Summary

We are interested in how growth factors, signaling pathways, and gene expression programs control normal cell proliferation and cancer cell metastasis. The development and maintenance of multicellular organisms requires tight control over the proliferation, differentiation, movement, organization and death of their constituent cells. Intricate molecular communication networks have evolved to control these processes. Our work is focusing on how cells receive, read and relay such signals, and how disruptions in these processes lead to tumor formation and cancer metastasis. We are approaching these questions through the rich venue provided by the transforming growth factor- (TGFb) pathway, as well as through the direct identification of metastasis genes and functions.

Elucidating signaling TGFb networks

With nearly forty related members in the human genome, TGFb represents one of the most prominent and functionally versatile families of cytokines. This family -the TGFbs, nodals, activins, bone morphogenetic proteins (BMPs), myostatins, and anti-Muellerian hormone- exert profound effects on every aspect of cell behavior. Produced by many cell types or restricted to just a few, different TGFb family members guide early stages of embryo development and tissue homeostasis throughout life. We uncovered key steps in this signaling pathway by identifying the TGFb and BMP receptors and elucidating their mechanism of activation (Figure 1 -- see Figure 1 and all other figures in the Images subsection found in the blue navigation box above). These receptors are serine/threonine kinases and their substrates are the Smad transcription factors (Figure 2). We showed that Smad C-terminal phosphorylation by TGFb and BMP receptor kinases, and Smad linker phosphorylation by MAPK, GSK3 and other protein kinases, are key events in Smad regulation (Figure 3). We are identifying Smad interactions with transcription cofactors and ubiquitin ligases that are supported by these phosphorylation events. These findings are paving the way for new discoveries on the function and network integration of the TGFb and BMP pathways.

Discerning how cells read TGFb signals

The cellular response to TGFb depends on the cell type and the context. We propose that activated Smad factors regulate different genes in different ways in different cell types by associating with diverse DNA binding cofactors, whose availability depends on the cell type. These interactions generate distinct repertoires of Smad transcriptional complexes, each complex targeting a small set of genes for either activation or repression.  With this in mind, we recently identified FoxO, C/EBP, E2F4/5 and ATF3 as the Smad cofactors that mediate, respectively, the induction of p15INK4 and p21CIP1 and the repression of c-MYC and ID1(Figure 4). These gene responses underlie one of the most important effects of TGFb, namely, growth inhibition (Figure 5).

From cytostasis to metastasis

TGF is the most prominent cytokine inhibitor of cell proliferation. This effect involves the repression of cell proliferation genes such as c-MYC and ID1, and the induction of cyclin-dependent kinase inhibitors, two of which -p27Kip1 and p57Kip2- we co-discovered. Besides providing a coherent program for cell cycle inhibition, these transcriptional mechanisms are allowing us to identify alterations that are responsible for the evasion of the TGF cytostatic action in breast cancer and brain tumors (Figure 5). Moreover, once relieved from the growth inhibitory action, tumor cells may use TGFb to escape immune surveillance. We recently found that tumor-derived TGFb inhibits the expression of five tumor cell-killing genes in cytotoxic T lympocytes. And, not content with loosing TGFb growth inhibitory responses, tumor cells may alter the Smad pathway to undertake metastasis to the bones and lungs in breast cancer patients (Figure 6). We are currently identifying the TGFb target genes and mechanisms that mediate metastasis to these organs. In sum, as tumors develop they turn TGFb from a tumor suppressor signal into an accomplice in metastasis.

Identifying metastasis genes and functions

Metastasis is the cause of 90% of deaths from cancer, and yet little is know about its underlying mechanisms. We are addressing this problem by exploiting the fact that tumors have distinct patterns of organ-specific colonization, each with a distinct biology and clinical evolution. We are identifying clinically relevant genes that mediate tumor microenvironment interactions, cancer cell entry and exit from the circulation, and cancer stem cell colonization of various organs. By combining in vivo selection of human metastatic cells, transcriptomic profiling and functional testing, we have identified genes that selectively mediate breast cancer metastasis to bones or lungs (Figure 7). Some of these genes serve dual functions, providing growth advantages both in the primary tumor and in the metastasis microenvironment.  Others contribute to aggressive growth selectively in a particular organ (Figure 8). Bioinformatics analysis of hundreds of primary tumors has resulted in a lung metastasis gene expression signature (LMS) that predicts clinical outcome in patients with ER-negative breast cancer. We are investigating the role of the LMS in other tumors that metastasize to the lungs.

Deconstructing metastasis for clinical intervention

Using genetic and pharmacological approaches, we recently found that several LMS genes, including the epidermal growth factor receptor ligand epiregulin, the cyclooxygenase COX2, and the matrix metalloproteinases 1 and 2, collectively facilitate the assembly of new tumor blood vessels, the release of tumor cells into the circulation, and the breaching of lung capillaries by circulating tumour cells to seed pulmonary metastasis (Figure 9). These findings reveal how aggressive primary tumorigenic functions can be mechanistically coupled to greater lung metastatic potential, and how such biological activities may be therapeutically targeted with specific drug combinations.