Pilot study of vascular health in survivors of Hodgkin lymphoma.

BACKGROUND: Vascular-related toxicities have been reported among survivors of Hodgkin lymphoma (HL), but their genesis is not well understood. PROCEDURE: Fasting blood samples from 25 previously irradiated HL survivors were analyzed for biomarkers that can reveal underlying inflammation and/or endothelial cell activation: high-sensitivity C-reactive protein (hsCRP), triglycerides, total cholesterol, high-density lipoprotein (HDL), apolipoprotein ß, lipoprotein (a), fibrinogen, circulating endothelial cells (CECs), and vascular cell adhesion molecule-1 (VCAM-1) expression. Values were compared to subjects in the Coronary Artery Risk Development in Young Adults (CARDIA) study. CECs and VCAM-1 were compared to healthy controls. RESULTS: Survivors (76% male), median age 17.6 years (5-33) at diagnosis, 33.0 years (19-55) at follow-up, included stages IA (n = 6), IIA (n = 10), IIB (n = 2), IIIA (n = 4), and IVA (n = 3) patients. Twenty-four received at least chest radiation therapy (RT) (median dose 3,150 cGy; range: 175-4,650 cGy), one received neck only; 14 (56%) had a history of anthracycline exposure (median dose: 124 mg/m(2) range: 63-200 mg/m2). Compared to CARDIA subjects, mean hsCRP (3.0 mg/L ± 2.0 vs. 1.6 ± 1.9), total cholesterol (194.1 mg/dl ± 33.2 vs. 179.4 ± 32.9), lipoprotein (a) (34.2 mg/dl ± 17.5 vs. 13.8 ± 17.5), and fibrinogen (342.0 mg/dl ± 49.1 vs. 252.6 ± 48.4) were significantly elevated. CECs (2.3 cells/ml ± 1.5 vs. 0.34 ± 1.4) were significantly elevated compared to controls. No difference in VCAM-1 expression (51.1% ± 36.8 vs. 42.3 ± 35.6) was detected. CONCLUSION: HL survivors exposed to RT have evidence of vascular inflammation, dyslipidemia, and injury suggestive of early atherogenesis.

Predicting the Local Response of Metastatic Brain Tumor to Gamma Knife Radiosurgery by Radiomics With a Machine Learning Method.

PURPOSE: The current study proposed a model to predict the response of brain metastases (BMs) treated by Gamma knife radiosurgery (GKRS) using a machine learning (ML) method with radiomics features. The model can be used as a decision tool by clinicians for the most desirable treatment outcome. METHODS AND MATERIAL: Using MR image data taken by a FLASH (3D fast, low-angle shot) scanning protocol with gadolinium (Gd) contrast-enhanced T1-weighting, the local response (LR) of 157 metastatic brain tumors was categorized into two groups (Group I: responder and Group II: non-responder). We performed a radiomics analysis of those tumors, resulting in more than 700 features. To build a machine learning model, first, we used the least absolute shrinkage and selection operator (LASSO) regression to reduce the number of radiomics features to the minimum number of features useful for the prediction. Then, a prediction model was constructed by using a neural network (NN) classifier with 10 hidden layers and rectified linear unit activation. The training model was evaluated with five-fold cross-validation. For the final evaluation, the NN model was applied to a set of data not used for model creation. The accuracy and sensitivity and the area under the receiver operating characteristic curve (AUC) of the prediction model of LR were analyzed. The performance of the ML model was compared with a visual evaluation method, for which the LR of tumors was predicted by examining the image enhancement pattern of the tumor on MR images. RESULTS: By the LASSO analysis of the training data, we found seven radiomics features useful for the classification. The accuracy and sensitivity of the visual evaluation method were 44 and 54%. On the other hand, the accuracy and sensitivity of the proposed NN model were 78 and 87%, and the AUC was 0.87. CONCLUSIONS: The proposed NN model using the radiomics features can help physicians to gain a more realistic expectation of the treatment outcome than the traditional method.

Necrotizing leukoencephalopathy following CHOP chemotherapy.

