Ranjit K. Sahu

Fourier transform infrared spectroscopy in cancer detection

The rapid developments in the field of infrared spectroscopy in the past decade have demonstrated a potential for disease diagnosis using noninvasive technologies. Several earlier studies have highlighted the advantage of using infrared spectroscopy both in the near- and mid-infrared regions for diagnostic purposes at clinical levels. The areas of focus have been the distinction of premalignant and malignant cells and tissues from their normal state using specific parameters obtained from Fourier transform infrared spectra, making it a rapid and reagent-free method. While it still requires pilot studies and designed clinical trials to ensure the applicability of such systems for cancer diagnosis, substantial progress has been made in incorporating advances in computational methods into the system to increase the sensitivity of the entire setup, making it an objective and sensitive technique suitable for automation to suit the demands of the medical community. The development of fiber-optics systems for infrared spectroscopy have further opened up new and modern avenues in medical diagnosis at various levels of cells, tissues and organs under laboratory and clinical conditions.

Distinction of cervical cancer biopsies by use of infrared microspectroscopy and probabilistic neural networks

Fourier-transform infrared spectroscopy has shown alterations of spectral characteristics of cells and tissues as a result of carcinogenesis. The research reported here focuses on the diagnosis of cancer in formalin-fixed biopsied tissue for which immunochemistry is not possible and when PAP-smear results are to be confirmed. The data from two groups of patients (a control group and a group of patients diagnosed with cervical cancer) were analyzed. It was found that the glucose/phosphate ratio decreases (by 23-49%) and the RNA/DNA ratio increases (by 38-150%) in carcinogenic compared with normal tissue. Fourier-transform microspectroscopy was used to examine these tissues. This type of study in larger populations may help to set standards or classes with which to use treated biopsied tissue to predict the possibility of cancer. Probabilistic neural networks and statistical tests as parts of these biopsies predict the possibility of cancer with a high degree of accuracy (> 95%).

Early spectral changes of cellular malignant transformation using Fourier transform infrared microspectroscopy

Fourier transform infrared microspectroscopy (FTIR-MSP) is potentially a powerful analytical method for identifying the spectral properties of biological activity in cells. The goal of the present research is the implementation of FTIR-MSP to study early spectral changes accompanying malignant transformation of cells. As a model system, cells in culture are infected by the murine sarcoma virus (MuSV), which induces malignant transformation. The spectral measurements are taken at various postinfection time intervals. To follow up systematically the progress of the spectral changes at early stages of cell transformation, it is essential first to determine and validate consistent and significant spectral parameters (biomarkers), which can evidently discriminate between normal and cancerous cells. Early stages of cell transformation are classified by an array of spectral biomarkers utilizing cluster analysis and discriminant classification function techniques. The classifications indicate that the first spectral changes are detectable much earlier than the first morphological signs of cell transformation. Our results point out that the first spectral signs of malignant transformation are observed on the first and third day of postinfection (PI) (for NIH/3T3 and MEF cell cultures, respectively), while the first visible morphological alterations are observed only on the third and seventh day, respectively. These results strongly support the potential of developing FTIR microspectroscopy as a simple, reagent-free method for early detection of malignancy.

Can Fourier transform infrared spectroscopy at higher wavenumbers "mid IR… shed light on biomarkers for carcinogenesis in tissues?

Fourier transform infrared microspectroscopy (FTIR-MSP) has shown promise as a technique for detection of abnormal cell proliferation and premalignant conditions. In the present study, we investigate the absorbance in the sensitive wavenumber region between 2800 and 3000 cm(-1), which has been known to be due to the antisymmetric and symmetric stretching vibrations of CH2 and CH3 groups of proteins and lipids. We report common biomarkers from this region that distinguish between normal and malignant tissues and cell lines. Based on our findings, we propose that the wavenumber region around 2800 to 3000 cm(-1) in the FTIR spectra of cells and tissues could provide valuable scientific evidence at the onset of premalignancy and may be used for ex vivo and in vitro detection of carcinogenesis. To further examine the utility of these markers in cancer diagnosis and management, they are tested successfully in monitoring the changes occurring in leukemia patients during chemotherapy.

Fourier transform infrared microspectroscopy as a quantitative diagnostic tool for assignment of premalignancy grading in cervical neoplasia

The early diagnosis and proper identification of cervical squamous intraepithelial lesions plays an important role in a good prognosis for the patient. However, the present practice of screening based on PAP (Papanicolaou) smear and histopathology makes it tedious and prone to human errors. We assess the validity of FTIR microspectroscopy (FTIR-MSP) of biopsies as a method to properly assign the correct stage of premalignancy in patients with symptoms of cervical intraepithelial neoplasia. For the first time we evaluate the biopsies based on the FTIR spectra for different grades of neoplasia in tandem with probabilistic neural networks (PNNs) and histopathology. The results show that the grading of neoplasia based on FTIR-MSP and a PNN differentiates the normal from premalignant with a high level of accuracy. The false positive identification of the normal as cervical intraepithelial neoplasia 1 (CIN1), CIN2, and CIN3 patients is 9.04, 0.01, and 0.01%, respectively. The false negative identification of CIN2 patients as normal and CIN1 patients is 0.01 and 4.4%, respectively. Similarly, the false negative identification of CIN3 patients as normal, CIN1, and CIN2 is 0.14, 6.99, and 9.61%, respectively. The small errors encountered in the grading are comparable to current methods, encouraging advanced studies for the development of mechanized equipment for the diagnosis and grading of premalignant cervical neoplasia.

