"Cell" sub-issue: Cancer cells "play for their lives" drug resistance!

One of the most common features of cancer cells is the aneuploid karyotype, which may be caused by chromosomal instability (CIN) [1]
.

CIN drives cell-to-cell heterogeneity and has profound effects on cancer cell genomes, tumor pathogenesis, tumor evolution, metastatic spread, and the choice and success of treatments [2-4]
.

Recently, a research team led by Dr.
Jason M.
Sheltzer from Cold Spring Harbor Laboratory published important research results in the famous journal Development Cell [5]
.

They found that chromosomal instability (CIN) can accelerate the development of tumor drug resistance
.

In recent years, the continuous improvement of cancer therapy has significantly improved the survival rate and quality of life of patients with various malignancies
.

However, the resistance of tumors to therapeutic drugs is one of the main reasons hindering the prolongation of patients' survival[6,7]
.

Drug resistance is mainly attributed to new mutations generated by tumor cells during treatment, which can restore the function of the therapeutic target during drug treatment, or bypass the effect of drugs [8]
.

We still have many questions about how cancer develops drug resistance
.

Studies have found that aneuploidy will cause yeast to reduce the sensitivity of various antifungal agents [9-13]
.

So, is there an association between aneuploidy and drug resistance in human tumors? To test whether CIN affects tumor drug resistance, Sheltzer's team chose to induce transient CIN in cancer cell lines.
They treated the cancer cells with the Mps1 inhibitor AZ3146 and found that the cancer cells had significantly increased mitotic error rates
.

Sheltzer designed a competitive strategy: the same cancer cell line (such as human melanoma cell line A375) was divided into two groups of equal numbers, one transfected with blue fluorescent protein (BFP) and one transfected with red fluorescent protein (RFP) ); the BFP group and the RFP group were divided into two groups, and a group of cells (BFP2, RFP1) were selected to induce CIN by Mps1i treatment, and the Mps1i was washed away after 24 hours, and the non-Mps1i-treated cells (RFP2, BFP1) ) co-culture, that is, BFP1 and RFP1 group/BFP2 and RFP2 group cells were co-cultured; the next passage was about 30 days, and flow cytometry was used to detect the abundance of cell populations at each passage to observe the effect of various conditions on cancer cell growth.
effect (Figure 1A)
.

Figure 1A Schematic diagram of competition experiment First, under standard culture conditions, Sheltzer induced transient CIN using Mps1 inhibitor, which made cancer cells (human melanoma cell line A375) aneuploid, and observed that CIN was harmful to cancer cells; However, in the presence of a certain concentration of the therapeutic drug (vemurafenib), it was observed that CIN accelerated the development of tumor resistance (Fig.
1B)
.

Figure 1B A375 cell population competition experiment Sheltzer separated the cell populations generated after the competition experiment, and tested the sensitivity of different drugs respectively.
It was found that the sensitivity of different treated cell populations to vemurafenib decreased to varying degrees, while for There was no resistance to paclitaxel and doxorubicin (Fig.
1C)
.

This indicates that the cells pretreated with Mps1i will not develop cross-resistance to other drugs after developing resistance to specific drugs
.

Figure 1C Response of cell populations to various drugs after 24 hours of competition experiments This phenomenon was confirmed in cells of various genetic backgrounds or when different targeted drugs were used (Figure 2)
.

The Sheltzer team treated the A375 cell line with paclitaxel (Fig.
2A); the Colo205 cell line with vemurafenib and paclitaxel, respectively (Fig.
2B); and the RPE1 cell line with paclitaxel (Fig.
2C); all observed that CIN accelerated resistance in these cell lines production of medicinal properties
.

Figure 2.
Mps1i accelerates the acquisition of drug resistance in different genetic backgrounds.
In the competition experiment, the Mps1i-treated cell population initially decreased substantially, and after a period of passage, it began to increase, gradually surpassing the untreated Mps1i-treated cell population and occupying the population of dominance
.

What is the reason for this phenomenon? Prof.
Sheltzer hypothesized that treatment with Mps1i would result in an initial population of cells with different karyotypes; most aneuploidies are detrimental to cells and cannot compete with untreated normal cell populations at the beginning of the experiment; but some rare Aneuploidy can promote the resistance of cancer cells to specific drugs, and under the pressure of the presence of drugs, cancer cells with these aneuploidies are selected
.

(Figure 3A) Figure 3A Aneuploidy screening schema.
To test this hypothesis, Sheltzer performed single-cell sequencing of cells from different competition experiments
.

