Staged Stereotactic Radiosurgery Decreases Symptomatic Radionecrosis in Large Brain Metastasis.

BACKGROUND: Limited brain metastasis is treated definitively with stereotactic radiosurgery when surgical resection is not indicated. Although this has historically been performed in a single fraction, multi-fraction approaches such as fraction radiosurgery (FSRS) and staged radiosurgery (SSRS) have been recently examined as alternative approaches for larger lesions to permit better tumor control without increased toxicity. CASE REPORT: We present the case of a patient who developed symptomatic radionecrosis in two brain metastasis, 2.3 cm and 2.1 cm in size, which were treated with 18 Gy in one fraction, but no radionecrosis in a 3.3 cm lesion treated in two fractions of 15 Gy nor in two punctate lesions that were treated in one fraction of 20 Gy. Although she did not respond to steroids, she responded to bevacizumab symptomatically and on neuroimaging. CONCLUSION: Congruent with other recent studies, our report suggests that large brain metastasis should be considered for FSRS/SSRS.

TTK inhibitor promotes radiosensitivity of liver cancer cells through p21.

The monopolar spindle 1 ((hMps1/TTK) is a serine/threonine kinase that plays an important role in spindle assembly checkpoint signaling. To explore the possible relationship between TTK inhibition and radiosensitivity, we examined whether TTK inhibition influences cellular susceptibility of radiation. And we further revealed its mechanisms. We found that the expression of TTK was obviously higher in liver cancer tissues compared to the normal liver tissues. Kaplan-Meier Plotter demonstrated that patients with low TTK expression levels had a longer overall survival than patients with high TTK expression levels. TTK inhibitor AZ3146 could simulated liver cancer cells to accumulate in the G2/M phase, which ultimately enhances DNA damage with more γ-H2AX foci and more apoptosis and necrosis induced by radiation, which prompted that TTK inhibition sensitized liver cancer cells to radiation. In addition, TTK inhibition altered cell-cycle progression and exacerbated centrosome abnormalities, resulting in enhanced mitotic catastrophe (MC) induced by radiation in a p21-mediated manner. In this study, we present evidences that the TTK inhibitor promotes the radiosensitivity of liver cancer cells through regulating cell cycle in p21-mediated manner in vitro, indicating that TTK inhibitor may be an attractive radiosensitizer for the patients with liver cancer.

Effects of photobiomodulation therapy on the local experimental envenoming by Bothrops leucurus snake.

Bothrops leucurus is the major causative agent of snakebites in Brazil's Northeast. The systemic effects of its venom are effectively neutralized by antivenom therapy, preventing bitten patients' death. However, antivenom fails in neutralizing local effects that include intense pain, edema, bleeding, and myonecrosis. Such effects can lead to irreversible sequels, representing a clinically relevant issue for which there is no current effective treatment. Herein, the effects of photobiomodulation therapy (PBMT) were tested in the local actions induced by B. leucurus venom (BLV) in mice (n = 123 animals in 20 experimental groups). A continuous emission AlGaAs semiconductor diode laser was used in two wavelengths (660 or 780 nm). Mechanical nociceptive thresholds were assessed with the electronic von Frey apparatus. Local edema was determined by measuring the increase in paw thickness. Hemorrhage was quantified by digital measurement of the bleeding area. Myotoxicity was evaluated by serum creatine kinase (CK) activity and histopathological analysis. PBMT promoted anti-hypernociception in BLV-injected mice; irradiation with the 660 nm laser resulted in faster effect onset than the 780 nm laser. Both laser protocols reduced paw edema formation, whether irradiation was performed immediately or half an hour after venom injection. BLV-induced hemorrhage was not altered by PBMT. Laser irradiation delayed, but did not prevent myotoxicity caused by BLV, as shown by a late increase in CK activity and histopathological alterations. PBMT was effective in the control of some of the major local effects of BLV refractory to antivenom. It is a potential complementary therapy that could be used in bothropic envenoming, minimizing the morbidity of these snakebite accidents.

Combination of Trastuzumab Emtansine and Stereotactic Radiosurgery Results in High Rates of Clinically Significant Radionecrosis and Dysregulation of Aquaporin-4.

