Cell Journal: Meal Replacement for Cancer Treatment?

(TNBC)（BC），BC，，
。-，，(CSCs)，TNBC
。

(TNBC)（BC），BC，，
。-，，(CSCs)，TNBC
。

CSCs are a slowly circulating cancer cell characterized by low levels of intracellular reactive oxygen species (ROS), which contributes to their self-renewal potential and resistance to DNA and other macromolecular damage induced by chemotherapy or radiotherapy
.

As hypoxia-inducible factor (HIF-1) mediates the up-regulation of glucose transporter (GLUT) and glycolytic enzymes (such as hexokinase (HK)), this metabolic plasticity enables CSCs to survive in unfavorable tumor microenvironments Survival, making it the most resistant subgroup of cancer cells

CSCs are a slowly circulating cancer cell characterized by low levels of intracellular reactive oxygen species (ROS), which contributes to their self-renewal potential and resistance to DNA and other macromolecular damage induced by chemotherapy or radiotherapy

In recent years, studies have shown that the fasting/simulated fasting (FMD) cycle improves the efficacy of standard therapies for different types of cancers (including BC), and at the same time protects normal cells from the toxicity caused by the treatment

Since CSCs play a key role in promoting tumor initiation, progression and resistance to treatment, this study investigated the effect of FMD on the survival of CSC, and how it will affect the growth of TNBC and the response to targeted therapy
.

Since CSCs play a key role in promoting tumor initiation, progression and resistance to treatment, this study investigated the effect of FMD on the survival of CSC, and how it will affect the growth of TNBC and the response to targeted therapy
.

1, fast / FMD by reducing hypoglycemic levels TNBC reducing human stem cells

1, fast / FMD by reducing hypoglycemia reducing human stem cells TNBC level 1, fast / FMD by reducing hypoglycemia reducing human stem cells TNBC level 1,fast / FMD by reducing hypoglycemia reduce levels of human stem cells TNBChypoglycaemia

Compared with free diet (AL), FMD cycling reduced the number of primary mammary glands in vitro and the percentage of cells expressing acetaldehyde dehydrogenase 1 (ALDH1) (Figures 1A and 1B)
.

Compared with AL, FMD reduces the frequency of cells that can promote TNBC and increases tumor progression-free survival (Figure S1E)

Compared with free diet (AL), FMD cycling reduced the number of primary mammary glands in vitro and the percentage of cells expressing acetaldehyde dehydrogenase 1 (ALDH1) (Figures 1A and 1B)

To evaluate the effect of glucose supplementation in the body, 3% glucose was added to the drinking water of mice fed the FMD cycle, which is based on the normal daily glucose concentration assumed by the mice through a standard diet

Figure 1 SUM159 human triple negative breast cancer (with TNBC) in, the FMD can reduce breast cell growth ball group, the CSC marker expression and frequency of stem cells, which may be conducted by 2-deoxy -d - glucose ( 2DG) enhancement
.

Figure 1 SUM159 human triple negative breast cancer (with TNBC) in, the FMD can reduce breast cell growth ball group, the CSC marker expression and frequency of stem cells, which may be conducted by 2-deoxy -d - glucose ( 2DG) enhancement
.

2DG)

2.

In the syngeneic TNBC model, FMD can reduce TNBC CSCs and delay tumor progression

2.
In the syngeneic TNBC model, FMD can reduce TNBC CSCs and delay tumor progression 2.
In the syngeneic TNBC model, FMD can reduce TNBC CSCs and delay tumor progression

To confirm the data in the syngeneic mouse TNBC model, the effect of fasting/FMD in 4T1 mouse transplantation was tested in BALB/c female mice with strong immunity
.

Compared with AL conditions, 4 FMD cycles delayed tumor progression, reduced the number of mammary glands and the percentage of CD44CD24+-and ALDH1 cells, thus confirming the anti-CSC effect previously reported in the human SUM159 model (Figure 2A)

To confirm the data in the syngeneic mouse TNBC model, the effect of fasting/FMD in 4T1 mouse transplantation was tested in BALB/c female mice with strong immunity

Since metastasis is the main cause of death in TNBC patients, and CSCs play a key role in the spread of metastasis, we evaluated the effect of circulating FMD, alone or in combination with 2DG, on the formation of metastasis in TNBC mice

Figure 2 shows that in the syngeneic TNBC model, FMD functions independently of the immune system and reduces metastasis

Figure 2 shows that in the syngeneic TNBC model, FMD acts independently of the immune system and reduces metastasis .