Toxic leukoencephalopathy syndromes are rare disorders of cerebral injury characterized by changes in the white matter and accompanying neurologic dysfunction. They have been reported in association with a variety of clinical etiologies, most commonly including severe hypertension, cranial irradiation, and environmental toxins. However, they have also been described in conjunction with immunosuppressive and chemotherapeutic agents. There has been one case of fatal leukoencephalopathy reported following CHOP chemotherapy for non-Hodgkin lymphoma. We report a second case of fatal necrotizing leukoencephalopathy following the administration of CHOP chemotherapy.

Cumulative Doses to Brain and Other Critical Structures After Multisession Gamma Knife Stereotactic Radiosurgery for Treatment of Multiple Metastatic Tumors.

PURPOSE: Repeat stereotactic radiosurgery (SRS) is an attractive alternative to whole brain radiation therapy (WBRT) for treatment of recurrent brain metastases (BM). The purpose of this study is to determine the cumulative doses to the brain and critical normal structures in patients who underwent repeat courses of Gamma Knife (GK) SRS. MATERIALS AND METHODS: We retrospectively identified ten patients who received at least three GK-SRS sessions for multiply recurrent BM at our institution from 2013 to 2016. We used Velocity™ 3.1.0 software to co-register the magnetic resonance imaging images and the dose data of all treatment sessions for each patient. The cumulative doses to brain, lenses, eyes, brainstem, optic nerves, chiasm, and hippocampi were calculated. Dose-volume histograms, as well as the mean, median and maximum doses of these structures, were analyzed. RESULTS: The median number of SRS was five sessions (range = 3-7 sessions) per patient over a median treatment span of 510 days (112-1,197 days), whereas the median number of metastatic tumors treated per patient was 25.0 (10-63). The median of the total tumor volume was 9.5 cc (2.3-75.9 cc). The median of the mean cumulative dose to the whole brain was 4.1 Gy (1.7-16.4 Gy). The medians of the maximum doses to the critical structures were as follows: brainstem, 6.1 Gy (2.2-28.9 Gy), chiasm, 3.9 Gy (1.8-10.8 Gy), right optic nerve, 2.9 Gy (1.2-9.0 Gy), and left optic nerve, 2.6 Gy (1.0-6.5 Gy). The medians of the mean and maximum cumulative doses to the hippocampi were 3.4 Gy (1.0-14.4 Gy) and 13.8 Gy (1.5-39.3 Gy), respectively. The median survival for the entire cohort was 26.7 months, and no patients developed radiation necrosis. CONCLUSION: Our study demonstrated that multisession GKSRS could be delivered with low cumulative doses to critical normal structures. Further studies are required to fully establish its role as an alternative treatment strategy to WBRT for the treatment of multiply recurrent BM.

Role of Consolidative Radiation Therapy After Autologous Hematopoietic Cell Transplantation for the Treatment of Relapsed or Refractory Hodgkin Lymphoma.

PURPOSE: To evaluate the role of the addition of consolidative radiation therapy after high-dose chemotherapy and autologous hematopoietic cell transplantation (AHCT) for relapsed or refractory Hodgkin lymphoma (HL). METHODS AND MATERIALS: Medical records were reviewed from a total of 80 consecutive patients who underwent high-dose chemotherapy with AHCT treated under a single protocol at University of Minnesota between November 2005 and January 2014. Of these, 32 patients received radiation therapy after AHCT as planned consolidation. RESULTS: At a median follow-up of 25 months, the 2-year overall survival (OS) and progression-free survival (PFS) for the entire cohort was 96% and 52%, respectively. Consolidative radiation therapy was found to significantly improve the 2-year PFS (67% vs 42%, P<.01) without a significant change in OS (100% vs 93%, P=.15). On subgroup analysis, consolidative radiation therapy was shown to improve PFS in patients with bulky disease (62% vs 39%, P=.02), B-symptoms (48% vs 28%, P=.05), primary refractory disease (47% vs 32%, P=.02), and those with a partial response on pretransplant imaging (47% vs 32%, P=.02). The improvement seen on 2-year PFS with consolidative radiation therapy remained significant on multivariate analysis (hazard ratio 4.64, 95% confidence interval 1.98-10.88). Minimal toxicity was observed among the patients receiving radiation therapy. CONCLUSIONS: The addition of consolidative radiation therapy after high-dose chemotherapy and AHCT demonstrated a significant improvement in 2-year PFS and no impact on OS. Radiation therapy was well tolerated, with minimal toxicity. Our study supports a role of consolidative radiation therapy in patients with HL treated with AHCT.