PURPOSE: Patients with human EGFR2-positive (HER2+) breast cancer have a high incidence of brain metastases, and trastuzumab emtansine (T-DM1) is often employed. Stereotactic radiosurgery (SRS) is frequently utilized, and case series report increased toxicity with combination SRS and T-DM1. We provide an update of our experience of T-DM1 and SRS evaluating risk of clinically significant radionecrosis (CSRN) and propose a mechanism for this toxicity. EXPERIMENTAL DESIGN: Patients with breast cancer who were ≤45 years regardless of HER2 status or had HER2+ disease regardless of age and underwent SRS for brain metastases were included. Rates of CSRN, SRS data, and details of T-DM1 administration were recorded. Proliferation and astrocytic swelling studies were performed to elucidate mechanisms of toxicity. RESULTS: A total of 45 patients were identified; 66.7% were HER2+, and 60.0% were ≤ 45 years old. Of the entire cohort, 10 patients (22.2%) developed CSRN, 9 of whom received T-DM1. CSRN was observed in 39.1% of patients who received T-DM1 versus 4.5% of patients who did not. Receipt of T-DM1 was associated with a 13.5-fold (P = 0.02) increase in CSRN. Mechanistically, T-DM1 targeted reactive astrocytes and increased radiation-induced cytotoxicity and astrocytic swelling via upregulation of Aquaporin-4 (Aqp4). CONCLUSIONS: The strong correlation between development of CSRN after SRS and T-DM1 warrants prospective studies controlling for variations in timing of T-DM1 and radiation dosing to further stratify risk of CSRN and mitigate toxicity. Until such studies are completed, we advise caution in the combination of SRS and T-DM1.

A volume-equivalent spherical necrosis-tumor-normal liver model for estimating absorbed dose in yttrium-90 microsphere therapy.

PURPOSE: Primary hepatocellular carcinoma and metastatic liver tumors are highly malignant tumors in Asia. The incidence of fatal liver cancer is also increasing in the United States. The aim of this study was to establish a spherical tumor model and determine its accuracy in predicting the absorbed dose in yttrium-90 (Y-90) microsphere therapy for liver cancer. METHODS: Liver morphology can be approximated by a spherical model comprising three concentric regions representing necrotic, tumor, and normal liver tissues. The volumes of these three regions represent those in the actual liver. A spherical tumor model was proposed to calculate the absorbed fractions in the spherical tumor, necrotic, and normal tissue regions. The THORplan treatment planning system and Monte Carlo N-particle extended codes were used for this spherical tumor model. Using the volume-equivalent method, a spherical tumor model was created to calculate the total absorbed fraction [under different tumor-to-healthy-liver ratios (TLRs)]. The patient-specific model (THORplan) results were used to verify the spherical tumor model results. RESULTS: The results for both the Y-90 spectrum and the Y-90 mean energy indicated that the absorbed fraction was a function of the tumor radius and mass. The absorbed fraction increased with tumor radius. The total absorbed fractions calculated using the spherical tumor model for necrotic, liver tumor, and normal liver tissues were in good agreement with the THORplan results, with differences of less than 3% for TLRs of 2-5. The results for the effect of TLR indicate that for the same tumor configuration, the total absorbed fraction decreased with increasing TLR; for the same shell tumor thickness and TLR, the total absorbed fraction was approximately constant; and for tumors with the same radius, the total fraction absorbed by the tumor increased with the shell thickness. CONCLUSIONS: The results from spherical tumor models with different tumor-to-healthy-liver ratios were highly consistent with the reference results (THORplan). These findings indicate that a spherical tumor model can provide good estimates of Y-90 doses in microsphere therapy and can be considered a first approximation for dose estimation in Y-90 microsphere therapy.

Radionecrosis versus progresión de la enfermedad en metástasis cerebrales: valor del 18F-DOPA PET/TC/RM / Radionecrosis versus disease progression in brain metastasis. Value of 18F-DOPA PET/CT/MRI

La utilización del 18F-DOPA PET/TC junto a la superposición con imágenes de resonancia magnética y el empleo de métodos de análisis visual y semicuantitativo permitió diferenciar entre las alteraciones posradiocirugía vs. sospecha de progresión de la enfermedad en un paciente con metástasis cerebrales de melanoma, permitiendo tomar una conducta quirúrgica correcta precozmente (AU)

The use of 18F-DOPA PET/CT with magnetic resonance imaging fusion and the use of visual methods and quantitative analysis helps to differentiate between changes post-radiosurgery vs. suspicion of disease progression in a patient with brain metastases from melanoma, thus facilitating taking early surgical action (AU)

Cell viability, reactive oxygen species, apoptosis, and necrosis in myoblast cultures exposed to low-level infrared laser.