Figure 2 shows that in the syngeneic TNBC model, FMD acts independently of the immune system and reduces metastasis.

3.
Low baseline blood sugar is associated with increased survival in patients with metastatic TNBC

3.
Low baseline blood glucose is associated with an increase in the survival rate of patients with metastatic TNBC 3.
Low baseline blood glucose is associated with an increase in the survival rate of patients with metastatic TNBC 3.
Low baseline blood glucose is associated with an increase in the survival rate of patients with metastatic TNBC

In order to determine whether the findings in mice are related to the progression of human TNBC, we studied the correlation of baseline blood glucose concentration to the overall survival (OS) of patients with advanced TNBC, and retrospectively evaluated the platinum exposure between October 2006 and January 2020.
Advanced BC patients undergoing basic chemotherapy (Figure S4A)
.
Among 418 patients with different BC subtypes, we selected 81 patients with advanced TNBC who received continuous first-line carboplatin-paclitaxel or carboplatin-gemcitabine treatment (Table S1)
.
If the baseline blood glucose level is less than 100 mg/dL, the blood glucose is considered normal
.
Compared with patients with normoglycemia, patients with hyperglycemia had significantly lower OS (Figure 3A)
.
The multivariate model confirmed the independent negative prognostic effect of hyperglycemia on the OS of patients (HR hyperglycemia patients vs.
euglycemia patients = 1.
95; 95% CI 1.
05-3.
64; p = 0.
035) (Figure 3B)
.

In order to determine whether the findings in mice are related to the progression of human TNBC, we studied the correlation of baseline blood glucose concentration to the overall survival (OS) of patients with advanced TNBC, and retrospectively evaluated the platinum exposure between October 2006 and January 2020.
Advanced BC patients undergoing basic chemotherapy (Figure S4A)
.
Among 418 patients with different BC subtypes, we selected 81 patients with advanced TNBC who received continuous first-line carboplatin-paclitaxel or carboplatin-gemcitabine treatment (Table S1)
.
If the baseline blood glucose level is less than 100 mg/dL, the blood glucose is considered normal
.
Compared with patients with normoglycemia, patients with hyperglycemia had significantly lower OS (Figure 3A)
.
The multivariate model confirmed the independent negative prognostic effect of hyperglycemia on the OS of patients (HR hyperglycemia patients vs.
euglycemia patients = 1.
95; 95% CI 1.
05-3.
64; p = 0.
035) (Figure 3B)
.
In order to determine whether the findings in mice are related to the progression of human TNBC, we studied the correlation of baseline blood glucose concentration to the overall survival (OS) of patients with advanced TNBC, and retrospectively evaluated the platinum exposure between October 2006 and January 2020.
Advanced BC patients undergoing basic chemotherapy (Figure S4A)
.
Among 418 patients with different BC subtypes, we selected 81 patients with advanced TNBC who received continuous first-line carboplatin-paclitaxel or carboplatin-gemcitabine treatment (Table S1)
.
If the baseline blood glucose level is less than 100 mg/dL, the blood glucose is considered normal
.
Compared with patients with normoglycemia, patients with hyperglycemia had significantly lower OS (Figure 3A)
.
The multivariate model confirmed the independent negative prognostic effect of hyperglycemia on the OS of patients (HR hyperglycemia patients vs.
euglycemia patients = 1.
95; 95% CI 1.
05-3.
64; p = 0.
035) (Figure 3B)
.

Figure 3 In patients with advanced TNBC, hyperglycemia is associated with poor overall survival

Figure 3 In patients with advanced TNBC, hyperglycemia is associated with poor overall survival

4.
PKA activation reverses STS-dependent mammary gland shrinkage

4.
PKA activation reverses the reduction of STS-dependent mammary glands 4.
PKA activation reverses the reduction of STS-dependent mammary glands

Next, we will study the mechanism by which glucose consumption sensitizes CSCs
.
The activity of PKA was detected by detecting the phosphorylation of CREB, the classical downstream substrate of PKA, and the expression of KLF5 in SUM159 and 4T1 tumors
.
In both FMD mouse models, pCREB and KLF5 were down-regulated in tumors, indicating that the PKA pathway was inhibited (Figure 4A)
.
In addition, RNA sequencing analysis of SUM159 tumor masses revealed that FMD-induced PKA pathway inhibition selectively occurred in CD44CD24+−, but not in CD44CD24 differentiated cells (Figure 4B)
.