Evolving role of radiation therapy for hematologic malignancies.

Radiation therapy continues to play a paramount role in the therapy of hematologic malignancies, whether as definitive therapy, as consolidation after chemotherapy, as part of bone marrow transplantation protocols, or in palliation. During the past 2 decades, significant advances in radiation therapy have occurred, including the evolution of involved-field irradiation and the adoption of conformal radiation administration. It is hoped that modern techniques will reduce the long-term sequelae associated with radiation-based treatments.

Geometrical accuracy of a 3-tesla magnetic resonance imaging unit in Gamma Knife surgery.

OBJECT: The authors sought to evaluate and improve the geometrical accuracy of a 3-tesla magnetic resonance (MR) imaging unit used for Gamma Knife surgery (GKS). METHODS: To evaluate the geometrical accuracy of a Siemens Magnetom Trio 3-tesla MR imaging unit, two phantoms were used. Both phantoms were imaged with computed tomography (CT), a 1.5-tesla MR imaging unit (Siemens Avanto), and the 3-tesla MR imaging unit. A pair of orthogonal films was obtained with a radiotherapy simulator to validate the spatial coordinates of the marker positions determined with CT. The coordinates of the markers were measured using the GammaPlan treatment planning software. Magnetic resonance imaing was performed using three-dimensional (3D) magnetization-prepared rapid acquisition gradient echo (MPRAGE) and fast low-angle shot sequence (FLASH) pulse sequences. The voxel size was 1 x 1 x 1 mm3. CONCLUSIONS: The root-mean-square error of MR images was 2 +/- 0.73 mm for 3D MPRAGE. The error was reduced to 1.5 +/- 0.64 mm for FLASH. The errors were decreased further by applying an image distortion correction method (the field-of-view filter) to the images acquired with FLASH. The mean errors were 1.3 +/- 0.39 mm and 1.5 +/- 0.77 mm for the two phantoms. The errors increased from 1 mm to 3.1 mm as the measurement points approached the caudal edge of the head coil (larger z value). Proper selection of a pulse sequence together with a geometrical distortion correction improved the geometrical accuracy of MR images. However, further study is needed to increase the geometrical accuracy of 3-tesla MR imaging units for radiosurgical applications.

Heat-induced up-regulation of NAD(P)H:quinone oxidoreductase potentiates anticancer effects of beta-lapachone.

PURPOSE: The purpose of the present study was to evaluate the efficacy of mild hyperthermia to potentiate the anticancer effects of beta-lapachone (3,4-dihydro-2,2-dimethyl-2H-naphthol[1,2-b]pyran-5,6-dione) by up-regulating NAD(P)H:quinone oxidoreductase (NQO1) in cancer cells. EXPERIMENTAL DESIGN: Effects of beta-lapachone alone or in combination with mild heating on the clonogenic survival of FSaII fibrosarcoma cells of C3H mice and A549 human lung tumor cells in vitro was determined. Effects of heating on the NQO1 level in the cancer cells in vitro were assessed using Western blot analysis for NQO1 expression, biochemical determination of NQO1 activity, and immunofluorescence microscopy for NQO1 expression. Growth of FSaII tumors in the hind legs of C3H mice was determined after treating the host mice with i.p. injection of 45 mg/kg beta-lapachone followed by heating the tumors at 42 degrees C for 1 hour every other day for four times. RESULTS: Incubation of FSaII tumor cells and A549 tumor cells with beta-lapachone at 37 degrees C reduced clonogenic survival of the cells in dose-dependent and incubation time-dependent manner. NQO1 level in the cancer cells in vitro increased within 1 hour after heating at 42 degrees C for 1 hour and remained elevated for >72 hours. The clonogenic cell death caused by beta-lapachone increased in parallel with the increase in NQO1 levels in heated cells. Heating FSaII tumors in the legs of C3H mice enhanced the effect of i.p.-injected beta-lapachone in suppressing tumor growth. CONCLUSION: We observed for the first time that mild heat shock up-regulates NQO1 in tumor cells. The heat-induced up-regulation of NQO1 enhanced the anticancer effects of beta-lapachone in vitro and in vivo.

BRAF Mutation Is Associated with Improved Local Control of Melanoma Brain Metastases Treated with Gamma Knife Radiosurgery.