Low-level infrared laser is considered safe and effective for treatment of muscle injuries. However, the mechanism involved on beneficial effects of laser therapy are not understood. The aim was to evaluate cell viability, reactive oxygen species, apoptosis, and necrosis in myoblast cultures exposed to low-level infrared laser at therapeutic fluences. C2C12 myoblast cultures at different (2 and 10 %) fetal bovine serum (FBS) concentrations were exposed to low-level infrared laser (808 nm, 100 mW) at different fluences (10, 35, and 70 J/cm(2)) and evaluated after 24, 48, and 72 h. Cell viability was evaluated by WST-1 assay; reactive oxygen species (ROS), apoptosis, and necrosis were evaluated by flow cytometry. Cell viability was decreased atthe lowest FBS concentration. Laser exposure increased the cell viability in myoblast cultures at 2 % FBS after 48 and 72 h, but no significant increase in ROS was observed. Apoptosis was decreased at the higher fluence and necrosis was increased at lower fluence in myoblast cultures after 24 h of laser exposure at 2 % FBS. No laser-induced alterations were obtained at 10 % FBS. Results show that level of reactive oxygen species is not altered, at least to those evaluated in this study, but low-level infrared laser exposure affects cell viability, apoptosis, and necrosis in myoblast cultures depending on laser fluence and physiologic conditions of cells.

Combination of ionising radiation with hyperthermia increases the immunogenic potential of B16-F10 melanoma cells in vitro and in vivo.

Mild hyperthermia (HT) (41.5 °C for 30-60 min) has been shown in various cell culture systems, preclinical and clinical models to be a very potent radiosensitiser. Recent research suggests that local HT application in combination with standard tumour therapies such as radiotherapy (RT) and/or chemotherapy may not only improve local tumour control but also lead to systemic and immune mediated anti-tumour responses. Melanoma has been proven to be rather radioresistant and mostly only the addition of immunotherapy is capable of inducing beneficial anti-melanoma responses. This work therefore focuses on whether HT increases the immunogenic potential of B16-F10 mouse melanoma cells in combination with RT. The in vitro experiments revealed that combination of RT with HT resulted in an increased percentage of apoptotic and necrotic melanoma cells and an increased release of the danger signal heat shock protein 70 (Hsp70) and high mobility group box protein 1 (HMGB1). HT alone was also capable of inducing this release. We set up local irradiation and heating procedures of B16-F10 tumour-bearing C57/BL6 mice and revealed that the tumour growth of tumours treated with RT plus HT was significantly retarded compared to tumours treated only with RT. This combined treatment generated a beneficial tumour microenvironment by enhancing the infiltration of CD11c + /MHCII + /CD86+ dendritic cells, CD8+ T cells, and NK cells, and decreasing that of regulatory T cells and myeloid-derived suppressor cells. We conclude that HT in combination with RT has an immune-stimulating potential that might result in anti-tumour immunity.

Monte Carlo simulations of dose distributions with necrotic tumor targeted radioimmunotherapy.

Radio-resistant hypoxic tumor cells are significant contributors to the locoregional recurrences and distant metastases that mark failure of radiotherapy. Due to restricted tissue oxygenation, chronically hypoxic tumor cells frequently become necrotic and thus there is often an association between chronically hypoxic and necrotic tumor regions. This simulation study is the first in a series to determine the feasibility of hypoxic cell killing after first targeting adjacent areas of necrosis with either an α- or ß-emitting radioimmunoconjugate.

Hyperbaric oxygen for neurologic indications--action plan for multicenter trials in: stroke, traumatic brain injury, radiation encephalopathy & status migrainosus.

The 2008 Toronto Hyperbaric Medicine Symposium was convened to discuss research into neurologic indications for hyperbaric oxygen therapy (HBO2T). Four topics were particularly addressed: acute ischemic stroke; acute traumatic brain injury; brain radiation necrosis; and status migrainosus. Four multicenter trials were designed and proposed to evaluate the efficacy of HBO2T for these indications and are presented here in addition to brief reviews of the rationale behind each.

Immunogenicity of 56 degrees C and UVC-treated prostate cancer is associated with release of HSP70 and HMGB1 from necrotic cells.

