Rapid generation of in vitro multicellular spheroids for the study of monoclonal antibody therapy

Our plain-English summary of Rapid generation of in vitro multicellular spheroids for the study of monoclonal antibody therapy, by Y.T. Phung, D. Barbone, V.C. Broaddus, et al (Journal of Cancer, 2011).

The study

• Authors: Y.T. Phung, D. Barbone, V.C. Broaddus, M. Ho
• Journal: Journal of Cancer
• Published: 2011;2:507-14
• Cited by: 69 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Tumor microenvironments present significant barriers to penetration by antibodies and immunoconjugates and are difficult to study in vitro. Cells cultured as monolayers typically exhibit less resistance to therapy than those grown in vivo. Therefore, it is important to develop an alternative research model that better represents in vivo tumors. We have developed a protocol to produce multicellular spheroids, a simple and more relevant model of in vivo tumors that allows for further investigations of the microenvironmental effects on drug penetration and tumor cell killing. The protocol is used to produce in vitro three-dimensional tumor spheroids from established human cancer cell lines and primary cancer cells isolated from patients without the use of any extracellular components. To study the ability of tumor-targeting immunoconjugates to penetrate these tumor spheroids in vitro, we have used an immunotoxin targeting mesothelin, a surface protein expressed in malignant mesotheliomas. This method for producing consistent, reproducible 3D spheroids may allow for improved testing of novel monoclonal antibodies and other agents for their ability to penetrate solid tumors for cancer therapy.

In plain English

This technical report described a laboratory method for growing cancer cells as small three-dimensional clusters, called multicellular spheroids, instead of flat layers. The aim was to test how well antibody-based drugs get into tumor-like cell masses. It did not study asbestos exposure. Cells were placed in 96-well plates coated with a non-stick material (poly-HEMA) and spun in a centrifuge. They included NCI-H226 cells, two liver cancer lines, and two mesothelioma lines taken from the abdominal fluid of malignant mesothelioma patients treated at the National Cancer Institute. The mesothelioma and liver cancer cells formed spheroids well and were ready within 2 days. The report tested SS1P, a drug aimed at mesothelin, a protein on mesothelioma cells. After 72 hours, the concentration needed to inhibit half the cells (IC50) was over 1000 ng/mL for spheroids. That was at least 100 times the roughly 10 ng/mL needed for flat layers. Silencing the protein E-cadherin increased sensitivity to SS1P. An antibody blocking E-cadherin helped only if added before the spheroids formed. The authors concluded the method may allow improved testing of antibodies for their ability to penetrate solid tumors.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Technical report describing a laboratory (in vitro) protocol for making 3D tumor spheroids
• Who was studied: Human cancer cell lines: NCI-H226, the liver cancer (HCC) lines HepG2 and Hep3B, and two primary mesothelioma cell lines (NCI-M-03 and NCI-M-13) isolated from the ascites of malignant mesothelioma patients treated at the NCI
• Exposure: No asbestos exposure was studied. Cells were treated with SS1P, a recombinant immunotoxin targeting mesothelin. BL22 served as a negative control and cycloheximide as a positive control.
• Main results:
  • The mesothelioma and liver cancer (HCC) cells used formed spheroids well.
  • Spheroids were ready for use within 2 days of seeding the cells.
  • After 72 hours, the IC50 of SS1P was more than 1000 ng/mL for spheroids. This was at least 100 times the IC50 for monolayers, about 10 ng/mL.
  • More than 50% of cancer cells from spheroids were not killed by SS1P concentrations as high as 1,000 ng/mL.
  • In primary lines from malignant mesothelioma patients, SS1P was also far less effective on spheroids.
  • Silencing E-cadherin with siRNA reduced its protein expression by more than 80% and was followed by increased sensitivity to SS1P.
  • An inhibitory antibody against E-cadherin enhanced anti-tumor activity only if added before, not after, spheroid formation.
• The authors' conclusion: The authors concluded that this method for producing consistent, reproducible 3D spheroids may allow improved testing of new monoclonal antibodies and other agents for their ability to penetrate solid tumors.
• Limitations the authors state:
  • Spheroids do not model all features of solid cancers, such as the influence of stroma and immune cells.

Topics

Medical terms in this study, explained in our Lexicon: Monoclonal Antibody, Tumor Microenvironment, Mesothelin, Mesothelioma.

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