OBJECTIVES: Evidence has implicated a possible role of tumor mutation status on local control (LC) with radiotherapy. BRAF is a proto-oncogene that is mutated in approximately 50% of patients with melanoma. We sought to analyze the influence of BRAF status on LC of melanoma brain metastases (MBM) following Gamma Knife radiosurgery (GK). METHODS: Among 125 patients treated with GK for MBM at our institution between 2006 and 2015, we identified 19 patients with 69 evaluable metastases whose BRAF mutation status was known and follow-up imaging was available. LC of individual metastases was compared based on BRAF mutation status using statistical techniques to control for measurements of multiple metastases within each patient. CNS progression was defined as either local failure or development of new lesions. RESULTS: Of the 69 metastases, BRAF was mutated in 30 and wild-type in 39. With a median follow-up of 30 months for all patients and a median follow-up of 5.5 months for treated lesions, 1-year LC was significantly better among metastases with mutated vs. wild-type BRAF (69 vs. 34%, RR = 0.3, 95% CI = 0.1-0.7, p = 0.01). BRAF mutation was found to be a significant predictor of LC after stereotactic radiosurgery (SRS) in both univariate [RR = 0.3 (95% CI 0.1-0.7, p = 0.01)] and multivariate [RR = 0.2 (95% CI 0.1-0.7, p = 0.01)] analyses. There was also a trend toward improved CNS progression free survival (PFS) at 1 year (26 vs. 0%, p = 0.06), favoring BRAF-mutated patients. CONCLUSION: In this retrospective study, MBM treated with GK had significantly improved LC for patients with BRAF mutation vs. wild-type. Our data suggest that BRAF mutation may sensitize tumors to radiosurgery, and that BRAF wild-type tumors may be more radioresistant.

A phase I feasibility study of multi-modality imaging assessing rapid expansion of marrow fat and decreased bone mineral density in cancer patients.

PURPOSE: Cancer survivors are at an increased risk for fractures, but lack of effective and economical biomarkers limits quantitative assessments of marrow fat (MF), bone mineral density (BMD) and their relation in response to cytotoxic cancer treatment. We report dual energy CT (DECT) imaging, commonly used for cancer diagnosis, treatment and surveillance, as a novel biomarker of MF and BMD. METHODS: We validated DECT in pre-clinical and phase I clinical trials and verified with water-fat MRI (WF-MRI), quantitative CT (QCT) and dual-energy X-ray absorptiometry (DXA). Basis material composition framework was validated using water and small-chain alcohols simulating different components of bone marrow. Histologic validation was achieved by measuring percent adipocyte in the cadaver vertebrae and compared with DECT and WF-MRI. For a phase I trial, sixteen patients with gynecologic malignancies (treated with oophorectomy, radiotherapy or chemotherapy) underwent DECT, QCT, WF-MRI and DXA before and 12months after treatment. BMD and MF percent and distribution were quantified in the lumbar vertebrae and the right femoral neck. RESULTS: Measured precision (3mg/cm(3)) was sufficient to distinguish test solutions. Adiposity in cadaver bone histology was highly correlated with MF measured using DECT and WF-MRI (r=0.80 and 0.77, respectively). In the clinical trial, DECT showed high overall correlation (r=0.77, 95% CI: 0.69, 0.83) with WF-MRI. MF increased significantly after treatment (p<0.002). Chemotherapy and radiation caused greater increases in MF than oophorectomy (p<0.032). L4 BMD decreased 14% by DECT, 20% by QCT, but only 5% by DXA (p<0.002 for all). At baseline, we observed a statistically significant inverse association between MF and BMD which was dramatically attenuated after treatment. CONCLUSION: Our study demonstrated that DECT, similar to WF-MRI, can accurately measure marrow adiposity. Both imaging modalities show rapid increase in MF following cancer treatment. Our results suggest that MF and BMD cannot be used interchangeably to monitor skeletal health following cancer therapy.

Cytotoxicity of perillyl alcohol against cancer cells is potentiated by hyperthermia.

