Humanin: A novel therapeutic target for TNBC

 

An aggressive form of Invasive Breast Cancer which accounts for about 10 to 20 percent of all Breast Cancers, is the Triple Negative Breast Cancer (TNBC). Triple Negative Breast Cancer grows and spreads rapidly as compared to other types of Breast Cancer. It is also a metastatic breast cancer, which means, it would grow and spread beyond the breast. New treatment strategies have been developed for breast tumors that express hormone receptors, and/or human epidermal growth factor receptor 2 (Her2), but for TNBC there are no therapeutic options, and chemotherapy/radiotherapy remains the first-line treatment. The chances of this cancer returning after treatment is also one of the factors, as to why the treatment methods for it are more limited than other Breast Cancers.

Multiple studies are being conducted to find potential novel therapeutic targets for the treatment of TNBC. In this study, the role of Humanin on tumor progression in TNBC is analyzed. Humanin (HN) is a mitochondrial-derived peptide with a cytoprotective effect in many tissues. HN was initially discovered in 2001 by Hashimoto using a cDNA library made from tissues of a healthy portion of Alzheimer's patient brain. HN was the first small open reading frame (ORF) identified within the mitochondrial DNA, encoded within the 16S rRNA gene (MT-RNR2). HN has been proven to protect normal tissues from chemotherapy, but its role in tumor progression is still not very clear.

Since the expression of HN in breast cancer had not been evaluated before, an assessment of the presence of HN and its mRNA in human and murine breast tumor cell lines was carried out. Flow cytometry analysis revealed the presence of HN in human MCF7 and T47D luminal breast tumor cells and MDA-MB-231 TNBC cells. Similar findings were also observed in murine breast cell lines. HN expression in human and murine breast cell lines was also evaluated by RT-PCR. The expression of HN in TNBC cells was confirmed through immunofluorescence. The expression of HN in sections of normal human mammary gland obtained from women undergoing cosmetic breast reduction, was evaluated. It was found that the expression of HN was stronger and wide-spread in TNBC cells as compared to normal breast cells.

To study the effect of HN on the response of murine and human TNBC cells to cytotoxic stimuli, different dosages of DOXO was used as treatment. DOXO (Doxorubicin) is a type of chemotherapy drug, which slows or stops the proliferation of cancer cells by blocking the topoisomerase II enzyme. 4T1 TNBC cells were used for the experiment, and incubated with various doses of liposomal DOXO. Cell viability was evaluated by MTT Assay and it was noticed that the cells were resistant to 100nM DOXO, but higher doses decreased the cell viability. To check the effects of HN on the cytotoxic effects of DOXO, 4T1 cells treated with 500nM DOXO were transfected with HN. It was observed that HN impaired both the anti-proliferative and pro-apoptotic effects of DOXO. Furthermore, the 4T1 cells were subjected to serum starvation. Serum starvation inhibited the proliferation of 4T1 cells. Low doses of HN (0.5 and 5 µM) did not show any significant changes, but an increased dosage of 10 µM HN reverted the anti-proliferative effects of serum starvation. The proliferation rate of 4T1 cells were restored back to control levels.

Fig 1: Effect of HN on chemosensitivity of TNBC Cells

To assess the effects of HN on tumor progression and chemosensitivity of TNBC in vivo, an experimental murine model was utilized. DOXO treatment was administered and later incubated with HN. HN reduced the number of apoptotic cells in tumors from both control and DOXO-treated cells. Furthermore, it was noticed that the administration of HN alone promoted the development of metastases as compared to control cells.

The bioinformatic analysis of transcriptomic data from different bioprojects available in NCBI showed that HN expression was found in all samples, which included normal and tumor breast cells, luminal and basal cell lines, and primary breast tumors of all types and their metastases.

To conclude, this study showed that HN facilitates tumor progression and chemo-resistance in experimental models of Triple Negative Breast Cancer (TNBC). It was found that HN and its receptors are upregulated in TNBC, and that exogenous and endogenous HN protects these cells from multiple cytotoxic and anti-proliferative stimuli. The study thus suggests that HN could possibly serve as a novel therapeutic target to improve the efficacy of chemotherapy in TNBC.

 

 

References:

1.Moreno Ayala MA, Gottardo MF, Zuccato CF, Pidre ML, Nicola Candia AJ, Asad AS, et al. Humanin Promotes Tumor Progression in Experimental Triple Negative Breast Cancer. Scientific Reports [Internet]. 2020 May 22 [cited 2022 Nov 7];10(1):8542. Available from: https://www.nature.com/articles/s41598-020-65381-7

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Image References:

1.    https://images.app.goo.gl/QZtVeviozqm4Z9Xc6

2.    Moreno Ayala MA, Gottardo MF, Zuccato CF, Pidre ML, Nicola Candia AJ, Asad AS, et al. Humanin Promotes Tumor Progression in Experimental Triple Negative Breast Cancer. Scientific Reports [Internet]. 2020 May 22 [cited 2022 Nov 7];10(1):8542. Available from: https://www.nature.com/articles/s41598-020-65381-7