Trastuzumab Induced Chemobrain, Atorvastatin Rescued Chemobrain with Enhanced Anticancer Effect and without Hair Loss-Side Effect

Abstract

The authors identified that chemo-brain was induced after trastuzumab (TZB) therapy. In addition, atorvastatin (ATV) could rescue chemo-brain during trastuzumab (TZB) therapy. Enhanced therapeutic effect of TZB was confirmed after ATV therapy. We also investigated that there was no hair loss side effect due to ATV therapy. In an animal model, 150 μg TZB and five serial doses of 20 mg/kg ATV were administered. ¹⁸F-fluorodeoxyglucose Positron Emission Tomography (PET) and Magnetic Resonance Imaging (MRI) data were acquired. Statistical parametric mapping analysis and voxel-based morphometry analysis were performed to identify differences in glucose metabolism and gray matter concentration. The enhanced therapeutic efficacy of TZB after ATV treatment was assessed using a human epidermal growth factor receptor 2-positive gastric cancer model. We found a decrease in cerebral glucose metabolism and gray matter concentration in the frontal lobe following TZB therapy (p < 0.005). After subsequent ATV administration, glucose metabolism and regional gray matter concentration were rescued (p < 0.005). Cognitive impairment due to TZB and the rescue effect of ATV were confirmed using a passive avoidance test and quantitative real-time reverse transcription PCR. Furthermore, the penetration and accumulation of TZB in tumors increased by 100% after ATV co-administration, which resulted in an enhanced anti-cancer effect. Our study collectively demonstrates that ATV co-administration with TZB rescued the TZB-induced chemo-brain and enhances the therapeutic efficacy of TZB in tumors. We also showed that there was no hair loss during ATV therapy.

Keywords: anti-cancer effect; atorvastatin; chemo-brain; radiomics; trastuzumab therapy.

Figure 1

PET evidence of ATV effect during TZB therapy. (A) The experimental design. (B) A decrease in cerebral glucose metabolism was observed in the region of the bilateral frontal lobe following administration of TZB or (C) CTX relative to baseline (p < 0.005). (D) No significant difference was found between ATV treatment and baseline (p < 0.005). (E) An increase in cerebral glucose metabolism was observed in the region of the medial prefrontal cortex following administration of ATV (p < 0.005). Decreased glucose metabolism after TZB treatment was rescued after administration of ATV. PET, positron emission imaging. FDG PET, ¹⁸F-fluorodeoxyglucose positron emission tomography. ATV, atorvastatin. TZB, trastuzumab. CTX, cyclophosphamide. (n = 10 per group).

Figure 2

MR VBM evidence of ATV effect during TZB therapy. (A) The experimental design. (B) A decrease in gray matter concentration was observed in the region of the left frontal association cortex during TZB therapy and in the region of the frontal association cortex and hippocampus on the left side. (C) During CTX therapy relative to the baseline (p < 0.005). (D) No significant difference between ATV and the baseline was observed (p < 0.005). (E) Decreased regional gray matter concentration in the region of the frontal association cortex during TZB therapy was rescued when ATV was administered (p < 0.005). MR, magnetic resonance imaging. T2w, T2-weighted imaging. 3D, three-dimensional. VBM, voxel-based morphometry. ATV, atorvastatin. TZB, trastuzumab. CTX, cyclophosphamide. (n = 7 per group).

Figure 3

Behavioral study and pertaining the level of cytokines. (A) Schematic of passive avoidance test. Passive avoidance tests were performed to evaluate memory impairment in all groups. (B) The mean time of latency. The ATV co-administration group significantly extended the mean time of step-through latency (time of latency was 259 s) compared with that of the single TZB administration group. Values are means ± standard deviation of the mean (n = 6). (** p < 0.005). (C) Quantitative real-time reverse transcription polymerase chain reaction. (* p < 0.05 and ** p < 0.005). ATV, atorvastatin. TZB, trastuzumab.

Figure 4

Evidence of no side effect of ATV on hair regrowth. (A) The design of the experiment performed to test the in vivo side effect of ATV on hair regrowth in C57BL/6 mice. (B) Representative images of back skin gloss of mice and cropped image with higher magnification. ATV, atorvastatin. PBS, phosphate-buffered saline. CTX, cyclophosphamide. (n = 7 per group).

Figure 5

Effect of ATV on TZB accumulation in the tumor tissue. (A) The experimental design of TZB penetration into tumors. (B) In vivo localization of Alexa-488-TZB in NCI-N87 tumor tissue (scale bar = 500 μm). (C) Alexa-488-TZB accumulation per tumor tissue area (** p < 0.005), (D) In vivo penetration of Alexa-488-TZB after co-administration with ATV in NCI-N87 tumor tissue. Representative images of Alexa-488-TZB (green), a rhodamine-lectin stained functional blood vessel (red), and green and red merged with DAPI staining (blue) (scale bar = 50 μm). (E) In vivo penetration of Alexa-488-TZB is represented by histograms of Alexa-488 fluorescent intensity. (F) Images of functional vessel and segmented functional vessel. (G) Density of rhodamine-lectin positive functional blood vessel (N.S.). ATV, atorvastatin. TZB, trastuzumab. CTX, cyclophosphamide. (n = 7 per group).

Figure 5

Effect of ATV on TZB accumulation in the tumor tissue. (A) The experimental design of TZB penetration into tumors. (B) In vivo localization of Alexa-488-TZB in NCI-N87 tumor tissue (scale bar = 500 μm). (C) Alexa-488-TZB accumulation per tumor tissue area (** p < 0.005), (D) In vivo penetration of Alexa-488-TZB after co-administration with ATV in NCI-N87 tumor tissue. Representative images of Alexa-488-TZB (green), a rhodamine-lectin stained functional blood vessel (red), and green and red merged with DAPI staining (blue) (scale bar = 50 μm). (E) In vivo penetration of Alexa-488-TZB is represented by histograms of Alexa-488 fluorescent intensity. (F) Images of functional vessel and segmented functional vessel. (G) Density of rhodamine-lectin positive functional blood vessel (N.S.). ATV, atorvastatin. TZB, trastuzumab. CTX, cyclophosphamide. (n = 7 per group).

Figure 6

Cytotoxicity of ATV and effect of ATV on apoptosis and tumor growth delay. (A) Ex vivo image of cell nuclei. (B) The edge contours of DAPI-stained cell nuclei were extracted using a Laplacian edge-finding algorithm. (C) Representative images of cell nuclei when CTX or ATV was co-administered with TZB. (D,E) Circularity and fractal dimension of the cell nucleus in tumor tissue (mean ± standard deviation) (** p < 0.005 and * p < 0.05). ATV, atorvastatin. TZB, trastuzumab. CTX, cyclophosphamide. FD, fractal dimension. (F) Cytotoxicity assay according to the concentration of ATV. (G) Cell viability according to the concentration of TZB with ATV (* p < 0.05). (H) Cell lysates were immunoblotted with the indicated antibodies. (I) Relative intensity of Bcl-2. (J) Relative intensity of cleaved PARP. (n = 3, mean ± standard deviation, * p < 0.05). (K) Tumor growth delay for 30 days (n = 3–5 per group). ATV, atorvastatin. TZB, trastuzumab. PARP, poly (ADP-ribose) polymerase 1.