Cancer Stem Cells: New Targets for Cancer Therapy

Interested in receiving more content like this? Join over 8,000 of your peers who have already signed up for the Cell Informer newsletter.


 

The Cancer Stem Cell Hypothesis

Are stem cells involved in cancer? Cancer stem cells (CSC’s) are subpopulations of cancer cells that can self-renew, generate diverse cells in the tumor mass, and sustain tumorigenesis1. Cancer researchers hypothesizes that tumors arises from cancer stem cells that originate as a result of mutational hits on normal stem cells, by the transformation of restricted progenitor cells or even the differentiated cells that acquire self-renewing capacity2. These cancer stem cells drive tumor progression and recurrence after chemotherapy treatments and have recently become targets for cancer stem cell therapies3. Since both normal stem cells and cancer cells possess ability to self-renew, many pathways that are classically associated with cancer are also involved in the regulation of normal stem cell development including Notch, Wnt, Shh and classical pluripotency transcription factor pathways Oct-4, Sox-2 and Nanog4.

An Illustration of the Cancer Stem Cell Hypothesis

An Illustration of the Cancer Stem Cell Hypothesis

 

3D Spheroid Culture of CSCs

Cancer stem cells (CSCs) seek out a niche created within the tumor 3D microenvironment5. A miniscule number of cells within an in vitro 3D tumor spheroid or an in vivo tumor exhibit “stem cell-like” characteristics including 1) slowed proliferation rate 2) self-renewal capabilities and 3) undifferentiated phenotype that can undergo multilineage differentiation. CSCs, which can reconstitute tumors following drug treatment, have been described in both 3D spheroid models and in patients undergoing chemotherapy6. Increased expression of “stemness-related genes” was observed when comparing solid tumor cell lines grown as 3D spheroids to monolayers7. While clearly not definitive proof of the presence of CSCs, these types of results illustrate the tendency for the in vitro 3D microenvironment to create a “stem cell-like” population similar to the in vivo population believed to be responsible for tumor drug resistance.

Protocol Guide: Tumorsphere Formation Protocol

Cancer Stem Cell Media

Traditionally, cancer stem cells have been isolated and grown from cancer cell lines and tumor biopsies in undefined serum containing media using 3D tumorsphere aggregates. However, using undefined culture conditions has led to experimental variability and suboptimal cell growth. 3dGRO™ Spheroid Medium is a new chemically defined and serum-free cancer stem cell media that supports the expansion of CSC’s in 3D tumorsphere cell cultures. The media supports serial passaging of cancer cell lines as tumorspheres leading to an overall enrichment of CSC population overtime.

Features and Benefits

  • Chemically defined, Serum-free formulation.
  • High tumorsphere formation efficiency.
  • Allows extended serial tumorsphere passaging (>10).
  • Proven to enrich CSCs from numerous cancer cell lines.

Cell Lines tested with 3dGRO™ Spheroid Medium

MCF-7, A-431, Panc-1, LNCaP, HT-29, A-549, U-87 MG, HT1080, E006AA

Application Note: Cancer Stem Cell Proliferation in Human Prostate and Breast Cancer Cell Lines Utilizing a New Defined Serum-Free 3D Spheroid Media

3D Tumorsphere formation of MCF7 breast cancer and E006AA prostate cancer cells at Passage 5 cultured in 3dGRO™ Spheroid Medium

Figure 1. 3D Tumorsphere formation of MCF7 breast cancer and E006AA prostate cancer cells at Passage 5 cultured in 3dGRO™ Spheroid Medium.

 

3D Spheroid Formation Plates

Corning® Costar® Ultra-Low attachment multi-well plates are designed to inhibit cell attachment to efficiently form 3D spheroids, including cancer stem cells. The surface is coated with a highly hydrophilic and neutrally charged coating that binds to polystyrene forcing seeded cells into a suspended floating state. The coating is stable, nontoxic, biologically inert and non-degradable.

 

 

Cancer Stem Cell Markers

Small populations of stem-like cells have been isolated from most leukemias as well as from many solid tumors such as brain glioblastomas, medulloblastomas, breast, cervical, colorectal, gastrointestinal, hepatocellular, lung, pancreatic, prostate, and skin carcinomas8. Cancer stem cells display the same cell surface markers as their normal counterparts, but demonstrate uncontrolled proliferation, perhaps due to a reduced responsiveness to negative growth regulators or to the loss of contact inhibition and gap junction intercellular communication (GJIC).
 