It was found by sequencing: on the 0th and 6th day of the experiment, the cell heterogeneity after Mps1i treatment was significantly increased; on the 21st day of the experiment, when the Mps1i-treated cell population exceeded the untreated cell population, the Mps1i treatment The cellular heterogeneity decreased and the karyotype stabilized (Fig.
3B,C)
.

At the same time, Sheltzer found that the same anticancer drug can select different aneuploidy in different genetic backgrounds, rather than similar karyotypes (Fig.
3D,E)
.

Figure 3B-E Single-cell sequencing results at different time points after Mps1i treatment Sheltzer further screened out a variety of drug-resistant karyotypes through single-cell sequencing (Table 1)
.

Table 1.
Summary of single-cell karyotypes of Mps1i-treated cell lines.
Could the appearance of these aneuploidies repeatedly confer drug resistance in cancer cells? Or is it random and does not recur? To answer this question, Sheltzer isolated two karyotypes in p53-/- RPE1 cells, the two most common karyotypes (Mono10c2, c4) in paclitaxel cultures (Fig.
4A, B)
.

At the same time, a karyotype (Mono13) that was never observed in the competition experiment was selected as a control
.

Then, competition experiments were performed with these three types of cells against their original cells
.

The experiment found that Mono13-RPE1 cells were not significantly different from normal cells; while the competitiveness of Mono10-RPE1 cells in standard medium was significantly reduced; in the medium containing paclitaxel, the abundance of Mono10-RPE1 cells increased from 46% to 89% %, which indicates that the phenomenon that the same karyotype accelerates the drug resistance of cancer cells is repeatable (Fig.
4C)
.

Figure 4.
The drug resistance caused by aneuploidy is reproducible.
So, does endogenous CIN have the same effect as exogenous CIN? Sheltzer analyzed a dataset of patient xenografts (PDX) and found that 11% of "low-CIN tumors" had a significant response to treatment, compared with only 3% of "high-CIN tumors" (Fig.
5A); 69% of "High-CIN tumors" had progressive disease after treatment, while only 58% of "low-CIN tumors" (Fig.
5B); "high-CIN tumors" had shorter survival when treated with drugs (Fig.
5D-F)
.

Figure 5A-F Results of analysis of xenograft (PDX) datasets Sheltzer also analyzed clinical and copy number data from The Cancer Genome Atlas (TCGA)
.

Since the CIN of the TCGA samples could not be measured, the researchers evaluated the fractional genome alteration (FGA) as an indicator
.

The analysis found that at the end of the observation period, individuals with high-FGA tumors were significantly more likely to have tumors than those with low-FGA tumors (Fig.
5G)
.

High FGA was also significantly associated with disease progression in colorectal, glioma, prostate and endometrial cancers (Fig.
5H)
.

The relationship between FGA and patient prognosis was evaluated by regression analysis, and it was found that FGA was significantly associated with shorter survival in patients with some cancers, such as breast cancer, acute myeloid leukemia, papillary renal cell carcinoma, etc.
(Fig.
5I)
.

Figure 5G-I Results of The Cancer Genome Atlas (TCGA) dataset analysis These studies found that high levels of endogenous CIN in the absence of Mps1 inhibitors are associated with progressive disease and treatment resistance in multiple cancer types increase related
.

Through these findings, we can use Mps1 as a potential therapeutic target and develop related Mps1 inhibitors as new tumor drugs
.

However, these findings also suggest that Mps1 inhibitors should be used with caution in the clinic to avoid inadvertently accelerating the development of drug resistance in cancer patients
.