BACKGROUND: Prostate hyperthermia and photodynamic therapy can be delivered by a variety of procedures which result in a wide range of temperatures and light energy and cause different kinds of cell death. METHODS: We have addressed the immunogenic effect of heating and UVC irradiation on the prostate cancer (PCa) cell line LNCaP, by studying the release of Danger Associated Molecule Pattern (DAMP) molecules HSP70 and HMGB1 and the dendritic cell (DC) antigen-presenting efficiency. RESULTS: Intracellular upmodulation and extracellular release of HSP70 were inversely correlated. Mild temperatures (43-47 degrees C) induced an early increase of intracellular HSP70, whereas the highest temperature (56 degrees C) induced its extrusion from the cell. Likewise, UVC caused an immediate migration of HSP70 into the cell medium in the absence of any intracellular modulation. 56 degrees C and UVC also induced a robust release of HMGB1. The release of DAMP molecules was closely associated with post-apoptotic membrane damage, as shown by double Annexin V/propidium iodide staining, whereas beta-tubulin, a structural component of cell membranes, was specifically induced by 56 degrees C heating. Tumor uptake strongly impaired the cytokine-driven maturation of DCs and 56 degrees C heating led to a significant recovery of CD83 and CCR7 DC maturation markers, but did not influence the antigen cross-presentation activity. On the contrary, UVC-treated LNCaP had negligible effects on DC maturation, but increased the cross-priming of tumor specific CTL. CONCLUSIONS: These data may be of use in the design of effective non-surgical PCa ablations that combine tumor destruction with long lasting immunity.

Cirugía conservadora de riñón para tumores renales pequeños, rol de la radiofrecuencia / Neprhon-sparing surgery for the treatment of small renal tumors. The role of radiofrequency

OBJETIVO: La radiofrecuencia con asistencia laparoscópica es una opción de tratamiento minimamente invasivo para la conservación de parénquima renal, especialmente en pacientes con comorbilidad aumentada. Se presentan los resultados a corto plazo de los pacientes tratados por esta novedosa técnica.MÉTODOS: Las lesiones renales menores de 4 cm., sospechosas de malignidad o de metástasis a la TC o RNM son candidatas para radiofrecuencia. Bajo visión laparoscópica el tumor es identificado, realizándose una biopsia por punción percutánea. Según tamaño del tumor, se realiza un determinado número de punciones con aguja de radiofrecuencia, con el fin de lograr la necrosis del tumor durante al menos 1 ciclo de radiofrecuencia. El seguimiento es realizado con RNM en el día post operatorio 1 y luego con TC o RNM al mes, 3, 6 y 12 meses. La ausencia persistente de contraste o la necrosis vascular de la lesión es considerada una ablación satisfactoria sin recurrencia.RESULTADOS: Doce pacientes, 2 por enfermedad metastásica y 10 por lesiones primarias (edad promedio 60.8 años), con una o más lesiones sospechosas de malignidad fueron tratadas con radiofrecuencia. El ASA promedio fue de 2,4. El número de tumores tratados fue de 15 con un diámetro de 2,8 cm. Se utilizaron un promedio de 2,5 punciones con aguja de radiofrecuencia. Los resultados de la biopsia fueron metástasis de 1° tiroídeo: 1 paciente, metástasis de melanoma: 1 paciente y cáncer de células renales en 10 pacientes. La estadía hospitalaria promedio fue de 25.8 horas. No existieron complicaciones a corto plazo. Tiempo de seguimiento 8.8 meses. Hasta hoy no hay evidencias de recidiva en los controles imagenológicos(AU)

CONCLUSIONES: La radiofrecuencia es efectiva en erradicar lesiones renales pequeñas, tanto primarias como metastásicas, siendo especialmente útil en pacientes con comorbilidad aumentada. A pesar de que no hay una cantidad suficiente de pacientes con seguimiento adecuado, esta tecnología es prometedora. El abordaje bajo visión laparoscópica contribuye a una biopsia efectiva, evitando diseminación y permite una radiofrecuencia más certera al constatar bajo visión directa la necrosis del tumor(AU)

