ASCO / AACR / ESMO Publications
In collaboration with researchers and thought leaders around the world, OncoDiscover® uses scientific expertise and cutting-edge innovation to help drive cancer care to new frontiers. Below are key ASCO / AACR / ESMO publications.
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Journal of Clinical Oncology
Volume 40, No. 16 suppl, ASCO 2022Jayant Khandare, Gourishankar Aland, Sreeja Jayant, Pratim Chakraborti, Alain D'Souza, Avinash Kadam, Poonam Birari-Gawande, Yogesh Narayan Bendale
AIR Pvt Ltd, Pune, India; Actorius Innovations and Research, Pune, India; Actorius Innovations and Research Pvt. Ltd., Pune, India; Rasayu Biologics, Pune, India; Rasayu Cancer Clinic, Pune, India
Background: Circulating Tumor Cells (CTC) is a predictive biomarker for accounting disease progression and for minimal residual disease (MRD). Effect of conventional anticancer therapy on CTC count is well documented, however there is paucity of data related to effect of CAM based modality on CTC count in cancer patients. This study provides a preliminary observation about the effect of Ayurveda therapy on CTC count.
Methods: The retrospective study involved stratification of 72 cancer patients undergoing cancer and maintenance treatment in a non-conventional, Ayurveda cancer treatment in India. For monitoring of prognosis in cancer patients, CTC count was assesed in patients attending Rasayu Cancer clinic. 17 cancer types, namely, breast cancer, cervix and ovarian cancer, bladder, lung, Head and neck squamous carcinoma, follicular thyroid, diffuse B-cell lymphoma, Hodgkin’s and non-Hodgkin’s, colorectal, Hepatocellular, stomach with abdominal metastasis, metastatic prostate cancer, SCC with lung and skeletal metastasis etc. Total 33 (46%) males and 39 (54.1%) female patients of various types and stages were analyzed for the presence of CTCs retrospectively. CTCs were isolated and renumerated from 1.5 ml of patient’s blood sample using OncoDiscover Liquid Biopsy Technology platform enriched with anti-EpCAM antibody immunomagnetic kit, approved by Drug Controller General of India (DCGI). CTCs were confirmed for cytokeratin+ 18 (CK18), DAPI+ and CD45-. Subsequently CTCs were imaged using Zeiss Axio Observer 7 fluorescence microscope. In 28 patients (50%), CTC was accounted for at both pre and post treatment in duration of 3-6 months. 28 patients were assessed for quality of life measured by FACT-G questionnaire. The outcome was quantified for clinico-pathological parameters; age / gender, cancer types, and CTC distribution.
Results: The mean and median CTC distribution was observed to be 15.34 and 12.5, respectively. 8% patients showed the absence of any CTCs (6 subjects (1 male and 5 females). While 32 males (96%) and 34 females (87%) showed presence of CTCs. The correlation coefficient of CTC presence in male and female was significant 0.4799 (p < 0.05). The Ayurveda Rasayana therapy showed significant reduction in post interventional CTC count (-3.94±1.2) (P=0.02). In addition, this group of patients also showed significant improvement in health-related quality of life as measured by the FACT-G questionnaire (P<0.05).
Conclusions: CTC is a validated predictive biomarker for accounting minimal residual disease, both in pre and post cancer treatments. The enumeration of CTCs represents an effective prognostic biomarker in assessing the disease progression. Reduction in CTC count was seen to be associated with improvement in health-related quality of life (QoL) which needs to be investigated further to establish correlation.
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https://ascopubs.org/doi/abs/10.1200/JCO.2022.40.16_suppl.e14527 -
Journal of Clinical Oncology
Volume 40, No. 16 suppl, ASCO 2022Jayant Khandare, Alain D'Souza, Smriti Arora, Balram Singh, Gourishankar Aland, Narendra Kale, Isha Gore, Arnav Deshmukh, Rick Kamble, Vikas Jadhav, Pankaj Chaturvedi, Actorius Innovations and Research
Actorius Innovations and Research Pvt. Ltd., Pune, India; Actorius Innovations and Research, Pune, India; Actorius Innovations and Research, Simi Valley, CA; OneCell Diagnostics, Pune, India; Tata Memorial Hospital, Mumbai, India; Rasayu Biologics, Pune, India; Rasayu Cancer Clinic, Pune, India
Background: Metastatic progression accounts for nearly 90% of cancer-related deaths and has been directly correlated with the presence of circulating tumor cells (CTCs) in numerous carcinomas including breast, lung, ovarian, colorectal, and head and neck cancers. The removal of CTCs from cancer patient’s blood is directly implicated with reduction of extravasation and disease invasiveness to secondary organs.
