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Loss of p53 function can lead to a variety of cancers, including breast cancer. Mice heterozygous for the p53 gene (designated Trp53+/-) develop spontaneous mammary tumors, but this depends on the strain background and has been linked to a locus on chromosome 7 (designated SuprMam1). Mammary tumors are common in BALB/c-Trp53+/-females, but are rare in C57BL/6-Trp53+/- mice. Prevalence of genomic instability appears to contribute to the phenotype as loss of heterozygosity (LOH) is significantly more common among tumors arising in BALB/c-Trp53+/- mice compared to C57BL/6J-Trp53+/- mice. This increased LOH in BALB/c-Trp53+/- tumors was shown to be due to recombination events. The BALB/c strain has been shown to have a deficiency in non-homologous end joining (NHEJ) of DNA double strand breaks (dsb), however, this does not account for the increase of LOH events in tumors. Our hypothesis was that BALB/c-Trp53+/- mice are more susceptible to mammary tumors due to impaired Homologous Recombination Repair (HRR) leading to LOH. Using the COMET assay, we demonstrate that dsbs persist longer in BALB/c-Trp53+/- mouse embryonic fibroblasts (MEFs) compared to C57BL/6J-Trp53+/- MEFs. Similarly, co-localization of H2AX and the homologous recombination protein RAD51 remain at dsbs longer in BALB/c-Trp53+/- MEFs compared to C57BL/6-Trp53+/- MEFs. Palb2, a gene that lies within the SuprMam1 interval and has been shown to contribute to heritable breast cancer, was chosen as an initial candidate gene. No coding SNPs or expression differences of Palb2 were found in the mammary glands between the two strains. Additional fine mapping and use of a filtering criteria in the SuprMam1 region yielded an additional 34 candidate genes. We demonstrate no significant differences in any of these genes in whole mammary glands and primary mammary epithelial cells between the two strains. Finally, using a congenic mouse strain, we demonstrate the lack of irradiation (IR) sensitivity alleles within the SuprMam1 region. These results suggest a possible defect in HRR in the BALB/c strain that is unlikely related to Palb2. The gene or genes respsonsible for increased mammary tumor incidence in the BALB/c-Trp53+/- remain to be identified. vii

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<p>Growth factors and their receptors are frequently activated by mutations in human cancer. Platelet-derived growth factor (PDGF)-B and its tyrosine kinase receptor, the PDGF β-receptor, have been implicated in autocrine transformation as well as paracrine stimulation of tumor growth. The availability of clinically useful antagonists motivates evaluation of PDGF inhibition in these diseases.</p><p>In chronic myelomonocytic leukemia with t(5;12), parts of the transcription factor TEL and the PDGF β-receptor are fused, generating a constitutively signaling protein. Oligomerization and unique phosphorylation pattern of TEL-PDGFβR was demonstrated, as well as the transforming activity of TEL-PDGFβR, which was sensitive to PDGF β-receptor kinase inhibition.</p><p>Dermatofibrosarcoma protuberans (DFSP) is characterized by a translocation involving the collagen Iα1 and PDGF B-chain genes. The COLIA1-PDGFB fusion protein was processed to mature PDGF-BB and transformed fibroblasts in culture. The PDGF antagonist STI571 inhibited growth of <i>COLIA1-PDGFB</i> transfected cells and primary DFSP cells <i>in vitro</i> and <i>in vivo</i> through induction of apoptosis.</p><p>Paracrine effects of PDGF-DD, a ligand for the PDGF β-receptor, were evaluated in a murine model of malignant melanoma. PDGF-DD production accelerated tumor growth and altered the vascular morphology in experimental melanomas.</p><p>A validated immunohistochemical procedure for PDGF β-receptor detection was established and applied to normal tissues and more than 280 tumor biopsies. Perivascular and stromal expression was detected in 90% and 50%, respectively, of human tumors.</p><p>Recently, non-transformed cells in the tumor microenvironment have emerged as targets in cancer therapy. Selective sensitization of tumor fibroblasts to paclitaxel by STI571 was evaluated <i>in vitro</i> and in a xenograft model. Whereas neither drug alone caused growth inhibition, combination of the two significantly reduced tumor growth, suggesting anti-stromal therapy as a possible treatment modality in solid tumors.</p>

