Robust Engraftment of Acute Myeloid Leukemia (AML) in Humanized Scaffolds

Background

Acute myeloid leukemia (AML) is a malignant blood disorder that poses significant challenges in treatment and research. Patient-derived xenograft (PDX) models are crucial tools for studying AML, but existing PDX models are often complex and labor-intensive, limiting their widespread adoption. To address these issues, researchers have attempted to improve PDX models by implanting humanized niches. However, current methods typically require complex procedures and specific biomaterials, making them difficult to implement broadly.

This study aims to establish a simpler and more robust AML PDX model by streamlining procedures, optimizing biomaterial selection, and refining transplantation methods. The researchers hope that this approach will better mimic the growth environment of AML in humans, thereby providing a more reliable experimental platform for AML research and treatment.

Source of the Paper

This paper was co-authored by Daniel Busa, Zdenka Herudkova, Jan Hyl, and other researchers from the Faculty of Medicine at Masaryk University in the Czech Republic. The research team also included experts from multiple institutions, including the University Hospital Brno and the Brno University of Technology. The paper was published in 2025 in the journal Molecular Oncology with the DOI 10.¹⁰⁰²⁄₁₈₇₈-0261.13790.

Research Process and Results

1. Comparison and Selection of Biomaterials

The first step of the study was to evaluate the potential of different biomaterials in forming a humanized bone marrow niche. The researchers tested seven different biomaterials, including commercially available β-tricalcium phosphate (β-TCP) ceramic granules, extracellular matrix (ECM) gels from three different sources, and collagen-hydroxyapatite (HA) matrices and collagen fibers developed by academic teams.

Experimental Steps:

• Biomaterial Preparation: Each biomaterial was mixed with human bone marrow-derived mesenchymal stromal cells (MSCs) and implanted into immunodeficient mice either surgically or via injection.
• Scaffold Formation Assessment: After 8 weeks of implantation, the formation of bone tissue and bone marrow niches in the scaffolds was evaluated through histological examination and image analysis.

Results:

• Bone Tissue Formation: All materials were capable of forming bone tissue in the scaffolds, but the success rates varied. β-TCP ceramic granules and collagen-HA matrices showed the highest success rates.
• Bone Marrow Niche Formation: ECM gels and collagen-HA fibers performed best in forming bone marrow niches, supporting the growth of murine hematopoietic cells.

2. Evaluation of AML Engraftment

After identifying the optimal biomaterials, the researchers further assessed their impact on AML engraftment. Four biomaterials (β-TCP ceramic granules, Sigma ECM gel, collagen-HA matrix, and collagen-HA fiber) were selected, and two AML samples known to engraft well in untreated mice were implanted into these scaffolds.

Experimental Steps:

• AML Transplantation: AML cells were transplanted via intravenous (i.v.) injection or direct injection into the scaffolds (i.sc.).
• Engraftment Assessment: The engraftment of AML cells in the scaffolds and murine bone marrow was evaluated using flow cytometry and histological analysis.

Results:

• AML Engraftment: All biomaterials supported complete infiltration of AML cells, with no significant differences in engraftment efficacy between materials.
• Primitive AML Cell Proportion: The proportion of primitive CD34+ AML cells was higher in the scaffolds, indicating that the scaffolds better maintained the primitive state of AML cells.

3. Comparison of Transplantation Routes

The researchers also compared three different AML transplantation routes: intravenous (i.v.) injection, direct intra-scaffold (i.sc.) injection, and co-implantation of AML cells with the scaffold mixture (mix). The results showed that the i.v. route provided the most robust AML engraftment in both scaffolds and murine tissues, while direct intra-scaffold injection offered no significant advantage.

Results:

• Advantages of i.v. Injection: The i.v. route enabled widespread distribution of AML cells in both scaffolds and murine bone marrow, with a simpler procedure.
• T-cell Expansion Issue: T-cell expansion was observed in the i.sc. and mix routes, which may have affected AML engraftment.

4. Optimization of Scaffold Formation

To shorten the scaffold formation time, the researchers evaluated scaffold formation at different time points (4, 6, and 8 weeks). The results showed that 6 weeks were sufficient for bone marrow niche formation, while 4 weeks were inadequate.

Results:

• 6-week Scaffold Formation: Six weeks were sufficient for bone marrow niche formation, with results comparable to those at 8 weeks.
• Enhancement with BMP-2: The addition of bone morphogenetic protein 2 (BMP-2) during scaffold formation significantly increased scaffold size and bone marrow niche formation.

5. Engraftment of Non-engrafting AML Samples

Finally, the researchers tested whether the scaffolds could support the engraftment of non-engrafting or poorly engrafting AML samples. The results showed that the i.v. route supported engraftment in ²⁄₄ non-engrafting AML samples, while the i.sc. route supported only ¹⁄₄.

Results:

• Success Rate of i.v. Injection: The i.v. route demonstrated a higher success rate in supporting non-engrafting AML samples.
• Challenge of T-cell Expansion: T-cell expansion in the i.sc. route may have contributed to AML engraftment failure.

Conclusions and Significance

This study successfully established a simple and robust AML PDX model by streamlining procedures, optimizing biomaterial selection, and refining transplantation methods. The results demonstrated that the i.v. route is the most effective method for AML transplantation, while ECM gels and collagen-HA fibers are the optimal biomaterial choices. Additionally, the study found that adding BMP-2 significantly enhances scaffold formation.

The scientific value of this research lies in providing a more reliable experimental platform for AML studies, better mimicking the growth environment of AML in humans. Its practical value lies in offering a more efficient experimental tool for AML treatment and drug screening.

Research Highlights

• Streamlined Procedures: This study significantly simplified the establishment of AML PDX models by optimizing biomaterial selection and transplantation methods.
• Advantages of i.v. Injection: The study confirmed the superiority of the i.v. route in AML engraftment, providing important insights for future research.
• Enhancement with BMP-2: The addition of BMP-2 significantly improved scaffold formation, offering new ideas for scaffold optimization.