Methods: We designed and printed 3 Dimensional (3D) microchannel devices using biocompatible polymer and packed it with anti-EpCAM (EpCAM) mediated glass-based (G) compositions (G-EpCAM). The computational fluid dynamic (CFD) analysis simulation was explored to optimize the hemodynamic effect of the G-EpCAM device for measuring the pressure and velocity difference for blood along the spiral flow microchannels. Red blood cell (RBC) hemolysis was estimated using G-EpCAM compositions packed in a device to determine optimal biocompatibility. We assessed cancer cell lines (breast cancer MCF7, lung cancer A549) interactions and capture with varying incubation time points, effect of anti-EpCAM concentrations, number of G-EpCAMs, and series of devices. We evaluated G-EpCAM-on-device’s CTC capture capability and biocompatibility using head and neck, colorectal, lung, and ductal breast cancer patient’s blood samples. All G-EpCAM captured CTCs were immuno-stained for cytokeratin 18 (CK18) expression and the optimal fluorescence acquisition intensity was quantified.
Results: Extracorporeal G-EpCAM microchannel device was 3D printed and consisted of interlocking top lid and bottom base with inlet and outlet channels. The path length of the spiral device consisted of 20 microchannels with 6.0-feet length. Device accommodated 28 gm of non-hemolytic G-EpCAM compositions. CFD analysis showed 3.8 mm as the ideal channel diameter and 2mm as the superlative G-EpCAM diameter for maximal cells and CTC capture with minimal blood hemolysis (less than 1%) as compared to control. Series 1 and 2 device indicated 90% and 85% cell capture efficiency, respectively using G-EpCAM devices indicating highest interactions and efficiency with cells. Conversely, the first device in series captured the highest cells. In addition, the efficiency improved as the number of G-EpCAM compositions was increased. We accounted device to capture CTCs with specificity having G-EpCAM composition and observed no hemolysis and non-specific interactions with other blood cells like RBCs or leukocytes.
Conclusions: Continuous CTC removal from cancer patient’s blood circulation using such device offers promising therapeutic utility in stemming aggressive metastatic invasion and progression for improving the overall survival of epithelial origin cancer patients. Clinical trial information: CTRI U1111/1192-3951.
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https://ascopubs.org/doi/abs/10.1200/JCO.2022.40.16_suppl.e14522 -
Journal of Clinical Oncology
Volume 40, No. 16 suppl, ASCO 2022Nirmal Vivek Raut, Atul Bharde, Aland Gourishankar, Sreeja Jayant, Meghana Garbhe, Sayali Gosavi, Apoorva Janorkar, Alain D'Souza, Aravindan Vasudevan, Jayant Khandare
School of Consciousness, Dr. Vishwanath Karad MIT-World Peace University, Pune, India; Savitribai Phule Pune University, Pune, India; Actorius Innovations and Research Pvt. Ltd., Pune, India; Actorius Research and innovations, Pune, India; Actorius Research and innovations Co., Cupertino, CA; Onecell Diagnostics, Cupertino, CA
Background: To analyze the role of Circulating tumor cells (CTC) as a confirmatory personalized biomarker for monitoring the disease progression, disease burden, and minimal residual disease in epithelial origin cancers.
Methods: In this retrospective study, 127 patients with colorectal, breast, and ovarian cancer at stage III and IV were analyzed. The patients were at various stages of intensive chemo and radiotherapy while the CTCs were isolated and enumerated from 1.5 ml of blood. The decision to continue chemotherapy or change to oral metronomic therapy was based on the presence of circulating tumor cells in Stage III. While in stage IV, serial measurement of CTC guided therapy. CTCs were isolated using OncoDiscover platform possessing EpCAM antibody based immunomagnetic targeting of magnetic nanoparticles after RBC lysis. CTCs were imaged and identified as CK18+ and CD45- cells showing a well-defined nucleus using a motorized fluorescence microscope operational with a monochrome camera. CTCs were enumerated using automated image analysis software and counts were expressed as number per 1.5 ml of blood.
