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Background <p>L-asparaginase has been a widely employed as antitumor enzyme for the treatment of acute lymphoblastic leukemia for almost three decades. The enzyme takes advantage of the inability of tumor cells to synthesize the L-asparagine and is killed by L‐asparagine deprivation. Despite the availability of bacterial sources for L-asparaginase, there is a growing interest in identifying new microbial sources with improved therapeutic properties. Therefore, this study aims to investigate the production of L-asparaginase from a fungal source, to explore its potential as a novel alternative enzyme for cancer treatment.</p> Results <p><i>Fusarium falciforme</i> AUMC 16563 was used to produce L-asparaginase (123.42 U/mL) after 5 days, 0.2% glucose and 1.0% asparagine; were used at 25 <b>˚</b>C and pH 8.0. Employing two columns of chromatography (DEAE-cellulose and Sephacryl S 200 HR), the enzyme was purified 14.26-fold, reaching a maximum activity of 5109.4 U/mg. SDS-PAGE revealed a 46.06&#xa0;kDa asparaginase. The <i>K</i><sub>m</sub> and <i>V</i><sub>max</sub> values for pure asparaginase using asparagine was 5.77 × 10<sup>− 2</sup> mM and 128.22 µmol/min. Additionally, <i>Fusarium falciforme</i> AUMC 16563’ pure asparaginase demonstrated anticancer activity against PC-3 (a prostate cell line) with an IC<sub>50</sub> of 78.6 µg/mL, HePG-2 (a human hepatocellular carcinoma cell line) with an IC<sub>50</sub> of 69.6 µg/mL, HCT-116 (a colon cell line) with an IC<sub>50</sub> of 51.5 µg/mL and MCF-7 (a breast cancer cell line) with an IC<sub>50</sub> of 32.8 µg/mL. The expression levels of proapoptotic genes (BAX and p53) were significantly greater in the breast cancer cell lines treated with asparaginase than in the negative control breast cancer cell lines.The degree of DNA fragmentation in MCF-7 cells treated with <i>Fusarium falciforme</i> 16563’ pure asparaginase was 27.2 ± 0.69%, and that in MCF-7 cells treated with the drug Doxorubicin 24.1 ± 0.86% was significantly greater than that in the corresponding negative control cells 9.1 ± 1.01%. Finally, the biochemical profiles revealed no impact on the liver or the kidneys. These results suggested that asparaginase had relatively little effect on liver function. All hematological parameters were within normal range during the experiment.</p> Conclusions <p>The results of the present study revealed a potent L-ASNase from endophytic <i>F. falciforme</i> isolated from <i>Trifolium alexandrinum</i>, which performs well under a variety of environmental circumstances and can be used in a number of commercial applications.</p>

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L-asparaginase from the novel Fusarium falciforme AUMC 16563: extraction, purification, characterization, and cytotoxic effects on PC-3, HePG-2, HCT-116, and MCF-7 cell lines

  • 期刊论文

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  • 发表日期：

    2025年3月17日

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  • Abdullah Abobakr Saleh,
  • Hamdy M. El-Aref,
  • Azza M. Ezzeldin,
  • Rania M. Ewida,
  • Osama A. M. Al-Bedak

摘要

Background

L-asparaginase has been a widely employed as antitumor enzyme for the treatment of acute lymphoblastic leukemia for almost three decades. The enzyme takes advantage of the inability of tumor cells to synthesize the L-asparagine and is killed by L‐asparagine deprivation. Despite the availability of bacterial sources for L-asparaginase, there is a growing interest in identifying new microbial sources with improved therapeutic properties. Therefore, this study aims to investigate the production of L-asparaginase from a fungal source, to explore its potential as a novel alternative enzyme for cancer treatment.

Results

Fusarium falciforme AUMC 16563 was used to produce L-asparaginase (123.42 U/mL) after 5 days, 0.2% glucose and 1.0% asparagine; were used at 25 ˚C and pH 8.0. Employing two columns of chromatography (DEAE-cellulose and Sephacryl S 200 HR), the enzyme was purified 14.26-fold, reaching a maximum activity of 5109.4 U/mg. SDS-PAGE revealed a 46.06 kDa asparaginase. The Km and Vmax values for pure asparaginase using asparagine was 5.77 × 10− 2 mM and 128.22 µmol/min. Additionally, Fusarium falciforme AUMC 16563’ pure asparaginase demonstrated anticancer activity against PC-3 (a prostate cell line) with an IC50 of 78.6 µg/mL, HePG-2 (a human hepatocellular carcinoma cell line) with an IC50 of 69.6 µg/mL, HCT-116 (a colon cell line) with an IC50 of 51.5 µg/mL and MCF-7 (a breast cancer cell line) with an IC50 of 32.8 µg/mL. The expression levels of proapoptotic genes (BAX and p53) were significantly greater in the breast cancer cell lines treated with asparaginase than in the negative control breast cancer cell lines.The degree of DNA fragmentation in MCF-7 cells treated with Fusarium falciforme 16563’ pure asparaginase was 27.2 ± 0.69%, and that in MCF-7 cells treated with the drug Doxorubicin 24.1 ± 0.86% was significantly greater than that in the corresponding negative control cells 9.1 ± 1.01%. Finally, the biochemical profiles revealed no impact on the liver or the kidneys. These results suggested that asparaginase had relatively little effect on liver function. All hematological parameters were within normal range during the experiment.

Conclusions

The results of the present study revealed a potent L-ASNase from endophytic F. falciforme isolated from Trifolium alexandrinum, which performs well under a variety of environmental circumstances and can be used in a number of commercial applications.