Protein in Embryonic Stem Cells Controls Malignant Tumor Cells

A protein that governs development of human embryonic stem cells (hESCs) also inhibits the growth and spread of malignant melanoma, the deadliest skin cancer, Northwestern University researchers have discovered. Metastatic melanoma, which develops from the transformation of skin pigment cells or melanocytes, has a death rate of more than 80 percent and a median survival of less than 7.5 months.

The Northwestern scientists, led by researcher Mary J. C. Hendrix, additionally found that the protein, called Lefty, prevents aggressive breast cancer cells from metastasizing. Death from metastatic breast cancer exceeded 40,000 in 2007, with over 180,000 new cases diagnosed in the United States.

Importantly, Lefty is secreted only in hESCs, and not in any other stem cell type tested – including stem cells isolated from amniotic fluid, cord blood or adult bone marrow – or placental cells.

Results of the study, described in an article in the March 3rd online version of The Proceedings of the National Academy of Sciences, build on an elegant body of research by the Hendrix lab to identify the genes and cellular pathways involved in cancer metastasis.

Hendrix is president and scientific director of the Children’s Memorial Research Center and professor in The Robert H. Lurie Comprehensive Cancer Center of Northwestern University and at Northwestern's Feinberg School of Medicine. Lynne-Marie Postovit, who was first author on the study and a post-doctoral trainee in the Hendrix lab, is currently an assistant professor at the University of Western Ontario, Canada.

Embryonic stem cells are pluripotent, meaning they can become any of 200-plus cell types in the adult body, depending on the signals they receive from their microenvironment (surrounding cells, tissues and vasculature). During cancer progression, malignant cells also receive and release signals from their microenvironment, cues that promote tumor growth and metastasis.

Groundbreaking work by Hendrix and colleagues is elucidating how, by becoming more like unspecialized stem cells, aggressive melanoma cells gain enhanced abilities to migrate, invade and metastasize while remaining virtually undetected by the immune system.

Hendrix and co-researchers previously demonstrated that a three-dimensional matrix conditioned by hESCs induced metastatic melanoma cells to revert to a normal, skin cell-like type with the ability to form colonies in the manner of hESCs (Postovit and Seftor et al, Stem Cells 24:501-505, 2006).

“This observation allowed us to appreciate the powerful influence of the hESC microenvironment on the reprogramming of metastatic melanoma cells,” Hendrix said.

In subsequent experiments, Hendrix, Postovit and co-researchers found that aggressive melanoma and breast cancer produce a “morphogenic” protein called Nodal, which is essential for human embryonic stem cell pluripotency (Topczewska et al, Nature Medicine 12:925-932, 2006). Other researchers have found that Nodal also is present in testicular cancer.

“Thus, Nodal may serve as a prognostic marker of aggressive behaviors in human cancers,” Hendrix said.

As described in the PNAS study, the Lefty protein inhibits production of Nodal and therefore plays a major role in embryonic cell differentiation and development – under normal circumstances.

Hendrix and colleagues discovered that metastatic tumor cells do not express Lefty, allowing them to overproduce Nodal in an unregulated manner.

However, when the group exposed metastatic tumor cells to the microenvironment of hESCs containing Lefty, they witnessed dramatically reduced Nodal expression (production) in these cancer cells together with decreased tumor cell growth and invasiveness and an increase in apoptosis, or programmed cell suicide.

Although exposure to a hESC microenvironment inhibited Nodal expression and tumor growth in both metastatic melanoma and breast cancer cells, the breast cancer cells underwent more complex reprogramming. Melanoma cells responded to the hESC-derived factors within three days, but breast cancer cells required two additional days to achieve the most significant reduction in Nodal.

This discrepancy is likely due to differences in signaling mechanisms between the two cell types. Yet, despite the inherent differences between melanoma cells and breast cancer cells, these divergent tumor types both underwent cell suicide following exposure to the hESC microenvironment.

“The remarkable similarity of the responses of the two tumor types is likely attributable to the commonality of plasticity (for example, the aberrant and unregulated expression of Nodal) that indiscriminately unifies highly aggressive cancer cells, regardless of their tissue of origin,” Hendrix said.

“Further, the tumor suppressive effects of the hECs microenvironment, by neutralizing the expression of Nodal in aggressive tumor cells, provide previously unexplored novel therapeutic modalities for cancer treatment,” Hendrix said.