Results: In this retrospective study we analyzed blood sample from 127 patients with the advanced stage epithelial cancers (breast- 50 %, ovarian -27 %, colorectal- 23 %) for the presence of CTCs. Amongst those, 52 % showed the presence of CTCs (breast- 52 %, ovarian -46 %, colorectal- 58 %). The CTC count ranged between 1-5 / 1.5 ml of blood with mean and median value of 2 and 1. Among the CTC positive population, majority had CTC count of 1 (44.4 %), while more than 2 CTCs were observed in 11 % of population. CTC clusters were detected in 13 % of population which predominantly were stage IV patients. 67 % among the follow up patients showed decrease in CTC count from the baseline due to the prescribed treatment, while 22 % patients showed increase in CTC count from the baseline. 11 % patients did not show change in CTC count from the baseline. When CTCs count was investigated as an independent variable to monitor the therapeutic response, it correlated well with the positive or negative outcome. In few representative cases, the reduction of CTC number from the basal value was indicative of an effective treatment. Exceptionally, in a representative colorectal cancer case, PET showed no primary as well secondary tumor burden, but the presence of CTCs in blood led further investigating an abdominal MRI that indicated multiple liver lesions suggesting micro-metastasis. Subsequent to SIRT treatment, the patient showed complete tumor regression and absence of CTCs in peripheral blood.
Conclusions: Our data suggest that CTC can serve as a dynamic intermittent biomarker for monitoring the disease progression in advanced stages and assess the therapeutic response, thus emphasizing the role of CTCs in personalized cancer management.
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https://ascopubs.org/doi/abs/10.1200/JCO.2022.40.16_suppl.e15021 -
Journal of Clinical Oncology
Volume 40, No. 16 suppl, ASCO 2022Gowhar Shafi, Aarthi Ramesh, Krithika Srinivasan, Atul Bharde, Burhanuddin Qayyumi, Gourishankar Aland, Sreeja Jayant, Alain D'Souza, Aravindan Vasudevan, Mohan Uttarwar, Pankaj Chaturvedi, Jayant Khandare
Indx Technology, Mumbai, India; School of Science, Monash University Malaysia, Malaysia, Malaysia, Malaysia; iNDX.Ai, Mumbai, India; Actorius Innovations and Research Pvt. Ltd, Pune, India; TMH, Mumbai, India; Actorius Innovations and Research, Pune, India; Actorius Innovations and Research Pvt. Ltd., Pune, India; Indx Technology, Cupertino, CA; Department of Head and Neck Surgical Oncology, Tata Memorial Hospital, Mumbai, India, Mumbai, India; AIR Pvt Ltd, Pune, India
Background: Oncology implicates highest precision using next generation diagnostics and progressive therapies assisted by predictive tools. If validated clinically, machine learning (ML) can provide better insights in precision oncology. Furthermore, it longitudinally may stratify the progression of cancer disease burden in a real time. We have developed, Circulating Tumor Cells (CTCs) driven ML model as a predictor for the treatment decision strategy for both surgery and adjuvant therapy in head and neck squamous cell carcinoma (HNSCC) patients.
Methods: In this study, a total of 380 HNSCC patients who underwent either surgery alone or surgery plus adjuvant therapy were accounted for. CTCs in patients were stratified based on clinicopathological parameters and using OncoDiscover platform having anti EpCAM antibody system regulated by the Drug Controller of India. Following this, we explored the predictive performance of the ML model on the usefulness of adjuvant therapy in HNSCC patients after the surgery. The available data was randomly divided into two subsets. First, 75%, of the original data was applied for Training the ML, and rest 25% of the data was used as a Test set. Survival curves were generated by Kaplan–Meier method and calculated through the Log rank test.
Results: XGBoost machine learning classifier was superior to Random Forest and SVM-based analyses in predicting the usefulness of adjuvant therapy post-surgery using CTC alone or in combination with other clinical parameters in HNSCC patients. Machine learning algorithms were compared for predicting the accuracy of patients stratification. The results for each model were: XGBoost model (Accuracy = 0.84, ROC value = 0.73, Kappa = 0.43); Random Forest model (Accuracy = 0.81 ROC value = 0.70, Kappa = 0.41); SVM model (Accuracy = 0.76, ROC value = 0. 69, Kappa = 0.40). The ROC value of the XGBoost model was highest (0.73) while the ROC value for the SVM model was lower (0.69). We observed that when CTCs were combined with clinicopathological parameters, the accuracy, kappa values and AUC-ROC drastically improved in predicting the usefulness of adjuvant therapy post-surgery. A similar trend was observed when CTCs were combined with clinicopathological parameters in predicting the line of chemotherapy, post-surgery.
