Induced Pluripotent Stem Cells from Bone Cancer Patients Provide Crucial Insights into the Genesis of Bone Cancer

A team of Mount Sinai researchers have utilized induced pluripotent stem cells (iPSCs) to elucidate the genetic changes that seem to convert a well-known anti-cancer signaling gene into a driver bone cancers. When it comes to bone cancers, the survival rate has not improved in 40 years despite advances in treatment. Since this study might provide new targets and suggest new strategies for attacking such cancers. it represents a welcome addition to the cancer literature.

This study, which was published in the journal Cell, revolves around iPSCs, which were discovered in 2006 by Nobel laureate Shinya Yamanaka. iPSCs use genetic engineering and cell culture techniques to reprogram mature, adult cells to become like embryonic stem cells. These iPSCs are “pluripotent,” which means that they are able to differentiate into any adult cell type and can also divide in culture indefinitely.

For therapeutic purposes, iPSCs can be derived from a patient’s own cells, differentiated into the cells the patient needs to be replaced, and then implanted into the patient’s body to augment tissue healing or even organ reconstruction. Since iPSCs can be successfully differentiated into heart muscle, nerve cells, bone, and other cell types, they have the potential advance the field of regenerative medicine by leaps and bounds.

iPSCs have already made their presence known in the clinic by serving as model systems for research and diagnosis. The new Mount Sinai study used iPSCs to construct an accurate model of a genetic disease “in a dish.” The culture dishes contain self-renewing patient-specific iPSCs or a specific cell line that enable in-depth study diseases that are driven by each person’s genetic differences. When matched with patient records, iPSCs and iPSC-derived target cells have the ability to help physicians predict a patient’s prognosis and whether or not a given drug will be effective for him or her.

In this study, skin cells from healthy patients and patients with a genetic disease called Li-Fraumeni syndrome were isolated and reprogrammed into patient-specific iPSC lines. These iPSCs were then differentiated into bone-making cells (osteoblasts), which are the cells where particular rare and common bone cancers start. Li-Fraumeni syndrome greatly predisposes patients to a variety of cancers in several different types of tissues.

The patient-derived osteoblasts were then tested for their tendencies to become tumor cells and to make bone. This particular bone cancer model did a better job of recapitulating the characteristics of bone cancer than previously used mouse or cellular models.

LFS iPSCs for stem cell production

“Our study is among the first to use induced pluripotent stem cells as the foundation of a model for cancer,” said lead author Dung-Fang Lee, PhD, a postdoctoral fellow in the Department of Developmental and Regenerative Biology, Icahn School of Medicine at Mount Sinai. “This model, when combined with a rare genetic disease, revealed for the first time how a protein known to prevent tumor growth in most cases, p53, may instead drive bone cancer when genetic changes cause too much of it to be made in the wrong place.”

The Mount Sinai disease model research uses a simple fact of human life as its basis: human genes undergo mutations at a certain rate that tends to increase as we age, and the formation of new mutations in relentless and constant. Some mutations make no difference, a few some confer advantages, and others cause disease. Beyond inherited mutations that contribute to cancer risk, the combination of random, accumulated DNA changes in our cells as we age also increase our cancer risk.

The current study focused on those genetic pathways involved in Li-Fraumeni Syndrome or LFS, a rare genetic disease that causes high risk for many cancers in affected families. Osteosarcoma (bone cancer) is a common cancer observed in LFS patients and many of them are diagnosed before the age of 30. Additionally, osteosarcoma is the most common type of bone cancer in all children, and after leukemia, the second leading cause of cancer death for them.


Importantly, about 70 percent of LFS families have a mutation in their copy of a genes called TP53, which encodes the p53 protein. P53 is a “the tumor suppressor,” which means that it functions to preserve the integrity of the genome and keep the rate of cell division in check. Common forms of osteosarcoma, which are driven by somatic or inherited mutations, have also been closely linked by past studies to defects in p53 when mutations interfere with the ability of the protein to function properly.

Crystal Structure of p53 protein bound to DNA


Rare genetic diseases like LFS provide excellent model systems because they tend to result from a change in a single gene, instead of the diverse and overlapping mutations observed in common diseases, and, in this case, more common, non-inherited bone cancers. The LFS-iPSC based modeling highlights the contribution of p53 alone to osteosarcoma.

By analyzing iPSC lines, and bone cancer driven by p53 mutations in LFS patients, the Munt Sinai research team showed, for the first time, that the LFS bone cancer results from an overactive p53 gene. Too much p53 in osteoblasts dampens the function of a gene, H19, and a related protein, decorin, that would otherwise help stem cells differentiate into normal osteoblasts.

The inability of cells to differentiate makes them vulnerable to genetic mistakes that drive cancer, since more “stemness” means a tendency toward rapid, abnormal growth, like that observed in tumors. One tragic feature of osteosarcoma is the rapid, error-prone production of weaker bone by cancerous bone-making cells, where a young person surprisingly breaks a bone to reveal undiagnosed, advanced cancer.

Dung-Fang Lee and his colleagues discovered that the H19 gene seems to control a network of interconnected genes that fine-tune the balance between cell growth and resistance to growth. Decorin is a protein that is part of connective tissue like bone, but that also plays a signaling role, interacting with growth factors to slow the rate that cells divide and multiply, unless turned off by too much p53.