Reference: [1] Vasudevan A, Schukken KM, Sausville EL, Girish V, Adebambo OA, Sheltzer JM.
Aneuploidy as a promoter and suppressor of malignant growth.
Nat Rev Cancer.
2021;21(2):89-103.
doi :10.
1038/s41568-020-00321-1[2] Dürrbaum M, Kuznetsova AY, Passerini V, Stingele S, Stoehr G, Storchová Z.
Unique features of the transcriptional response to model aneuploidy in human cells.
BMC Genomics.
2014;15 :139.
Published 2014 Feb 18.
doi:10.
1186/1471-2164-15-139[3]Oromendia AB, Dodgson SE, Amon A.
Aneuploidy causes proteotoxic stress in yeast.
Genes Dev.
2012;26(24):2696- 2708.
doi:10.
1101/gad.
207407.
112[4]Donnelly N, Passerini V, Dürrbaum M, Stingele S, Storchová Z.
HSF1 deficiency and impaired HSP90-dependent protein folding are hallmarks of aneuploid human cells.
EMBO J.
2014;33( 20):2374-2387.
doi:10.
15252/embj.
201488648[5] Lukow DA, Sausville EL, Suri P, et al.
Chromosomal instability accelerates the evolution of resistance to anti-cancer therapies [published online ahead of print, 2021 Aug 2].
Dev Cell.
2021;S1534-5807(21)00592-X.
doi:10.
1016/j.
devcel.
2021.
07.
009 [6] Baudino TA.
Targeted Cancer Therapy: The Next Generation of Cancer Treatment.
Curr Drug Discov Technol.
2015;12(1):3-20.
doi:10.
2174/1570163812666150602144310[7] Johnson SB, Park HS, Gross CP, Yu JB.
Use of Alternative Medicine for Cancer and Its Impact on Survival.
J Natl Cancer Inst.
2018;110(1):10.
1093/jnci/djx145.
doi:10.
1093/jnci/djx145[8] Sullivan RJ, Flaherty KT.
Resistance to BRAF-targeted therapy in melanoma.
Eur J Cancer.
2013;49(6):1297-1304.
doi:10.
1016/j.
ejca.
2012.
11.
019[9] Chen G, Bradford WD, Seidel CW, Li R.
Hsp90 Stress potentiates rapid cellular adaptation through induction of aneuploidy.
Nature.
2012;482(7384):246-250.
Published 2012 Jan 29.
doi:10.
1038/nature10795 [10] Chen G, Mulla WA, Kucharavy A, et al.
Targeting the adaptability of heterogeneous aneuploids.
Cell.
2015;160(4):771-784.
doi:10.
1016/j .
cell.
2015.
01.
026[11] Pavelka N, Rancati G, Zhu J, et al.
Aneuploidy confers quantitative proteome changes and phenotypic variation in budding yeast.
Nature.
2010;468(7321):321-325.
doi:10.
1038/ nature09529[12] Selmecki AM, Dulmage K, Cowen LE, Anderson JB, Berman J.
Acquisition of aneuploidy provides increased fitness during the evolution of antifungal drug resistance.
PLoS Genet.
2009;5(10):e1000705.
doi:10.
1371/journal .
pgen.
1000705[13] Yang F, Teoh F, Tan ASM, Cao Y, Pavelka N, Berman J.
Aneuploidy Enables Cross-Adaptation to Unrelated Drugs.
Mol Biol Evol.
2019;36(8):1768-1782.
doi :10.
1093/molbev/msz104 Editor-in-Chief | Dai Siyu1038/nature10795 [10] Chen G, Mulla WA, Kucharavy A, et al.
Targeting the adaptability of heterogeneous aneuploids.
Cell.
2015;160(4):771-784.
doi:10.
1016/j.
cell.
2015.
01.
026[ 11] Pavelka N, Rancati G, Zhu J, et al.
Aneuploidy confers quantitative proteome changes and phenotypic variation in budding yeast.
Nature.
2010;468(7321):321-325.
doi:10.
1038/nature09529[12] Selmecki AM, Dulmage K, Cowen LE, Anderson JB, Berman J.
Acquisition of aneuploidy provides increased fitness during the evolution of antifungal drug resistance.
PLoS Genet.
2009;5(10):e1000705.
doi:10.
1371/journal.
pgen.
1000705[13] Yang F, Teoh F, Tan ASM, Cao Y, Pavelka N, Berman J.
Aneuploidy Enables Cross-Adaptation to Unrelated Drugs.
Mol Biol Evol.
2019;36(8):1768-1782.
doi:10.
1093/molbev/msz104 Liability Editor | Dai Siyu1038/nature10795 [10] Chen G, Mulla WA, Kucharavy A, et al.
Targeting the adaptability of heterogeneous aneuploids.
Cell.
2015;160(4):771-784.
doi:10.
1016/j.
cell.
2015.
01.
026[ 11] Pavelka N, Rancati G, Zhu J, et al.
Aneuploidy confers quantitative proteome changes and phenotypic variation in budding yeast.
Nature.
2010;468(7321):321-325.
doi:10.
1038/nature09529[12] Selmecki AM, Dulmage K, Cowen LE, Anderson JB, Berman J.