Conclusions: ML-enabled, CTCs driven predictions can be highly accurate and ascertain the patient treatments. CTCs can be a positive predictor for selecting patient’s treatment regimen in both surgery as well in type of treatment (e.g. surgery alone or surgery + adjuvant therapy). It can also implicate to classify the patients and determine who necessitates an additional adjuvant therapy. Further investigations in this direction are necessary to predict the treatment options based on ML that may improve the overall survival of cancer patients.
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https://ascopubs.org/doi/abs/10.1200/JCO.2022.40.16_suppl.1547 -
Journal of Clinical Oncology
Volume 40, No. 16 suppl, ASCO 2022Annie Baa, Raja Pramanik, Atul Sharma, Ahitagni Biswas, Sachidanand JeeBharti, Mayank Singh, Rajeev Kumar, Aland Gourishankar, Sreeja Jayant, Alain D'Souza, Vikas Jadhav, Jayant Khandare
All India Institute of Medical Science (AIIMS), New-delhi, India; All India Institute of Medical Sciences, New Delhi, India; Actorius Innovations and Research Pvt. Ltd., Pune, India; OneCell Diagnostics, Pune, India; AIR Pvt Ltd, Pune, India
Background: Head and neck cancer is a huge burden in South East Asia with frequent relapse after curative therapy while the rest presenting in advanced unresectable stages. Financial constraints for targeted and immunotherapy make it inaccessible for the bulk of population. Thus, low-cost but efficacious regimen is highly implicated. We assessed if readily available triplet therapy of EMF, is superior in terms of extending life and maintaining quality of life along with evaluation of CTCs as a predictive biomarker in such patients.
Methods: This was a single arm, phase II, investigator initiated interventional study, wherein 35 patients were enrolled. Platinum resistant/refractory patients of HNSCC were treated with combination of erlotinib 150mg daily, methotrexate 40 mg/m2 and 5-fluourouracil 500 mg/m2 (d1, d8) q28 days till progression or unacceptable toxicities. The primary endpoint was overall response rate(ORR) at 3 months; additional endpoints were disease control rate(DCR) at 3 months, overall survival (OS), progression free survival (PFS) safety and patient reported quality of life(QOL). The role of CTCs in gauging the responders and non-responders was monitored using anti Epithelial Cell Adhesion Molecule antibody based enrichment - OncoDiscover Drug Controller General of India (DCGI) approved platform.
Results: The ORR and DCR at 3 months was 45.7% and 68.5%, respectively. The median PFS was 5 months (95%CI: 3.9-6 months) and median OS was 9 months (95%CI:7.4 -10.5 months). The 3 and 6 months PFS rates was 86 + 6% and 45 + 9%, respectively, while OS rates at 3 and 6 months were 91+ 5% and 68+ 8%, respectively. Rash, mucositis and fatigue were common adverse events occurring in 23 (65%), 14 (40%) and 9 (25.7%) respectively. The grade 3 events seen were rash in 5 (14.2%) and diarrhea in 2 (5.7%). Clinically significant improvement was seen in domains of role functioning, social functioning, fatigue, pain and global health status, swallowing, dryness of mouth and feeling ill. The mean CTC count at baseline was 0.90 + 1.1 /1.5ml of blood. Responders showed decline in levels from 1.19 + 0.25 to 0.33 + 0.48, while non-responders had increasing trend: 0.29 + 0.48 to 1+ 0.10 at 3 months (p = 0.010); with concordance rates with response being 52.9%. Additionally, CTC clearance at 3 months had numerically better PFS ̃ 6 months (95% CI: 4.72-7.72) and OS of 10 months (95% CI: 2.3-5.65) vs 4 months (95% CI: 2.3-5.65), p = 0.258 and 8 months (95% CI: 4.3-11.6), p = 0.203 in those with persistence of CTCs.
Conclusions: The triplet regimen of EMF is a feasible, safe therapeutic option with favourable response rates and improved QOL in patients with platinum resistant/refractory HNSCC. CTCs have a promising futuristic role as a predictive biomarker and can be extrapolated in clinical upfront setting too. Clinical trial information: CTRI/2020/02/023378.
