is poorly understood. Recent findings provide insight into mechanisms underlying tumor growth in LNs. Tumor cells and their secreted molecules engage stromal, myeloid, and lymphoid cells within primary tumors and in the lymphatic system, decreasing antitumor immunity and promoting tumor growth. Understanding the mechanisms of cancer survival and growth in LNs is key to designing effective therapy for the eradication of LN metastases. In addition, uncovering the implications of LN metastasis for systemic tumor burden will inform treatment decisions. In this review, we discuss the current knowledge of the seeding, growth, and further dissemination of LN metastases.