Next, we will study the mechanism by which glucose consumption sensitizes CSCs
.
The activity of PKA was detected by detecting the phosphorylation of CREB, the classical downstream substrate of PKA, and the expression of KLF5 in SUM159 and 4T1 tumors
.
In both FMD mouse models, pCREB and KLF5 were down-regulated in tumors, indicating that the PKA pathway was inhibited (Figure 4A)
.
In addition, RNA sequencing analysis of SUM159 tumor masses revealed that FMD-induced PKA pathway inhibition selectively occurred in CD44CD24+−, but not in CD44CD24 differentiated cells (Figure 4B)
.
Next, we will study the mechanism by which glucose consumption sensitizes CSCs
.
The activity of PKA was detected by detecting the phosphorylation of CREB, the classical downstream substrate of PKA, and the expression of KLF5 in SUM159 and 4T1 tumors
.
In both FMD mouse models, pCREB and KLF5 were down-regulated in tumors, indicating that the PKA pathway was inhibited (Figure 4A)
.
In addition, RNA sequencing analysis of SUM159 tumor masses revealed that FMD-induced PKA pathway inhibition selectively occurred in CD44CD24+−, but not in CD44CD24 differentiated cells (Figure 4B)
.

Figure 4 FMD selectively down-regulates PKA in CSCs

Figure 4 FMD selectively down-regulates PKA in CSCs Figure 4 FMD selectively down-regulates PKA in CSCs

In SUM159 or 4T1 TNBC cells, there was no significant change in the expression of pCREB and KLF5 between CSCs and differentiated cancer cells under CTR conditions, while the expressions of pCREB and KLF5 in CSCs under STS conditions were selectively down-regulated (Figure 4C and 4D)
.
These results indicate that the PKA axis is selectively involved in the impact on CSCs, and PKA inhibits the role of stem cell regulation (Figure 5A, 5B and S5A)
.

In SUM159 or 4T1 TNBC cells, there was no significant change in the expression of pCREB and KLF5 between CSCs and differentiated cancer cells under CTR conditions, while the expressions of pCREB and KLF5 in CSCs under STS conditions were selectively down-regulated (Figure 4C and 4D)
.
These results indicate that the PKA axis is selectively involved in the impact on CSCs, and PKA inhibits the role of stem cell regulation (Figure 5A, 5B and S5A)
.

In SUM159 and 4T1 tumors, FMD down-regulated the levels of G9A and H3K9me2 (Figure 5C)
.
It is worth noting that exogenous activation of the PKA pathway by 8Br-cAMP reversed the STS/FMD-dependent down-regulation of KLF5 and H3K9me2 proteins.
Compared with STS alone, the expression of KLF5 and H3K9me2 proteins increased (Figure S5B)
.
Taken together, these data indicate that FMD-induced failure of TNBC CSCs is mediated at least through glucose reduction and PKA pathway inhibition
.

In SUM159 and 4T1 tumors, FMD down-regulated the levels of G9A and H3K9me2 (Figure 5C)
.
It is worth noting that exogenous activation of the PKA pathway by 8Br-cAMP reversed the STS/FMD-dependent down-regulation of KLF5 and H3K9me2 proteins.
Compared with STS alone, the expression of KLF5 and H3K9me2 proteins increased (Figure S5B)
.
Taken together, these data indicate that FMD-induced failure of TNBC CSCs is mediated at least through glucose reduction and PKA pathway inhibition
.
In SUM159 and 4T1 tumors, FMD down-regulated the levels of G9A and H3K9me2 (Figure 5C)
.
It is worth noting that exogenous activation of the PKA pathway by 8Br-cAMP reversed the STS/FMD-dependent down-regulation of KLF5 and H3K9me2 proteins.
Compared with STS alone, the expression of KLF5 and H3K9me2 proteins increased (Figure S5B)
.
Taken together, these data indicate that FMD-induced failure of TNBC CSCs is mediated at least through glucose reduction and PKA pathway inhibition
.
Taken together, these data indicate that FMD-induced failure of TNBC CSCs is mediated at least through glucose reduction and PKA pathway inhibition
.