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https://ascopubs.org/doi/abs/10.1200/JCO.2022.40.16_suppl.e18038
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Journal of Clinical Oncology
Volume 39, No. 15 suppl, ASCO 2021Alain D'Souza, Muhammad Mosaraf Hossain, Sreeja Jayant, Isha Gore, Pratim Chakraborti, Aland Gourishankar, Balram Singh, Smriti Arora, Swati Tripathi, Nitin Singh, Reecha Badave, Mohammad Ali Asgar Chowdhury, Rajib Kumar Shil, Shafiqul Islam, Ridwan Ahmed, Mohit Majumder, Srikanta Chowdhury, Abu Shadat Mohammod Noman, Pankaj Chaturvedi, Jayant Khandare
Actorius Innovations and Research Pvt. Ltd., Pune, India; Department of Biochemistry & Molecular Biology, University of Chittagong, Chattogram, Bangladesh; Department of Radiotherapy, Chittagong Medical College Hospital, Chittagong, Bangladesh; Department of Biochemistry and Molecular Biology, University of Chittagong, Chattogram, Bangladesh; Eukaryotic Gene Expression and Function (EuGEF) Research Group, Chattogram, Bangladesh; Department of Pathology, McGill University, Montreal, QC, Canada, Montreal, QC, Canada; Tata Memorial Hospital, Mumbai, India; Actorius Innovations and Research Co, Simi Valley, CA
Background: Tobacco consumption accounts for 1.6 million deaths annually in the South East Asia Region (SEAR). Notably, amongst 10-20% of the global population consuming the betel quid and tobacco, about 81% concentration is in SEAR regions, including India and Bangladesh. The prevalence of HNSCC in these regions is rising alarmingly. For example, HNCs account for 23% of total 156775 cancer incidences in Bangladesh. Liquid biopsy tools are unavailable and expensive for most patients in this region. However, early cancer detection using tumor biomarkers, for example, Circulating Tumor Cells (CTCs) is highly implicated. Furthermore, such biomarkers are being validated and have potential for screening of high-risk patients, such as genetic predisposition, tobacco consumption, etc. We report the first observational study in HNSCC patients in Bangladesh correlating the presence of CTCs to chronic tobacco consumption.
Methods: The study involved 70 cancer patients and 10 healthy volunteers (no prior cancer history). 87% of the patients had a specified history of chronic tobacco consumption. CTCs were isolated in 1.5 ml of blood using OncoDiscover Liquid Biopsy Test, which is clinically approved by the Drug Controller General of India, that contains an enriching anti-EPCAM antibody immunomagnetic kit. CTCs are qualified as CK18+, DAPI+and CD45-. Subsequently, CTCs were imaged using Zeiss Axio Observer 7 and quantified for Mean Fluorescence Intensity (MFI) for clinicopathological parameters; age/gender, HNSCC sub-population, and CTC distribution. Results: This is the 1st study on Bangladesh phenotype accounting for the presence of CTCs in HNSCC patients. In this population, 34 males (66%) and 10 females (52%) accounted for 91 CTCs. CTC distribution was 0 to 6 with mean and median ̃ 2.02 and 2, respectively. 25 patients (17 males, 8 females) were negative for any CTCs. Interestingly, 02 patients exhibited CTC clusters indicative of aggressive metastasis in which 01 patient had no prior tobacco usage or family cancer history. There was no correlation between CTC presence in males (66 %) and females (52 %). Healthy volunteer samples exhibited no false positives. The MFI values ranged between 23 and 766, with mean and median MFI values were 157 and 96, respectively, indicative of CK overexpression on CTCs of HNSCC patients. Conclusions: HNSCC patients with a history of chronic tobacco consumption in Bangladesh correlated the presence of CTCs in 64 % of the cases. Prospectively, CTCs may be validated as a biomarker for screening chronic tobacco users in Bangladesh to detect early cancers and HNSCC. Clinical trial information: BMRC/Grants/2018/99 (1-100).
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https://ascopubs.org/doi/10.1200/JCO.2021.39.15_suppl.e18011
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Journal of Clinical Oncology
Volume 38, No. 15 suppl, ASCO 2020Jayant Khandare, Smriti Arora, Balram Singh, Alain D'Souza, Nitin Singh, Narendra Kale, Shubham Bhide, Amrut Ashturkar, Aravindan Vasudevan, Gourishankar Aland
Actorius Innovations and Research Co, Simi Valley, CA; Actorius Innovations and Research, Pune, India; Actorius Innovations and Research Pvt. Ltd, Pune, India
Background: The presence of circulating tumor cells (CTC) in the vascular system is a tell-tale signature of metastasis in epithelial origin cancers including lung, breast, colorectal and head and neck cancers. Noteworthy, about 90% of cancer deaths are due to the progression of metastasis. Yet, cancer therapy is focussed on inhibiting tumour growth and there is a paucity of options that target metastasis. We demonstrate the POP ‘device’ that removes circulating tumour cells (CTC) from a patient’s blood to reduce the metastatic progression and improve overall survival.