Acquisition of aneuploidy provides increased fitness during the evolution of antifungal drug resistance.
PLoS Genet.
2009;5(10):e1000705.
doi:10.
1371/journal.
pgen.
1000705[13] Yang F, Teoh F, Tan ASM, Cao Y, Pavelka N, Berman J.
Aneuploidy Enables Cross-Adaptation to Unrelated Drugs.
Mol Biol Evol.
2019;36(8):1768-1782.
doi:10.
1093/molbev/msz104 Liability Editor | Dai SiyuTargeting the adaptability of heterogeneous aneuploids.
Cell.
2015;160(4):771-784.
doi:10.
1016/j.
cell.
2015.
01.
026[11] Pavelka N, Rancati G, Zhu J, et al.
Aneuploidy confers quantitative proteome changes and phenotypic variation in budding yeast.
Nature.
2010;468(7321):321-325.
doi:10.
1038/nature09529[12] Selmecki AM, Dulmage K, Cowen LE, Anderson JB, Berman J.
Acquisition of aneuploidy provides increased fitness during the evolution of antifungal drug resistance.
PLoS Genet.
2009;5(10):e1000705.
doi:10.
1371/journal.
pgen.
1000705[13] Yang F, Teoh F, Tan ASM, Cao Y, Pavelka N, Berman J.
Aneuploidy Enables Cross-Adaptation to Unrelated Drugs.
Mol Biol Evol.
2019;36(8):1768-1782.
editor in charge of doi:10.
1093/molbev/msz104 | Dai SiyuTargeting the adaptability of heterogeneous aneuploids.
Cell.
2015;160(4):771-784.
doi:10.
1016/j.
cell.
2015.
01.
026[11] Pavelka N, Rancati G, Zhu J, et al.
Aneuploidy confers quantitative proteome changes and phenotypic variation in budding yeast.
Nature.
2010;468(7321):321-325.
doi:10.
1038/nature09529[12] Selmecki AM, Dulmage K, Cowen LE, Anderson JB, Berman J.
Acquisition of aneuploidy provides increased fitness during the evolution of antifungal drug resistance.
PLoS Genet.
2009;5(10):e1000705.
doi:10.
1371/journal.
pgen.
1000705[13] Yang F, Teoh F, Tan ASM, Cao Y, Pavelka N, Berman J.
Aneuploidy Enables Cross-Adaptation to Unrelated Drugs.
Mol Biol Evol.
2019;36(8):1768-1782.
editor in charge of doi:10.
1093/molbev/msz104 | Dai SiyuRancati G, Zhu J, et al.
Aneuploidy confers quantitative proteome changes and phenotypic variation in budding yeast.
Nature.
2010;468(7321):321-325.
doi:10.
1038/nature09529[12] Selmecki AM, Dulmage K, Cowen LE , Anderson JB, Berman J.
Acquisition of aneuploidy provides increased fitness during the evolution of antifungal drug resistance.
PLoS Genet.
2009;5(10):e1000705.
doi:10.
1371/journal.
pgen.
1000705[13] Yang F, Teoh F , Tan ASM, Cao Y, Pavelka N, Berman J.
Aneuploidy Enables Cross-Adaptation to Unrelated Drugs.
Mol Biol Evol.
2019;36(8):1768-1782.
doi:10.
1093/molbev/msz104 Editor-in-Chief | Dai SiyuRancati G, Zhu J, et al.
Aneuploidy confers quantitative proteome changes and phenotypic variation in budding yeast.
Nature.
2010;468(7321):321-325.
doi:10.
1038/nature09529[12] Selmecki AM, Dulmage K, Cowen LE , Anderson JB, Berman J.
Acquisition of aneuploidy provides increased fitness during the evolution of antifungal drug resistance.
PLoS Genet.
2009;5(10):e1000705.
doi:10.
1371/journal.
pgen.
1000705[13] Yang F, Teoh F , Tan ASM, Cao Y, Pavelka N, Berman J.
Aneuploidy Enables Cross-Adaptation to Unrelated Drugs.
Mol Biol Evol.
2019;36(8):1768-1782.
doi:10.
1093/molbev/msz104 Editor-in-Chief | Dai SiyuAcquisition of aneuploidy provides increased fitness during the evolution of antifungal drug resistance.
PLoS Genet.
2009;5(10):e1000705.
doi:10.
1371/journal.
pgen.
1000705[13] Yang F, Teoh F, Tan ASM, Cao Y, Pavelka N, Berman J.
Aneuploidy Enables Cross-Adaptation to Unrelated Drugs.
Mol Biol Evol.
2019;36(8):1768-1782.
doi:10.
1093/molbev/msz104 Editor-in-Chief | Dai SiyuAcquisition of aneuploidy provides increased fitness during the evolution of antifungal drug resistance.
PLoS Genet.
2009;5(10):e1000705.
doi:10.
1371/journal.
pgen.
1000705[13] Yang F, Teoh F, Tan ASM, Cao Y, Pavelka N, Berman J.
Aneuploidy Enables Cross-Adaptation to Unrelated Drugs.
Mol Biol Evol.
2019;36(8):1768-1782.
doi:10.
1093/molbev/msz104 Editor-in-Chief | Dai Siyu