5.
Combining the FMD cycle with PI3K/AKT/mTOR inhibitors can make the animals survive long-term and reduce the side effects caused by treatment

5.
The combined use of FMD cycles and PI3K/AKT/mTOR inhibitors can make animals survive long-term and reduce the side effects caused by treatment.
5.
The combined use of FMD cycles and PI3K/AKT/mTOR inhibitors can make animals survive long-term and reduce treatment causes the side effects of5,5,the FMD period and PI3K / AKT / mTOR inhibitor in combination with long-term use can make animal survival and reduce side effects caused by treatment

Although CSCs play a vital role in tumor formation and progression, differentiated cells also play a vital role in tumor volume
.
In fact, circulating FMD alone leads to significant depletion of CSCs, which will only delay tumor growth, but will not lead to cancer-free survival or long-term progression-free survival
.

Although CSCs play a vital role in tumor formation and progression, differentiated cells also play a vital role in tumor volume
.
In fact, circulating FMD alone leads to significant depletion of CSCs, which will only delay tumor growth, but will not lead to cancer-free survival or long-term progression-free survival
.

Studies have shown that FMD leads to increased expression of pro-apoptotic molecules (such as BIM and ASK1)
.
RNA-seq analysis also showed that FMD cycle up-regulates the CCND-CDK4/6 signal axis while up-regulating the PI3K-AKT and mTOR signaling pathways, and down-regulating CCNB-CDK1 (Figure 6A and S6A)
.

Studies have shown that FMD leads to increased expression of pro-apoptotic molecules (such as BIM and ASK1)
.
RNA-seq analysis also showed that FMD cycle up-regulates the CCND-CDK4/6 signal axis while up-regulating the PI3K-AKT and mTOR signaling pathways, and down-regulating CCNB-CDK1 (Figure 6A and S6A)
.
Studies have shown that FMD leads to increased expression of pro-apoptotic molecules (such as BIM and ASK1)
.
RNA-seq analysis also showed that FMD cycle up-regulates the CCND-CDK4/6 signal axis while up-regulating the PI3K-AKT and mTOR signaling pathways, and down-regulating CCNB-CDK1 (Figure 6A and S6A)
.
FMD cycles up-regulate the CCND-CDK4/6 signal axis while up-regulating the PI3K-AKT and mTOR signal pathways, and down-regulating CCNB-CDK1 (Figure 6A and S6A)
.

Then we investigated whether drugs inhibiting PI3K/AKT and mTOR pathways can enhance the effect of FMD by blocking the "starvation escape pathway" (SEPs), thereby promoting the survival of differentiated TNBC cells during FMD
.
In the mouse xenotransplantation of SUM159, weekly FMD significantly increased the anti-tumor activity of each drug inhibitor, resulting in a significant increase in the survival rate of mice (Figure 6B and S7A)
.
However, the most significant effect appears in the combination of all three drugs with the FMD cycle
.
In particular, the three drugs targeting PI3K/AKT and mTOR have a strong effect on the progression-free survival of mice fed a standard diet, but the addition of the FMD cycle leads to significant anti-tumor activity of the triple combination of PI3K/AKT and mTOR pathway inhibitors Enhanced
.
In 85% of mice, the tumor growth time exceeded 150 days (Figure 7A)
.