Methods: We designed, multi-component glass beads enriched antibody EpCAM conjugate substrates as POP blood fluidic device. We characterized the substrate and accounted for the biocompatibility using whole blood of healthy volunteers. We evaluated, the acute toxicity of substrates using rat (Wistar Albino) whole blood (CPCSEA registration number: 941/PO/Re/S/06/CPCSEA; 31/07/2019) and further studied major histopathological tissues for any toxicity. Finally, we evaluated 06 cancer patients whole blood (1.5 mL) for capturing and for the elimination of CTCs. The captured cells were immuno-stained, and the optimal fluorescence acquisition intensity was critically quantified in accounting CK18 protein over-expression.
Results: The multi-component antibody EpCAM based substrate exhibited efficient CTC capture ability with a mean capture efficiency ranging from 40% to 100 % when compared to the OncoDiscover CTC test approved by CDSCO/ drug controller general of India (DCGI). Furthermore, the substrate indicated high biocompatibility primarily exhibited by the absence of haemolysis on whole human blood. Additionally, the preliminary animal experiments in rats showed a 100% survival rate and negligible toxicity to major organs.
Conclusions: Removal of circulating tumor cells as a therapeutics is highly implicated in improving the overall survival of epithelial cancer patients.
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https://ascopubs.org/doi/10.1200/JCO.2020.38.15_suppl.e15043 -
Journal of Clinical Oncology
Volume 38, No. 15 suppl, ASCO 2020Jayant Khandare, Burhanuddin Qayyumi, Atul Bharde, Gourishankar Aland, Sreeja Jayant, Swati Tripathi, Nitin Singh, Reecha Badave, Alain D'Souza, Balram Singh, Smriti Arora, Narendra Kale, Aravindan Vasudevan, Amrut Ashturkar, Kumar Prabhash, Pankaj Chaturvedi, Actorius
Actorius Innovations and Research Co, Simi Valley, CA; Tata Memorial Hospital, Mumbai, India; Actorius Innovations and Research Pvt. Ltd, Pune, India; Actorius innovations and research, Pune, India; Actorius Innovations and Research, Pune, India
Background: Head and neck squamous cell carcinoma is leading cancer in the India with Oral squamous cell carcinoma (OSCC) as the most frequent subtype. OSCC is classified as a locoregional disease and its increased frequency is attributed to lack of good biomarkers compared to other epithelial cancers. At the time of diagnosis, above 50% of cases present the manifestation of advanced-stage disease, and are predisposed to disease failure in spite of appropriate treatment. Thus, early diagnosis of OSCC can significantly reduce the disease burden. Here we describe regulatory approved method to establish Circulating tumor cells (CTCs) presence in OSCC Indian patients and its positive correlation with various clinicopathological parameters, suggesting the potential use of CTCs as a significant parameter to stratify oral cancer with respect to the disease advancement.
Methods: In a cross-sectional observational study, 230 OSCC patients at the different pathological stage of the disease and treatment mode were enrolled. CTCs were isolated using approved OncoDiscover liquid biopsy technology (Drug controller general of India approved), platform technology based on immunomagnetic CTC enumeration. CTCs were detected for CK18 presence and well-defined, DAPI-stained nuclei. Enumerated CTC subsequently analyzed for various clinic-pathological parameters such as pstage, extra-capsular spread (ECS), lymphovascular emboli (LVE), perineural invasion (PNI) and depth of invasion (DOI). CTC cut off values were obtained to differentiate early vs advanced stages with respect to different clinical stages and parameters.
Results: CTCs of OSCC patients correlated positively with the cancer stages (clinical as well as pathological) as well as aggressive pathological features. The presence of aggressive pathological features that often suggest the poor outcome of the disease, we observed a 25-50 % increase in CTC number. Early stage, treatment naïve patients had lower number of CTCs. Mean CTC number in advanced-stage patients was 50 % higher than early-stage OSCC patients.
Conclusions: Considering a positive correlation of CTC number with various pathophysiological features, CTC can be contemplated as a reliable parameter to predict the disease outcome in oral cancer. The consistent presence of CTC across all disease stages also suggests a probable nature of OSCC as a biological systematic disease. Clinical trial information: CTRI/2018/03/012905
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https://ascopubs.org/doi/10.1200/JCO.2020.38.15_suppl.e15541
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Clin Cancer Res (2020) 26 (11_Supplement): B30.