PURPOSE: Perillyl alcohol (POH) (4-isopropenyl-cyclohexenecarbinol) is a member of the monoterpenes, which are present in various fruits and vegetables. POH has been demonstrated to be cytotoxic against a variety of experimental cancer cells in vitro and in vivo. Phase I clinical trials have indicated that POH may be useful for human tumor treatment. The purpose of our study was to reveal whether the anticancer effect of POH could be enhanced by hyperthermia. METHODS AND MATERIALS: SCK mammary carcinoma cells of A/J mice were used. The effects of POH or hyperthermia alone were studied by incubating the cells during exponential growth phase in culture with 0.25-1.0 mM of POH at 37 degrees C for varying lengths of time or heating cells at 41-43 degrees C for varying lengths of time. The combined effect of POH and hyperthermia was investigated by heating the cells with 1 mM of POH at 41-43 degrees C for varying lengths of time. The effects of the treatments were evaluated using the clonogenic cell survival assay and three types of apoptosis assays. RESULTS: An incubation of SCK cells with 1 mM of POH at 37 degrees C for 60 min or hyperthermia at 43 degrees C for 1 h decreased clonogenic cell survival to 40% and 60%, respectively. When the cells were heated at 43 degrees C for 1 h in the presence of 1 mM of POH, clonogenic cell survival decreased to 0.2%, indicating that hyperthermia potentiated the effect of POH to cause clonogenic cell death. Hyperthermia also markedly increased the degree of POH-induced apoptosis. CONCLUSION: Hyperthermia synergistically potentiates the cytotoxicity of naturally occurring POH against cancer cells.

Radiation dose response for supratentorial low-grade glioma--institutional experience and literature review.

PURPOSE: To examine radiation dose response for low-grade glioma (LGG) based on our institutional experience and to review the literature on this topic. METHODS AND MATERIALS: Sixty-seven patients with supratentorial low-grade nonpilocytic astrocytomas (n=36) or oligodendrogliomas (n=31) were treated with postoperative radiation therapy (RT). Twenty-seven patients (group A) received 5520 cGy; 24 patients (group B) received 5940 cGy; and 16 patients (group C) received 6375 cGy. The corresponding median follow-up was 60, 35 and 91 months, respectively. RESULTS: The disease-specific survival (DSS) at 5 and 10 years were 90.2% and 56.2%, 67.6% and 47.3%, and 62.5% and 50% for groups A, B and C, respectively (P=0.40). Only a greater extent of surgical resection and absence of contrast enhancement predicted DSS on multivariate analyses. Patients receiving higher doses of RT had higher complication rates. CONCLUSION: Our data confirmed the lack of radiation dose response for supratentorial LGG as demonstrated in the previous randomized trials. The radiation dose should not exceed 5520 cGy because dose escalation did not result in an improvement of DSS and it also increased the complication rates. Future research should focus on the eradication of radioresistant clones either by the improvement of surgical resection or the use of cytotoxic agents that can target on the radioresistant tumor cells.

Single dose versus fractionated stereotactic radiotherapy for meningiomas.

OBJECTIVE: To evaluate the safety and efficacy of stereotactic radiosurgery (SRS) compared to fractionated stereotactic radiation therapy (FSRT) for meningiomas treated over a seven year period. METHODS AND MATERIALS: Of the 53 patients (15 male and 38 female) with 63 meningiomas, 35 were treated with SRS and the 18 patients with tumors adjacent to critical structures or with large tumors were treated with FSRT. The median doses for the SRS and the FSRT groups were 1400 cGy (500-4500 cGy) and 5400 cGy (4000-6000 cGy) respectively. Median target volumes for SRS and FSRT were 6.8 ml and 8.8 ml respectively. The median follow-up for the SRS and FSRT groups were 38 months (4.1-97 months) and 30.5 months (6.0-63 months) respectively. RESULTS: The five-year tumor control probability (TC) for benign versus atypical meningiomas were 92.7% vs. 31% (P = .006). The three-year TC were 92.7% vs. 93.3% for SRS vs. FSRT groups respectively (P = .62). For benign meningiomas, the three-year TC were 92.9% vs. 92.3% for the SRS group (29 patients) vs. FSRT group (14 patients) respectively (P = .77). Two patients in the SRS group and one in the FSRT group developed late complications. CONCLUSION: Preliminary data suggest that SRS is a safe and effective treatment for patients with benign meningiomas. Fractionated stereotactic radiation therapy with conventional fractionation appeared to be an effective and safe treatment alternative for patients not appropriate for SRS. A longer follow-up is required to determine the long-term efficacy and the toxicity of these treatment modalities.

Response of breast cancer cells and cancer stem cells to metformin and hyperthermia alone or combined.