Then we investigated whether drugs inhibiting PI3K/AKT and mTOR pathways can enhance the effect of FMD by blocking the "starvation escape pathway" (SEPs), thereby promoting the survival of differentiated TNBC cells during FMD
.
In the mouse xenotransplantation of SUM159, weekly FMD significantly increased the anti-tumor activity of each drug inhibitor, resulting in a significant increase in the survival rate of mice (Figure 6B and S7A)
.
However, the most significant effect appears in the combination of all three drugs with the FMD cycle
.
In particular, the three drugs targeting PI3K/AKT and mTOR have a strong effect on the progression-free survival of mice fed a standard diet, but the addition of the FMD cycle resulted in significant anti-tumor activity of the triple combination of PI3K/AKT and mTOR pathway inhibitors Enhanced
.
In 85% of mice, the tumor growth time exceeded 150 days (Figure 7A)
.
Then we investigated whether drugs inhibiting PI3K/AKT and mTOR pathways can enhance the effect of FMD by blocking the "starvation escape pathway" (SEPs), thereby promoting the survival of differentiated TNBC cells during FMD
.
In the mouse xenotransplantation of SUM159, weekly FMD significantly increased the anti-tumor activity of each drug inhibitor, resulting in a significant increase in the survival rate of mice (Figure 6B and S7A)
.
However, the most significant effect appears in the combination of all three drugs with the FMD cycle
.
In particular, the three drugs targeting PI3K/AKT and mTOR have a strong effect on the progression-free survival of mice fed a standard diet, but the addition of the FMD cycle resulted in significant anti-tumor activity of the triple combination of PI3K/AKT and mTOR pathway inhibitors Enhanced
.
In 85% of mice, the tumor growth time exceeded 150 days (Figure 7A)
.

In this case, the observed anti-tumor effect may be partly mediated by strong inhibition of the PKA signaling pathway
.
In fact, we found that three pictilisib + ipatasertib + rapamycin treatments combined with STS, instead of single (pictilisib) or double (pictilisib + ipatasertib) treatments, strongly down-regulated the expression of KLF5 and H3K9me2 proteins
.

In this case, the observed anti-tumor effect may be partly mediated by strong inhibition of the PKA signaling pathway
.
In fact, we found that three pictilisib + ipatasertib + rapamycin treatments combined with STS, instead of single (pictilisib) or double (pictilisib + ipatasertib) treatments, strongly down-regulated the expression of KLF5 and H3K9me2 proteins
.
In this case, the observed anti-tumor effect may be partly mediated by strong inhibition of the PKA signaling pathway
.
In fact, we found that three pictilisib + ipatasertib + rapamycin treatments combined with STS, instead of single (pictilisib) or double (pictilisib + ipatasertib) treatments, strongly down-regulated the expression of KLF5 and H3K9me2 proteins
.

Figure 7 FMD reverses the late progression of TNBC by enhancing the effect of kinase inhibitors and reversing the death caused by hyperglycemia

Figure 7 FMD reverses the late progression of TNBC by enhancing the effect of kinase inhibitors and reversing the death caused by hyperglycemia

Therefore, we investigated whether the prevention of fasting/FMD-induced hyperglycemia can help prolong survival
.
Interestingly, the FMD cycle protects mice from hyperglycemia induced by PI3K/AKT and mTOR inhibitors (Figure 7B).
In some animals, hyperglycemia directly leads to death of mice without relying on tumor growth (Figure 7A).
)
.
The FMD cycle restores blood glucose to a normal level shortly after the medication, so that the blood glucose level is normal during most drug treatments (Figure 7B)
.
Based on RNA-seq results to evaluate the efficacy of the combination of CDK4/6 inhibitor palbociclib and pictilisib, it was found that palbociclib + pictilisib significantly delayed tumor progression, and the addition of the FMD cycle further improved its anti-tumor activity and delayed the acquisition of drug resistance (Figure 7C and S7C)
.
In addition, compared with drug treatment alone, fpd-palbociclib-pictilisib triple therapy also reduced the formation of mammary gland cells after 8 weeks of treatment (Figure 7C)
.