Jayant Khandare; Burhanuddin Nuruddin Qayyumi; Atul Bharde; Gourishankar Aland; Ajit Sagare; Swati Tripathi; Nitin Singh; Sreeja Jayant; Ashish Muglikar; Reecha Badave; Aravindan Vasudevan; Kumar Prabhash; Pankaj Chaturvedi
Objectives: To establish a rapid, highly specific, efficient, sensitive, and affordable CTC enumeration liquid biopsy technology and to validate its efficacy to isolate CTC disseminating from epithelial tumors of HNC subpopulation in India. Furthermore, to study the correlation of CTC distribution from peripheral blood with respect to various clinicopathologic factors in these patients.
Materials and Methods: A cross-sectional study was conducted using peripheral blood from enrolled 350 HNC patients. CTC were isolated using DCGI, India-approved technology, that exploits EpCAM-based immunomagnetic separation. EpCAM+ tumor cells were isolated from only 1.5 mL blood and critically assayed for cytokeratin 18 (CK18) expression and quantified using fluorescence imaging of CTCs so as to obtain a threshold to further minimize nonspecific and false-positive enumeration. CTC enumeration was subsequently subjected to statistical correlation with various clinical and pathologic parameters.
Results: We detected CTCs from all HNC patients across its various subsites, and there was a minimum threshold of at least 12 CTC in early oral cancer patients according to their clinicopathologic signatures. Compared to early oral cancer patients, advanced nodal patients showed 40% escalation in CTC count, while up to 80% increase in CTC count was observed when associated with aggressive features such as lymphovascular emboli (LVE) and with extranodal extensions. Of note, laryngopharyngeal primary had the highest mean CTC count of 33 in 1.5mL blood. Conversely, patients with advanced disease had higher CTC count and this was staggered in comparison with nearly but not all of the clinical features. Remarkably, higher clinical N (nodal) stage statistically correlated with increased CTC count and a marked increase in CTC was seen in tumors that showed lymphovascular emboli on histopathology and extranodal extension. The CTC counts were independent of parameters such as the age, sex, T stage, perineural invasion, bone involvement, or skin involvement. There was a notable trend towards reduced CTC count after chemotherapy; however, it was not statistically significant.
Conclusion: Our rapid and efficient CTC platform has demonstrated and clinically validated its use in Indian HNC phenotypes. This is the first comprehensive study to show a staggering positive correlation of CTC with various clinicopathologic factors. It comprised the largest number of oral cancer patients throughout the entire spectrum of HNSCC, the most common cancer in India. High CTC counts among HNC patients could possibly be one of the reasons for their dismal outcomes, and further studies with a correlation of CTC with patient survival in HNC are warranted. However, this study strongly implicates a perspective utility of CTC as a tumor marker in establishing the clinical staging in HNC patients.
Citation Format: Jayant Khandare, Burhanuddin Nuruddin Qayyumi, Atul Bharde, Gourishankar Aland, Ajit Sagare, Swati Tripathi, Nitin Singh, Sreeja Jayant, Ashish Muglikar, Reecha Badave, Aravindan Vasudevan, Kumar Prabhash, Pankaj Chaturvedi. Clinical correlation of circulating tumor cells as a blood marker in Indian head and neck cancer patients [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr B30..
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https://aacrjournals.org/clincancerres/article/26/11_Supplement/B30/278359/Abstract-B30-Clinical-correlation-of-circulating
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Annals of Oncology (2021) 32 (suppl_6): S1345-S1371. 10.1016/annonc/annonc740
N.V. Raut, N. Kale, A. D'souza, Y. Patil, P. Chakraborty, K. Prabhash, P. Chaturvedi, J. Khandare
Bhaktivedanta Hospital, Thane/IN, Actorius Innovations and Research, 411057 - Pune/IN, Tata Memorial Hospital, 400012 - Mumbai/IN
Background: CTCs predicts an unfavourable prognosis and outcomes in cancers. Lowering of cytokeratin 18 expression is a hallmark of epithelial mesenchymal transition (EMT). Homogeneity and validation of CK18 expression in cancer cell lines and CTCs originating from distinct solid tumors is indeterminate and may contribute to non-specific counts. We hypothesize that the expression of CK18 in varied cell lines may differ quantitatively, and additionally may exhibit similar trends in CTCs enumerated from different tumor types.