However, while findings from the study suggest that hESC-derived Lefty may have potential to prevent metastasis, it is not the only tumor suppressive factor within the embryonic microenvironment.

Observations from the study highlight the potential utility of isolating factors within the hESC microenvironment responsible for influencing tumor cell fate and reversing the cancerous properties of metastatic tumor cells, such as melanoma and breast cancer.

Additional contributing authors on the study include N. Margaryan; E. Seftor; D. Kirschmann; D. Abbott; W. Wheaton; A. Lipavsky; and R. Seftor.

This study was supported by grants from the Illinois Regenerative Medicine Institute; the National Institutes of Health (CA50702 and CA121205); the Charlotte Geyer Foundation (to Dr. Hendrix); and a Canadian Institutes of Health Research Postdoctoral Fellowship (to Dr. Postovit).
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MIT researchers demonstrate protective role of microRNA

Snippets of genetic material that have been linked to cancer also play a critical role in normal embryonic development in mice, according to a new paper from MIT cancer biologists.

The work, reported in the March 7 issue of Cell, shows that a family of microRNAs--short strands of genetic material--protect mouse cells during development and allow them to grow normally. But that protective role could backfire: The researchers theorize that when these microRNAs become overactive, they can help keep alive cancer cells that should otherwise die--providing another reason to target microRNAs as a treatment for cancer.

Discovered only a decade ago, microRNAs bind to messenger RNAs (mRNAs), preventing them from delivering protein assembly instructions, thereby inhibiting gene expression. The details of how microRNAs act are not yet fully understood.

"The scientific community is busy trying to understand what specific biological functions these microRNAs affect," said Andrea Ventura, lead author of the paper and postdoctoral associate in the Koch Institute for Integrative Cancer Research at MIT (formerly known as the Center for Cancer Research).

Ventura--who works in the laboratory of Tyler Jacks, director of the Koch Institute--and her colleagues studied the function of a family of microRNAs known as the miR-17~92 cluster.

Previous research has shown that the miR-17~92 cluster is overactive in some cancers, especially those of the lungs and B cells.

To better understand these microRNAs' role in cancer, the researchers decided to study their normal function. Knocking out microRNA genes and observing the effects can offer clues into how microRNA helps promote cancer when overexpressed.

They found that when miR-17~92 was knocked out in mice, the animals died soon after birth, apparently because their lungs were too small. Also, their B cells, a type of immune cell, died in an early stage of cell development.

This suggests that miR-17~92 is critical to the normal development of lung cells and B cells. In B cells, these microRNAs are likely acting to promote cell survival by suppressing a gene that induces cell death, said Ventura.

"Understanding why these things are happening provides important insight into how microRNAs affect tumorigenesis," he said.

The researchers theorize that when miR-17~92 becomes overactive in cancer cells, it allows cells that should undergo programmed cell death to survive.

Blocking microRNAs that have become overactive holds promise as a potential cancer treatment. Research is now being done on molecules that prevent microRNAs from binding to their target mRNA.

More work needs to be done to make these inhibitors into stable and deliverable drugs, but Ventura said it's possible it could be done in the near future.

The exact genes targeted by miR-17~92 are not known, but one strong suspect is a gene called Bim, which promotes cell death. However, a single microRNA can have many targets, so it's likely there are other genes involved.

The researchers also studied the effects of knocking out two other microRNA clusters that are closely related to miR-17~92 but located elsewhere in the genome.

They found that if the other two microRNA clusters are knocked out but miR-17~92 remains intact, the mice develop normally. However, if miR-17~92 and one of these similar clusters are removed, the mice die before birth, suggesting there is some kind of synergistic effect between these microRNA families.

Other MIT authors of the paper are Amanda Young, graduate student in biology; Monte Winslow, postdoctoral fellow in the Center for Cancer Research (CCR); Laura Lintault, staff affiliate in the CCR; Alex Meissner, faculty member at the Broad Institute of MIT and Harvard; Jamie Newman, graduate student in biology; Denise Crowley, staff affiliate at the CCR; Rudolf Jaenisch, professor of biology and member of the Whitehead Institute for Biomedical Research; Phillip Sharp, MIT Institute Professor; and Jacks, who is also a professor of biology.

The research was funded by the National Institutes of Health and the National Cancer Institute.

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