Therefore, we investigated whether the prevention of fasting/FMD-induced hyperglycemia can help prolong survival
.
Interestingly, the FMD cycle protects mice from hyperglycemia induced by PI3K/AKT and mTOR inhibitors (Figure 7B).
In some animals, hyperglycemia directly leads to death of mice without relying on tumor growth (Figure 7A).
)
.
The FMD cycle restores blood glucose to a normal level shortly after the medication, so that the blood glucose level is normal during most drug treatments (Figure 7B)
.
Based on RNA-seq results to evaluate the efficacy of the combination of CDK4/6 inhibitor palbociclib and pictilisib, it was found that palbociclib + pictilisib significantly delayed tumor progression, and the addition of the FMD cycle further improved its anti-tumor activity and delayed the acquisition of drug resistance (Figure 7C and S7C)
.
In addition, compared with drug treatment alone, fpd-palbociclib-pictilisib triple therapy also reduced the formation of mammary gland cells after 8 weeks of treatment (Figure 7C)
.
Therefore, we investigated whether the prevention of fasting/FMD-induced hyperglycemia can help prolong survival
.
Prevention.
Interestingly, the FMD cycle protects mice from PI3K/AKT and mTOR inhibitors-induced hyperglycemia (Figure 7B).
In some animals, hyperglycemia directly leads to death of mice, independent of tumor growth (Figure 7B).
7A)
.
The FMD cycle restores blood glucose to a normal level shortly after the medication, so that the blood glucose level is normal during most drug treatments (Figure 7B)
.
Based on RNA-seq results to evaluate the efficacy of the combination of CDK4/6 inhibitor palbociclib and pictilisib, it was found that palbociclib + pictilisib significantly delayed tumor progression, and the addition of the FMD cycle further improved its anti-tumor activity and delayed the acquisition of drug resistance (Figure 7C and S7C)
.
In addition, compared with drug treatment alone, fpd-palbociclib-pictilisib triple therapy also reduced the formation of mammary gland cells after 8 weeks of treatment (Figure 7C)
.

In summary:

In summary: In summary:

1. Fasting/FMD consumes TNBC stem cells by lowering blood sugar levels
2. PKA activation reverses FMD-induced reduction in cancer stem cells
3. In cancer cells, FMD activates a targeted starvation escape mechanism
4. FMD+kinase inhibitor can make TNBC regress without drug toxicity

Fasting/FMD consumes TNBC stem cells by lowering blood sugar levels

Fasting/FMD consumes TNBC stem cells by lowering blood sugar levels

PKA activation reverses FMD-induced reduction in cancer stem cells

PKA activation reverses FMD-induced reduction in cancer stem cells

In cancer cells, FMD activates a targeted starvation escape mechanism

In cancer cells, FMD activates a targeted starvation escape mechanism

FMD+kinase inhibitor can make TNBC regress without drug toxicity

FMD+kinase inhibitor can make TNBC regress without drug toxicity

The results of this study show that fasting simulated meal replacement has a wide range of different effects on normal cells, cancer cells and cancer stem cells.
It can quickly activate the starvation escape signal transduction pathway and be blocked by kinase inhibitors corresponding to the target.
Fasting simulates Meal replacement + kinase inhibitors can cause triple-negative breast cancer regression and reduce drug toxicity.
It provides a new method that may be applicable to many malignant cancers.
Therefore, it is necessary to conduct further human clinical studies for verification
.

The results of this study show that fasting simulated meal replacement has a wide range of different effects on normal cells, cancer cells and cancer stem cells.
It can quickly activate the starvation escape signal transduction pathway and be blocked by kinase inhibitors corresponding to the target.
Fasting simulates Meal replacement has a wide range of different effects on normal cells, cancer cells and cancer stem cells.
It can quickly activate the starvation escape signal transduction pathway and be blocked by the kinase inhibitors of the corresponding target.
Fasting simulated meal replacement has effects on normal cells and cancer cells.
It has a wide range of different effects from cancer stem cells.
It can quickly activate the starvation escape signal transduction pathway and be blocked by the kinase inhibitor corresponding to the target.
Fasting simulated meal replacement + kinase inhibitor can cause the regression of triple-negative breast cancer and reduce Drug toxicity provides a new method that may be applicable to many malignant cancers, so it is necessary to conduct further human clinical studies for verification
.
Fasting simulated meal replacement + kinase inhibitors can cause triple-negative breast cancer regression and reduce drug toxicity.
It provides a new method that may be applicable to many malignant cancers.
Therefore, it is necessary to conduct further human clinical studies for verification
.

Original source:

Giulia Salvadori, et al.
Fasting-mimicking diet blocks triple-negative breast cancer and cancer stem cell escape.
Cell Metab.
2021 Nov 2;33(11):2247-2259.
e6.

Giulia Salvadori, et al.
Fasting-mimicking diet blocks triple-negative breast cancer and cancer stem cell escape.
Cell Metab.
2021 Nov 2;33(11):2247-2259.
e6.

DOI: 10.
1016/j.
cmet.
2021.
10.
008

DOI: 10.
1016/j.
cmet.
2021.
10.
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