“Our experiments showed that restoring H19 expression hindered by too much p53 restored “protective differentiation” of osteoblasts to counter events of tumor growth early on in bone cancer,” said co-author, Ihor Lemischka, PhD, Director of The Black Family Stem Cell Institute within the Icahn School of Medicine. “The work has implications for the future treatment or prevention of LFS-associated osteosarcoma, and possibly for all forms of bone cancer driven by p53 mutations, with H19 and p53 established now as potential targets for future drugs.”

Genomic Imprinting Maintains A Reserve Pool of Blood-Forming Stem Cells

Hematopoietic stem cells or HSCs reside in the bone marrow and give rise to the wide variety of specialized blood cells that inhabit our bloodstreams. Within the bone marrow, HSCs come in two varieties: an active arm of HSCs that proliferate continually to replace our blood cells and a reserve arm that sits and quietly waits for their time to come.

New research from the Stowers Institute at Kansas City, Mo, in particular a research team led by Linheng Li, discovered a mechanism that helps maintain the balance between those HSCs kept in reserve and those on active duty.

According to Dr. Li, genomic imprinting, a process that specifically shuts off one of the two gene copies found in each mammalian cell , prevents the HSCs held in reserve from being switched to active duty prematurely.

Li explained: “Active HSCs form the daily supply line that continually replenishes worn-out blood and immune cells while the reserve pool serves as a backup system that replaces damaged active HSCs and steps in during times of increased need. In order to maintain a long-term strategic reserve of hematopoietic stem cells that lasts a lifetime it is very important to ensure that the back-up crew isn’t mobilized all at once. Genomic imprinting provides an additional layer of regulation that does just that.”

Sexual reproduction produces progeny that have once set of chromosomes from the mother and one set of chromosomes from the father. The vast majority of genes are expressed from both sets of chromosomes. However, in placental mammals and marsupial mammals a small subset of genes are imprinted, which means that they receive a mark during the development of eggs and sperm and these marks shut down expression of those genes in either the sperm pronucleus or the egg pronucleus. Therefore, after the fusion of the sperm and the egg and the eventual fusion of the egg and sperm pronuclei, these imprinted genes are only expressed from one copy of genes. Some are only expressed from the paternal chromosomes and others are only expressed from the maternal chromosome. Imprinting is essential for normal development in mammals.

The importance of genetic imprinting is shown if an egg loses its pronucleus and is then fertilized by two sperms. The resulting zygote has two copies of paternal chromosomes and no copies of the maternal chromosomes. Such an embryo is called an andogenote, and the embryo fails to form but the placenta overgrows. If this occurs during human development, it can lead to a so-called “molar pregnancy” or “hydatiform mole.” This fast growing placental tissue can become cancerous and lead to uterine cancer. For that reason, molar pregnancies are usually dealt with expeditiously.

However, if the sperm that fertilizes the egg is devoid of a pronucleus, and the egg pronucleus duplicates, then the resulting zygotes can two copies of the maternal chromosomes, and this entity is known as a gynogenote, and it develops with a poorly formed placenta that dies early in development.

In previous experiments in mice, Li and his colleagues indicated that the expression of several imprinted genes changes as HSCs transition from quiescent reserve cells to multi-lineage progenitor cells.

In their current study, Li and other Stowers Institute researchers examined a differentially imprinted control region, which drives the reciprocal expression of a gene called H19 from the maternal chromosome and IGF2 (insulin-like growth factor-2) from the paternal chromosome.

The first author of this study, Aparna Venkatraman developed a mouse model that allowed her to specifically delete the imprinted copy from the maternal chromosome. Thus, in these mice, H19, which restricts growth, was no longer active and Igf2,, which promotes cell division, was now active from the paternal and the maternal chromosome. To access the effect of this loss of imprinting on the maintenance of HSCs, Venkatraman examined the numbers of quiescent HSCs and active HSCs. in mouse bone marrow.

Venkatraman explained: “A large number of quiescent HSCs was activated simultaneously when the epigenetic control provided by genomic imprinting was removed. It created a wave of activated stem cells that moved through different maturation stages.”

She followed this experiment with a closer look at the Igf2 gene. Misregulation of Igf2 leads to overgrowth syndromes such as Beckwith-Wiedmann Syndrome. It exerts its growth promoting effects through the Igf1 receptor, which induces an intracellular signaling cascade that stimulates cell proliferation.

IGF signaling pathway
IGF signaling pathway

The expression of the Igf1 receptor itself is regulated by H19, which encodes a regulatory microRNA (miR-675) that represses translation of the Igf1 receptor gene and therefore prevents production of Igf1 receptor protein. Venkatraman explained that once the “imprinting block is lifted, the Igf2-Igf1r signaling pathway is activated.” Venkatraman continued: “The resulting growth signal triggers the inappropriate activation and proliferation of quiescent HSCs, which eventually leads to the premature exhaustion of the reserve [HSC] pool.”

Interestingly, the roundworm, Caenorhabditis elegans, provided the first clues that diminished insulin/IGF signaling can increase lifespan and delay aging. Li again: “Here the IGF pathway is conserved by subject to imprinting, which inhibits its activation in quiescent reserve stem cells. This ensures the long-term maintenance of the blood system, which in turn supports the longevity of the host.”