Characteristic Absorbance of Nucleic Acids in the Mid-IR Region as Possible Common Biomarkers for Diagnosis of Malignancy

FTIR spectroscopy has been extensively used to understand the differences between normal and malignant cells and tissues. In the present study, FTIR microspectroscopy was performed on biopsies to evaluate parameters deduced from changes in nucleic acid absorbance monitored at various characteristic wavenumbers in the Mid-IR region. The data showed that there were differences in the spectra of normal and malignant tissues from several organs such as colon, cervix, skin and blood with respect to absorbance due to nucleic acids. Similar results were observed in the case of cell lines that were transformed to induce carcinogenesis. Of the several ratios examined for consistency in differentiating cancer and normal tissues, the I(996 cm−1)/I(966 cm−1) showed promise as a distinguishing parameter and was comparable to the I(1121 cm−1)/I(1020 cm−1) ratio reported in many earlier studies. The absorbance of nucleic acids is presented with an emphasis on the application of FTIR microspectroscopy for diagnosis of malignancy. Our results indicate that usage of nucleic acid absorbance yield statistically significant parameters, which could differentiate normal and cancerous tissues.

Detection of abnormal proliferation in histologically 'normal' colonic biopsies using FTIR-microspectroscopy.

Background: Abnormal crypt proliferation and development in the colon has been associated with premalignant stages of colon cancer. Conventionally, molecular markers are used to detect abnormal crypt proliferation. Methods: In the present work, feasibility studies of FTIR‐MSP to distinguish between normal and abnormal crypts from colon biopsies that show normal histopathological features have been undertaken. Results: The results indicate that abnormal crypts show deviations in the pattern of absorbance in the Mid IR region along the crypt height when compared with the normal crypts. The crypts could be empirically classified into three groups such as crypts having a normal absorbance pattern for all biochemical components, crypts with abnormal absorbance pattern for some biochemical components and crypts with completely abnormal absorbance pattern along the height for all or most biochemical components studied by FTIR. The utilization of FTIR‐MSP is proposed for diagnosis of abnormal metabolism at the molecular level of histologically completely normal‐looking crypts, especially from those biopsies that are taken from sites far away from cancer. Conclusions: This method could give rise to a reduction in false‐negative results during examination of biopsies using the conventional histopathological methods. The present method may be complementary to existing methods for precise demarcation of the zone of colostomy prior to colon cancer surgery.

Characterization of Malignant Melanoma Using Vibrational Spectroscopy

Malignant melanoma, a malignant neoplasm of epidermal melanocytes is the third most common skin cancer. In many cases, melanoma develops from nevus, which is considered as the nonmalignant stage. Fourier transform infrared microspectroscopy (FTIR-MSP), which is based on characteristic molecular vibrational spectra of cells, was used to investigate spectral differences between melanoma, nevus, and the corresponding normal epidermis. In the present work, FTIR-MSP was performed on formalin-fixed biopsies of melanoma and nevi along with the adjoining histologically normal epidermis to understand the biochemical variations from the epidermis and identify suitable parameters for differentiation of nevi from melanoma. The comparative analysis of various parameters calculated from the spectral data of the normal epidermis and the abnormal regions showed that the changes in the nucleic acids was a significant indicator of the abnormal nature of the tissues. The RNA/DNA ratio was decreased in case of both melanoma and nevus compared to the epidermis. The amide II/amide I ratio was greater for nevus and melanoma compared to the epidermis. In contrast to other organs, the analysis of carbohydrates was not found as a suitable indicator in case of malignant melanoma. Shifts in band wave number were found to be a major distinguishing feature between the melanoma and compound nevi. The present study helps in the identification of spectral features suitable for distinction of melanoma from nevus that appear similar even in FTIR spectral features and thus can pave the way for development of in vivo screening systems based on these diagnostic markers.

Nucleic acids absorbance in Mid IR and its effect on diagnostic variates during cell division: A case study with lymphoblastic cells

The differences in the absorbance in the mid infrared region (Mid IR) between normal and abnormal tissues have been shown to be a possible criterion for the detection of different forms of cancer. The present work aimed to study the effects of cell growth and DNA synthesis on the RNA/DNA ratio, which is a promising parameter for cancer diagnosis by inducing synthesis of DNA. Lymphoblastic cell lines were synchronized by methotrexate and harvested for Fourier transform infrared microscopy (FTIR-MSP) at 30-min interval after thymidine addition. The distribution of DNA in the cells/nuclei and variation of nuclear size was studied using the fluorescent dye propidium iodide (PI) and the nuclear volume was calculated from microscopy. We analyzed the ratio of RNA and DNA to quantify the relative amounts during the process by taking the ratios I(1121)/I(1020) (symmetric) and the I(1244)/I(1230) (antisymmetric) phosphate vibrations. The pattern of changes in the ratios overtime obtained in the symmetric and antisymmetric vibrations were similar though the magnitudes were different. Only a minor effect on the RNA/DNA ratio (due to nuclear volume changes) was observed. The effect due to the unfolding of DNA is greatly masked due to RNA absorbance. The effectiveness of this ratio may lie in the increased transcriptional levels in carcinogenic tissues rather than changes occurring due to unfolding and folding of DNA.

Spectral signatures of colonic malignancies in the mid-infrared region: from basic research to clinical applicability.

The process of carcinogenesis in the colon progresses through several overlapping stages, making the evaluation process challenging, as well as subjective. Owing to the complexity of colonic tissues and the search for a technique that is rapid and foolproof for precise grading and evaluation of biopsies, many spectroscopic techniques have been evaluated in the past few decades for their efficiency and clinical compatibility. Fourier-transform infrared spectroscopy, being quantitative and objective, has the capacity for automation and relevance to cancer diagnosis. This article highlights investigations on the application of Fourier-transform infrared spectroscopy (particularly microscopy) in colon cancer diagnosis and parallel developments in data analysis techniques for the characterization of spectral signatures of malignant tissues in the colon.