<p>Transforming growth factor β (TGFβ) is a multifunctional cytokine that is involved in many biological effects, such as proliferation, migration, differentiation and cell survival. TGFβ regulates cellular responses by binding to a heteromeric complex of type I and type II serine/threonine kinase receptors. The type I receptor, termed activin receptor-like kinase (ALK), acts downstream of the type II receptor and propagates the signal to the nucleus by phosphorylating receptor regulated Smads (R-Smads). The activated R-Smads can associate with the common partner Smad, Smad4, and this complex translocates to the nucleus where it participates in transcriptional regulation of target genes. TGFβ plays an important role in vascular morphogenesis. The aim of this study was to obtain more insight into the mechanisms by which TGFβ can act as an inhibitor or stimulator of angiogenesis Our findings show that in endothelial cells (ECs), TGFβ can activate two distinct type I receptor/Smad signalling pathways with opposite cellular responses. In most cell types, TGFβ signals via the TGFβ type I receptor, ALK5. However, ECs express a predominant endothelial type I receptor, named ALK1. Whereas the TGFβ/ALK1 signalling leads to activation, the TGFβ/ALK5 pathway results in an inhibition of the activation state. This suggests that TGFβ regulates the activation state of the endothelium via a fine balance between these two pathways. We identified genes that are specifically induced by TGFβ mediated ALK1 or ALK5 activation. Id1 was found to be the target gene of the ALK1/Smad1/5 pathway while induction of plasminogen activator inhibitor-1 was activated only by ALK5/Smad2 pathway. Furthermore, ALK1 activated ECs are highly invasive but this property is lost if Id1 expression is specifically knocked-down. ECs invasiveness is highly dependent on αv integrin binding to its extracellular matrix (ECM) protein partner and the invasion requires proteolytic cleavage of the ECM by metalloproteases (MMPs). Hence, TGFβ/ALK1/Id1 pathway may promote invasion by modulating the expression or activity of integrins and MMPs that are well known components of the ECM. Timing and duration of TGFβ signalling are important specificity determinants for its effect on cellular behaviour. After binding to ALK1, TGFβ induces a transient phosphorylation of Smad1/5 but a stable phosphorylation of Smad2 via ALK5. Our studies indicate that Smad7 is potently induced by ALK1 signalling and may recruit a PP1α/TIMAP phosphatase complex to ALK1 to dephosphorylate the receptor and thereby turning off phosphorylation resulting in a temporal activation of TGFβ/ALK1-induced Smad1/5 pathway. This mechanism enables an efficient and tightly temporally controlled activation resulting in the dominance of ALK5 upon prolonged exposure to TGFβ. Bone morphogenetic protein (BMP) is a member of the TGFβ superfamily and signals through Smad1/5. The BMP/Smad1/5 pathway was found to potently activate the endothelium. Id1 was identified as an important BMP target gene in ECs and was sufficient and necessary for BMP-induced EC migration. These studies not only provide new insights into possible molecular mechanisms that underlie activation and quiescence of ECs during physiological angiogenesis but may also explain the vascular phenotypes observed in mice and humans with perturbed TGFβ signalling pathways.</p>

Growth factors and their receptors are frequently activated by mutations in human cancer. Platelet-derived growth factor (PDGF)-B and its tyrosine kinase receptor, the PDGF β-receptor, have been implicated in autocrine transformation as well as paracrine stimulation of tumor growth. The availability of clinically useful antagonists motivates evaluation of PDGF inhibition in these diseases. In chronic myelomonocytic leukemia with t(5;12), parts of the transcription factor TEL and the PDGF β-receptor are fused, generating a constitutively signaling protein. Oligomerization and unique phosphorylation pattern of TEL-PDGFβR was demonstrated, as well as the transforming activity of TEL-PDGFβR, which was sensitive to PDGF β-receptor kinase inhibition. Dermatofibrosarcoma protuberans (DFSP) is characterized by a translocation involving the collagen Iα1 and PDGF B-chain genes. The COLIA1-PDGFB fusion protein was processed to mature PDGF-BB and transformed fibroblasts in culture. The PDGF antagonist STI571 inhibited growth of COLIA1-PDGFB transfected cells and primary DFSP cells in vitro and in vivo through induction of apoptosis. Paracrine effects of PDGF-DD, a ligand for the PDGF β-receptor, were evaluated in a murine model of malignant melanoma. PDGF-DD production accelerated tumor growth and altered the vascular morphology in experimental melanomas. A validated immunohistochemical procedure for PDGF β-receptor detection was established and applied to normal tissues and more than 280 tumor biopsies. Perivascular and stromal expression was detected in 90% and 50%, respectively, of human tumors. Recently, non-transformed cells in the tumor microenvironment have emerged as targets in cancer therapy. Selective sensitization of tumor fibroblasts to paclitaxel by STI571 was evaluated in vitro and in a xenograft model. Whereas neither drug alone caused growth inhibition, combination of the two significantly reduced tumor growth, suggesting anti-stromal therapy as a possible treatment modality in solid tumors.