Metformin, the most widely prescribed drug for treatment of type 2 diabetes, has been shown to exert significant anticancer effects. Hyperthermia has been known to kill cancer cells and enhance the efficacy of various anti-cancer drugs and radiotherapy. We investigated the combined effects of metformin and hyperthermia against MCF-7 and MDA-MB-231 human breast cancer cell, and MIA PaCa-2 human pancreatic cancer cells. Incubation of breast cancer cells with 0.5-10 mM metformin for 48 h caused significant clonogenic cell death. Culturing breast cancer cells with 30 µM metformin, clinically relevant plasma concentration of metformin, significantly reduced the survival of cancer cells. Importantly, metformin was preferentially cytotoxic to CD44(high)/CD24(low) cells of MCF-7 cells and, CD44(high)/CD24(high) cells of MIA PaCa-2 cells, which are known to be cancer stem cells (CSCs) of MCF-7 cells and MIA PaCa-2 cells, respectively. Heating at 42°C for 1 h was slightly toxic to both cancer cells and CSCs, and it markedly enhanced the efficacy of metformin to kill cancer cells and CSCs. Metformin has been reported to activate AMPK, thereby suppressing mTOR, which plays an important role for protein synthesis, cell cycle progression, and cell survival. For the first time, we show that hyperthermia activates AMPK and inactivates mTOR and its downstream effector S6K. Furthermore, hyperthermia potentiated the effect of metformin to activate AMPK and inactivate mTOR and S6K. Cell proliferation was markedly suppressed by metformin or combination of metformin and hyperthermia, which could be attributed to activation of AMPK leading to inactivation of mTOR. It is conclude that the effects of metformin against cancer cells including CSCs can be markedly enhanced by hyperthermia.

Gamma knife stereotactic radiosurgery for renal cell carcinoma and melanoma brain metastases-comparison of dose response.

BACKGROUND: Metastatic melanoma appears to have inferior local control (LC) than renal cell carcinoma (RCC) after stereotactic radiosurgery (SRS) to the brain. OBJECTIVE: To retrospectively examine RCC vs. melanoma LC dose response. METHODS: Follow-up data were available for 88 patients (RCC=38; melanoma=50) with 235 tumors (RCC=92; melanoma=143) treated with Gamma Knife SRS between Dec. 2005 to Aug. 2012. LC was compared among RCC vs. melanoma and then at each margin dose (≤18Gy, 20Gy, 22Gy, and 24Gy). Patient survival and toxicity were analyzed. Median follow-up was 9.8 months (RCC) and 5.4 months (melanoma). RESULTS: Patient characteristics were similar between RCC vs. melanoma with respect to gender, age, KPS, GPA, lesions per patient, and tumor volume. For all margin doses, LC at 6 months was 98.6% (RCC) vs. 79.2% (melanoma). When broken down by margin dose, at ≤18 Gy (P<0.0001) and 20 Gy (P=0.02), RCC had better LC compared to melanoma. At 22 Gy, LC were similar between the two histologies (P=0.19). At 24 Gy, melanoma had better LC than RCC (P=0.02). Tumor volumes were similar between RCC vs. melanoma at each margin dose (P>0.05). Small melanoma tumors (<4ml) exhibited LC dose dependence. Median survival was 16.1 months (RCC) and 9.6 months (melanoma). Toxicity was not significantly different between the two histologies and margin doses. CONCLUSIONS: RCC has significantly better LC than melanoma after SRS. Higher doses could be used for melanoma tumors <4ml to improve melanoma LC.

Genetic fuzzy classifier for sleep stage identification.

Soft-computing techniques are commonly used to detect medical phenomena and help with clinical diagnoses and treatment. In this work, we propose a design for a computerized sleep scoring method, which is based on a fuzzy classifier and a genetic algorithm (GA). We design the fuzzy classifier based on the GA using a single electroencephalogram (EEG) signal that detects differences in spectral features. Polysomnography was performed on four healthy young adults (males with a mean age of 27.5 years). The sleep classifier was designed using a sleep record and tested on the sleep records of the subjects. Our results show that the genetic fuzzy classifier (GFC) agreed with visual sleep staging approximately 84.6% of the time in detection of wakefulness (WA), shallow sleep (SS), deep sleep (DS), and rapid eye movement (REM) stages.