OBJECTIVES: Laparoscopically assisted radiofrequency is a minimally invasive nephron-sparing treatment option for renal tumors, mainly in patients with high comorbidity. We present the short-term results of our series patients treated with this novel technique.METHODS: Renal lesions smaller than 4 cm, suspicious of malignancy or metastasis on CT scan or MRI are candidates for radiofrequency. Under laparoscopic vision the tumor is identified, and percutaneous biopsy is performed. Depending on the size of the tumor, a number of punctures with the radiofrequency needle are performed with the aim to achieve tumor necrosis during at least one cycle of radiofrequency. Follow-up is performed with MRI in the first postoperative day and then after CT scan or MRI at 1, 3, 6 and 12 months. The persistent absence of contrast or vascular necrosis of the lesion is considered a satisfactory ablation without recurrence.RESULTS: 12 patients, two with metastasis and ten with primary lesions (mean age 60.8 years), with one or more lesions suspicious of malignancy underwent radiofrequency. Mean ASA was 2.4. 15 tumors were treated, with a mean diameter of 2.8 cm. An average of 2.5 punctures was performed with the radiofrequency needle. Biopsy results showed: one case of thyroid cancer metastasis, one case of melanoma metastasis, and 10 cases of renal cell carcinoma. Mean hospital stay was 25.8 hours. There were not short-term complications. Follow-up time was 8.8 months. Today there is no evidence of recurrence in imaging tests.çCONCLUSIONS: Radiofrequency is effective eradicating small renal lesions, both primary and metastatic; it is especially useful in patients with high comorbidity. Despite the number of patients with adequate follow-up is not enough, the technology is promising. The approach under laparoscopic vision contributes to an effective biopsy, avoiding dissemination and enabling a more precise radiofrequency by direct vision control of tumor necrosis(AU)

New photonic technologies for the treatment and diagnosis of hepatic diseases: an overview of the experimental work performed in collaboration, between Physics Institute of São Carlos and Ribeirão Preto Faculty of Medicine of the University of São Paulo.

Recent advances in optical techniques have created a great range of possibilities for diagnosis and therapeutics in liver related diseases. With the uses of efficient light sources like lasers and LEDs (Light Emitting Diodes) it is possible to employ the light-tissue interaction to promote hepatic tissue regeneration after partial hepatectomy, to detect hepatocarcinoma and steatosis by utilizing optical fluorescence, to evaluate the metabolism of the liver during hepatic transplantation as well as to treat liver tumors. We present here an overview of the technique presently in development at the Ribeirâo Preto Faculty of Medicine-USP in cooperation with the Physics Institute of São Carlos-USP. The results obtained so far have been the subject of a list of publications and are here presented as an overview. A new perspective for modern application of optical techniques in different medical practices related to the liver is presented.

Beta-carboline derivatives: novel photosensitizers that intercalate into DNA to cause direct DNA damage in photodynamic therapy.

Novel 1,3,9-trisubstituted beta-carboline derivatives were found to exhibit DNA photocleavage properties under visible light irradiation in a cell-free system, which could be reduced by antioxidant vitamin E. Their photo-cytotoxicity to human tumor cell line HeLa was confirmed, in which apoptosis only contributed a small part to the cell death, and necrosis was the dominating outcome of HeLa cells in photodynamic therapy (PDT) using beta-carboline derivatives. Different from other clinical PDT drugs, beta-carboline derivatives were demonstrated to be able to distribute in the nucleus and intercalate into DNA, and consequently cause direct DNA damage by photochemical reaction products in PDT, which was proved by the distinct DNA tails in the comet assay and the considerable amount of DNA damaged cells quantified by flow cytometry. This mechanism could be the explanation for the delay of cell proliferation at DNA synthesis and mitosis.

Cell death induced by 131 I in a differentiated thyroid carcinoma cell line in vitro: necrosis or apoptosis?

AIM: The apoptotic and necrotic dose-response of thyroid carcinoma cells following irradiation with I was evaluated. METHODS: In our in-vitro model, cells of well-differentiated papillary thyroid carcinoma (B-CPAP) were incubated with increasing activity concentrations of I for 2 days. Changes in cell viability and the extents of necrosis and apoptosis were evaluated both immediately and 2 days after irradiation. RESULTS: Viability of B-CPAP cells diminished with increasing I activity concentration. No apoptosis was detectable immediately after irradiation. Two days after irradiation significant apoptosis was found. The lowest I activity concentration at which apoptosis was detectable corresponds to about 1 MBq . ml. At higher activity concentrations a larger percentage of cells became apoptotic but the proportion decreased again at activity concentrations >10 MBq . ml. Likewise, necrosis was minimal at low activity concentrations and showed an exponential increase with rising I activity concentrations (>5-10 MBq . ml). Necrosis was already detectable immediately after irradiation and was the predominant form of cell death at high activity concentrations. CONCLUSION: The data suggest that the nature of the cytotoxic effect of I and whether it leads to apoptotic or necrotic cell death is dose-dependent. High I doses seem to produce mainly necrotic phenomena, whereas at low I activity concentrations apoptotic phenomena prevail. The predominance of delayed apoptosis could explain why radioiodine therapy at lower doses is often linked to delayed onset and possible continuation of thyroid volume reduction over some months and even up to a year.