Markers Normal Stem Cells Cancer Stem Cells
Oct3/4, Sox2,
Nanog
Maintenance of pluripotency and self-renewal of ES and iPS cells Highly expressed in poorly differentiated tumors of breast cancer, glioblastomas and bladder cancers. Up regulation of transcription factors in stem-like cells
CD133
(Prominin-1)
Marker for hematopoietic stem cells, neural stem cells and somatic stem cells found in many tissues Higher clongenicity, invasiveness and tumorigenesis in human oral squamous cell carcinomas.
CD44 Self-renewal and differentiation Up-regulation in solid tumors and HNSCC cancers
ALDH Stem like properties including colony formation, self-renewal and tumorigenesis Marker for normal and malignant mammary stem cells. Prognostic marker for breast cancer and predictor of metastasis.
Side Populations (Hoechst) Enriched in hematopoietic stem cells Identified in solid tumors such as hepatocellular carcinoma, glioma, ovarian and breast cancers


Cancer Stem Cell Antibodies
 

CD133 CD44 CD24 CD29 CD90 Nestin
SCA-1 BMI-1 ALDH1 OCT3/4


Browse all Cancer Antibodies
Here

 

Cancer Stem Cell ALDH Detection

Cancer stem cells express elevated levels of aldehyde dehydrogenase (ALDH) and is considered a hallmark of all CSCs9. Historically, the ALDEFLUOR™ assay has been used to measure ALDH levels and has been influential to study rare CSCs populations. The ALDEFLUOR assay emits in the green region of the electromagnetic spectrum (512 nm). Thus, the reagent cannot be simultaneously utilized in cells or mice expressing green-fluorescent proteins emitting in the green fluorescent spectrum. AldeRed is a red-shifted fluorescent substrate for ALDH used for labelling viable ALDH positive cells leaving the green emission channel available for further experimentation10. The AldeRed ALDH Detection Assay addresses current limitations of the ALDEFLUOR assay.

Read about AldeRed in Nature

AldeRed Features and Benefits:

  • Red-shifted assay leaves green channel available for further experimentation
  • Live stem cell identification enables flow sorting of rare cell populations
  • Rapid enzymatic assay protocol

AldeRed 588-A

Figure 2. AldeRed 588-A is a fluorescent and non-toxic ALDH substrate that diffuses freely into live intact and viable cells, but remains trapped inside the cells once converted by ALDH into the corresponding acid.

Materials

     

References

  1. Dick JE. Evolution of the cancer stem cell model. Cell Stem Cell. 2014 Mar 6;14(3):275-91.
  2. K. Lenhard Rudolph. Aging-Induced Stem Cell Mutations as Drivers for Disease and Cancer. Cell Stem Cell. 2015 Jun 4; 16(6): 601–612.
  3. Xin Wei. Cancer stem cells and drug resistance: the potential of nanomedicine. Nanomedicine (Lond). 2012 Apr; 7(4): 597–615.
  4. Ailles L. Developmental signaling pathways in cancer stem cells of solid tumors. Biochim Biophys Acta. 2013 Feb;1830(2):2481-95.
  5. Zena Werb. The Cancer Stem Cell Niche: How Essential is the Niche in Regulating Stemness of Tumor Cells? Cell Stem Cell. 2015 Mar 5; 16(3): 225–238.
  6. Domingo-Domenech J. Targeting cancer stem cells to suppress acquired chemotherapy resistance. Oncogene. 2014 Sep 4;33(36):4451-63.
  7. Chang HY. Stemness, cancer and cancer stem cells. Cell Cycle. 2008 Dec;7(23):3622-4. Epub 2008 Dec 2.
  8. Eaves CJ. Cancer stem cells: an evolving concept. Nat Rev Cancer. 2012 Jan 12;12(2):133-43.
  9. Lee PW. Aldehyde dehydrogenase: its role as a cancer stem cell marker comes down to the specific isoform. Cell Cycle. 2011 May 1;10(9):1378-84.
  10. Pomper MG. A red-shifted fluorescent substrate for aldehyde dehydrogenase. Nat Commun. 2014 Apr 23;5:3662.

 

Related Links