Raman spectroscopy identifies radiation response in human non-small cell lung cancer xenografts.

External beam radiation therapy is a standard form of treatment for numerous cancers. Despite this, there are no approved methods to account for patient specific radiation sensitivity. In this report, Raman spectroscopy (RS) was used to identify radiation-induced biochemical changes in human non-small cell lung cancer xenografts. Chemometric analysis revealed unique radiation-related Raman signatures that were specific to nucleic acid, lipid, protein and carbohydrate spectral features. Among these changes was a dramatic shift in the accumulation of glycogen spectral bands for doses of 5 or 15 Gy when compared to unirradiated tumours. When spatial mapping was applied in this analysis there was considerable variability as we found substantial intra- and inter-tumour heterogeneity in the distribution of glycogen and other RS spectral features. Collectively, these data provide unique insight into the biochemical response of tumours, irradiated in vivo, and demonstrate the utility of RS for detecting distinct radiobiological responses in human tumour xenografts.

A Raman spectroscopic study of cell response to clinical doses of ionizing radiation.

The drive toward personalized radiation therapy (RT) has created significant interest in determining patient-specific tumor and normal tissue responses to radiation. Raman spectroscopy (RS) is a non-invasive and label-free technique that can detect radiation response through assessment of radiation-induced biochemical changes in tumor cells. In the current study, single-cell RS identified specific radiation-induced responses in four human epithelial tumor cell lines: lung (H460), breast (MCF-7, MDA-MB-231), and prostate (LNCaP), following exposure to clinical doses of radiation (2-10 Gy). At low radiation doses (2 Gy), H460 and MCF-7 cell lines showed an increase in glycogen-related spectral features, and the LNCaP cell line showed a membrane phospholipid-related radiation response. In these cell lines, only spectral information from populations receiving 10 Gy or less was required to identify radiation-related features using principal component analysis (PCA). In contrast, the MDA-MB-231 cell line showed a significant increase in protein relative to nucleic acid and lipid spectral features at doses of 6 Gy or higher, and high-dose information (30, 50 Gy) was required for PCA to identify this biological response. The biochemical nature of the radiation-related changes occurring in cells exposed to clinical doses was found to segregate by status of p53 and radiation sensitivity. Furthermore, the utility of RS to identify a biological response in human tumor cells exposed to therapeutic doses of radiation was found to be governed by the extent of the biochemical changes induced by a radiation response and is therefore cell line specific. The results of this study demonstrate the utility and effectiveness of single-cell RS to identify and measure biological responses in tumor cells exposed to standard radiotherapy doses.

Radiation-Induced Glycogen Accumulation Detected by Single Cell Raman Spectroscopy Is Associated with Radioresistance that Can Be Reversed by Metformin.

Altered cellular metabolism is a hallmark of tumor cells and contributes to a host of properties associated with resistance to radiotherapy. Detection of radiation-induced biochemical changes can reveal unique metabolic pathways affecting radiosensitivity that may serve as attractive therapeutic targets. Using clinically relevant doses of radiation, we performed label-free single cell Raman spectroscopy on a series of human cancer cell lines and detected radiation-induced accumulation of intracellular glycogen. The increase in glycogen post-irradiation was highest in lung (H460) and breast (MCF7) tumor cells compared to prostate (LNCaP) tumor cells. In response to radiation, the appearance of this glycogen signature correlated with radiation resistance. Moreover, the buildup of glycogen was linked to the phosphorylation of GSK-3ß, a canonical modulator of cell survival following radiation exposure and a key regulator of glycogen metabolism. When MCF7 cells were irradiated in the presence of the anti-diabetic drug metformin, there was a significant decrease in the amount of radiation-induced glycogen. The suppression of glycogen by metformin following radiation was associated with increased radiosensitivity. In contrast to MCF7 cells, metformin had minimal effects on both the level of glycogen in H460 cells following radiation and radiosensitivity. Our data demonstrate a novel approach of spectral monitoring by Raman spectroscopy to assess changes in the levels of intracellular glycogen as a potential marker and resistance mechanism to radiation therapy.