New photonic technologies for the treatment and diagnosis of hepatic diseases: an overview of the experimental work performed in collaboration, between Physics Institute of São Carlos and Ribeirão Preto Faculty of Medicine of the University of São Paulo / Novas tecnologias fotônicas no tratamento e diagnóstico de doenças hepáticas: uma revisão dos trabalhos experimentais realizados em colaboração, entre o Instituto de Física de São Carlos e a Faculdade de Medicina de Ribeirão Preto da Universidade de São Paulo

Recent advances in optical techniques have created a great range of possibilities for diagnosis and therapeutics in liver related diseases. With the uses of efficient light sources like lasers and LEDs (Light Emitting Diodes) it is possible to employ the light-tissue interaction to promote hepatic tissue regeneration after partial hepatectomy, to detect hepatocarcinoma and steatosis by utilizing optical fluorescence, to evaluate the metabolism of the liver during hepatic transplantation as well as to treat liver tumors. We present here an overview of the technique presently in development at the Ribeirâo Preto Faculty of Medicine - USP in cooperation with the Physics Institute of São Carlos -USP. The results obtained so far have been the subject of a list of publications and are here presented as an overview. A new perspective for modern application of optical techniques in different medical practices related to the liver is presented.

Recentes avanços em técnicas ópticas têm propiciado vasto campo de possibilidades tanto para o diagnóstico quanto para a terapêutica de doenças hepáticas.Com o uso de eficientes fontes de luz como o laser e Light emitting diodes (LED) é possível utilizar a interação luz-tecido para promover a regeneração hepática após hepatectomias parciais,detectar hepatocarcinoma, esteatose e outras alterações do fígado pelo uso da fluorescência óptica,para avaliar o metabolismo hepático durante o transplante de fígado e na abordagem diagnóstica e terapêutica de alterações hepatocelulares. Os autores apresentam uma ampla revisão de técnicas atualmente em desenvolvimento na Divisão de Gastroenterologia Cirúrgica da Faculdade de Medicina de Ribeirão Preto da Universidade de São Paulo num trabalho cooperativo com o Instituto de Física de São Carlos da USP. Os resultados obtidos até agora têm sido motivo de lista de publicações que são aqui apresentados em forma de revisão. Uma nova perspectiva de moderna aplicação de técnicas ópticas em várias situações clínico-cirúrgicas relacionadas com o fígado é apresentada e amplamente discutida.

Acral necrotizing mycosis fungoides.

A 65-year-old male with long-standing acral "eczema" and patches of mycosis fungoides suddenly developed a transformed, necrotizing ulcerative cutaneous T-cell lymphoma manifesting as necrotic lesions of the toe and lip. The tumor had brown and/or black eschars on the surface and grew multiple opportunistic organisms. Markers CD3, Epstein-Barr virus-encoded small RNA, and T-cell receptor g gene rearrangement were positive, with CD4, 8, 30, 56, and TIA-1 negative. In spite of initial response to radiation, the patient succumbed to infection within 3 months.

Differences in organization of metastatic and nonmetastatic tumors initiated by the same B16 melanoma clone in mature and young mice.

Subcutaneous transplants of mouse B16 melanoma clone G3.26 grow more slowly, and are markedly more metastatic to the lungs, in mature (greater than 12-month-old) mice than in young (2-month-old) mice. Previous studies suggested that tumors in young mice fail to disseminate viable tumor cells into the hematogenous circulation. To determine if changes in intratumor organization might accompany this altered tumor behavior, G3.26 tumors growing in young and mature mice were examined comparatively at progressive sizes relative to the onset of metastatic dissemination in the older mice. Although the degree of necrosis was comparable in both groups of tumors, vascular density, measured morphometrically in histological sections, was significantly lower in tumors from mature mice at a size when dissemination would be occurring. With the onset of reduced vascular density in tumors in mature mice, there was a substantial increase in the proportion of viable tumor cells that was hypoxic, based on radioresistance and incorporation of the hypoxic cell sensitizer, misonidazole. Quiescent tumor cells, identified by flow cytometry, were also more numerous in tumors from mature mice than in tumors from young mice. Although the importance of these differences in tumor organization to enhanced metastatic behavior is unclear, increased intratumor hypoxia might promote generation of metastatic variants. Alternately, dissemination of tumor cells might be facilitated through a reduced and possibly defective vasculature.