Although natural killer (NK) cells are thought to give the host a spontaneous resistance against tumours and have been postulated to act in vivo as surveillor cells, definitive data in support of these hypotheses has not been obtained. Recently the beige (bg) mouse, a morphological homologue of the human Chediak-Higashi (CH) syndrome, was shown to be deficient in NK activity. Specifically, spleen cells of bg mice were demonstrated to be incapable of in vitro natural cytotoxicity against tumour cells. We report here that a tumour line, modified to be sensitive to NK cytotoxicity by in vitro culture, demonstrated in vivo an increased growth rate, faster induction time and an increased metastatic capability in bg compared to control mice. This was not found with a tumour line insensitive to NK activity (without in vitro culture). In vivo activation of NK cells in bg and control mice resulted in a decrease in tumour growth rate and metastatic frequency. These results demonstrate that NK cells have an important function in the host's control of tumour growth and metastasis.

Purpose: The aim of this study was to show that novel photodynamic therapy (PDT) sensitizers can be activated by two-photon absorption in the near-IR region of the spectrum and to show, for the first time, that such activation can lead to tumor regressions at significant tissue depth. These experiments also evaluated effects of high-energy femtosecond pulsed laser irradiation on normal tissues and characterized the response of xenograft tumors to our PDT protocols. Experimental Design: Human small cell lung cancer (NCI-H69), non-small cell lung cancer (A549), and breast cancer (MDA-MB-231) xenografts were induced in SCID mice. Irradiation of sensitized tumors was undertaken through the bodies of tumor-bearing mice to give a treatment depth of 2 cm. Posttreatment tumor regressions and histopathology were carried out to determine the nature of the response to these new PDT agents. Microarray expression profiles were conducted to assess the similarity of responses to single and two-photon activated PDT. Results:Regressions of all tumor types tested were seen. Histopathology was consistent with known PDT effects, and no, or minimal, changes were noted in irradiated normal tissues. Cluster analysis of microarray expression profiling showed reproducible changes in transcripts associated with apoptosis, stress, oxygen transport, and gene regulation. Conclusions: These new PDT sensitizers can be used at a depth of 2 cm to produce excellent xenograft regressions. The tumor response was consistent with known responses to singlephoton activated PDT. Experiments in larger animals are warranted to determine the maximal achievable depth of treatment.Photodynamic therapy (PDT; ref. 1) is a protocol that uses light of the appropriate wavelength to activate photosensitizer accumulated in tumor tissue to its triplet state (2). The triplet excitation energy can be effectively transferred to molecular oxygen, resulting in the generation of singlet oxygen, superoxide radical, and other active oxygen species (2, 3). The singlet oxygen causes direct chemical damage to tumor and/or tumor endothelial cells (4), initiates a neutrophilic inflammatory response (5), and can stimulate both innate and specific antitumor immune responses (6 -8). Singlet-oxygen production, however, requires a sufficiently high energy of the triplet state of the sensitizer. Long-wavelength near-IR (NIR) light k ex > 750 nm has relatively low-photon energy, which restricts the range of processes that can be activated by one-photon excitation. The relation between the wavelength and the minimum excitation energy of singlet oxygen is shown in Fig. 1A. The dark gray shaded area corresponds to the excitation energy below the required minimum value, E so < 1.5 -1.6 eV (9, 10), which shows that sensitization by one-photon absorption (1PA) fails in the phototherapeutic window 780 to 950 nm, where tissues have maximum transparency to light. Sensitization by simultaneous two-photon absorption (2PA; ref. 11) combines the energy of two photons and can provide suffi...

The role of host mast cells in tumor-associated angiogenesis was investigated by comparing the angiogenic response of genetically mast-cell-deficient W/Wv mice and mast-cell-sufficient +/+ littermate mice to s.c. growing B16-BL6 tumors. The angiogenic response was found to be slower and initially less intense in W/Wv mice than in +/+ mice. Fewer W/Wv mice than +/+ mice developed spontaneous lung metastases and W/Wv mice exhibited fewer lung metastases per mouse. Bone-marrow repair of the mast-cell deficiency restored the angiogenic response of W/Wv mice and also restored the incidence of hematogenous metastases to approach that of +/+ mice. Differences in lymphatic metastasis were not detected between W/Wv and +/+ mice. These results demonstrate a role for mast cells in vivo during tumor angiogenesis, and suggest a role also for host mast cells in hematogenous metastasis.

The level of expression of the 67 kDa high-affinity laminin binding protein (LBP) correlates with the progression of many solid tumors. The cDNA clone for the 67 kDa LBP is sufficient to encode a polypeptide of only 32 kDa, and there is no readily identifiable mechanism for membrane association. We have overexpressed the transfected 67 kDa hamster LBP in quantities that have enabled us to analyze the membrane-bound form of the protein. Treatment of the purified LBP with methyl transesterification reagents, followed by GC-MS, identified the covalently bound fatty acids palmitate, stearate, and oleate. The fatty acid modification may provide a mechanism for membrane association. Molecular mass determination by MALDI-TOF MS demonstrated the true molecular mass of the protein to be 66.7 kDa, compatible with the SDS-PAGE observation of 67 kDa. Treatment of the LBP with neuraminidase, O-glycanase, or Endo-F glycosidase has no detectable effect on the apparent molecular mass of the protein, and the MALDI-TOF MS did not show evidence of mass heterogeneities typically observed with glycosylated proteins. Reduction with dithiothreitol or beta-mercaptoethanol had no effect on the apparent molecular mass on SDS-PAGE or on the relative quantities of molecular mass species on MALDI-TOF MS. The experimentally determined amino acid composition, however, was found to be consistent with the 67 kDa form being a homodimer of the 32 kDa precursor. Preliminary experiments also suggest that the high-affinity laminin binding characteristic of the protein may be modulated by an, as yet, unidentified membrane accessory molecule.

Six independent lines of evidence point to the existence of heme-containing compounds and͞or hemoglobin breakdown products in extracts of trabecular tissues of the large theropod dinosaur Tyrannosaurus rex. These include signatures from nuclear magnetic resonance and electron spin resonance that indicate the presence of a paramagnetic compound consistent with heme. In addition, UV͞visible spectroscopy and high performance liquid chromatography data are consistent with the Soret absorbance characteristic of this molecule. Resonance Raman profiles are also consistent with a modified heme structure. Finally, when dinosaurian tissues were extracted for protein fragments and were used to immunize rats, the resulting antisera reacted positively with purified avian and mammalian hemoglobins. The most parsimonious explanation of this evidence is the presence of blood-derived hemoglobin compounds preserved in the dinosaurian tissues.