Methods: CK 18 variance in epithelial cell lines (e.g. A549 +, MCF-7 +, and MEF – (n=192269 cells) and CTCs (n=63) of different phenotypes was analyzed and compared. The fluorescence intensity was measured post immunostaining, using motorized-automated, computer-assisted scanning, and through a customized ImageJ-macro tool. The effect of anti-CK 18 concentrations (0.06-6 μg/ml) and binding constants (Kb) was measured across all cell lines. CTCs were enumerated from head and neck squamous cell carcinoma (HNSCC) patient’s blood samples. (CTRI/2018/03/012905) and from clinical samples (e.g., breast, lung, colorectal (CRC), ovarian) using clinically relevant OncoDiscover platform.
Results: CK18 mapping revealed diverse fluorescence intensities distribution in three cell lines, as well as in HNSCC, lung, breast, ovarian, and colorectal cancer CTCs (Table). In addition, the protein binding assay showed 8.65 x 10 3 K b (M -1) for MCF7 and 7.9 x 10 3 for A549 cells indicating concentration-dependent binding for CK 18 expressing proteins on cells and may be varied in CTCs of different cancer types. Compared to CK - cell line (MEF), the normalized CK18 intensity was higher by 290 % and 310 %, respectively, in MCF7 (breast) and A549 (lung) cells, demonstrating the variation in CK18 expression. On the other hand, CTCs showed significant diversity in CK 18 expression with buccal mucosa revealing the lowest (67%), while CTCs of CRC origin demonstrated the highest expression (320%) (Table). CK18 intensity representing across the cell lines and on CTCs enumerated from different cancer types.
Conclusion: Non-regulated CTC enumeration platforms pre-requisite critical validations to eliminate the non-specificity of CTC counts, which are highly imperative to clinical decisions in cancer management.
Clinical trial identification
CTRI/2018/03/012905.Legal entity responsible for the study
Actorius Innovations and Research Pvt Ltd.Disclosure
N. Kale, A. d'Souza, Y. Patil, P. Chakraborty, J. Khandare: Financial Interests, Institutional, Full or part-time Employment: Actorius Innovations and Research Pvt. Ltd. All other authors have declared no conflicts of interest.Read more
https://oncologypro.esmo.org/meeting-resources/molecular-analysis-for-precision-oncology-congress-2021/validation-of-cytokeratin-ck18-protein-expression-in-epithelial-cell-lines-and-in-circulating-tumor-cells-ctcs
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Journal of Clinical Oncology
Volume 37, No. 15 suppl, ASCO 2019Jayant Khandare, Burhanuddin Qayyumi, Kumar Prabhash, Rohit Chavan, Aland Gourishankar, Zamila Khatun, Manish Mair, Arjun Singh, Aravindan Vasudevan, Ajit Sagare, Vamshi Chidara, Atul Bharde, Pankaj Chaturvedi
Actorius Innovations and Research Pvt. Ltd, Pune, India; Tata Memorial Hospital, Mumbai, India
Background: ‘Liquid biopsy’ technologies are unaffordable and unavailable in developing countries despite having highest cancer burden and mortality rates. Current Circulating Tumor Cell (CTCs) technologies sustain clinical concerns of a) non-specificity b) low efficiency c) high blood volume requirement d) long turn-around time, and d) exorbitant cost (~$900-1400). We report, an extremely low cost, innovative nanosystem for rapid enumeration of CTS with higher specificity and efficiency.
Methods: We designed a nanosystem mediated by conjugation of anti-EpCAM through multi-reactive glutathione spacer, carbon allotrope and amine terminated dendrimer. The platform was evaluated for enhanced aqueous dispersibility and increased interaction with CTCs for rapid isolation and enumeration in 100 Head and Neck Cancer (HNC) patients having primary tumor sub-sites-oral cavity, larynx, hypopharynx, oropharynx, nasopharynx, salivary gland, and thyroid. The captured cells were immuno-stained and the optimal fluorescence acquisition intensity was validated in accounting CTCs with CK18 protein expression. There was complete elimination of the false positive normal cells (NC) count to CTCs by our method. The analysis was performed with only 1.5 ml of collected blood samples.
Results: The CTC distribution in cohort study ranged from 1 - 85 cells per 1.5 mL of blood. In more than 80% of patient’s CTCs, the quantitative estimation of anti-CK18 protein over-expression indicated ~10-fold higher intensity over to NCs. As compared to treatment naive, recurrent, and disease-free patients, the spread of CTC number across the clinical range appeared to be tight (close to mean value). The CTC enumeration sensitivity linearity was ~99.2%, and the complete enumeration process time was under 03 hours/1.5 ml of blood. Consequently, efficient, rapid and yet affordable CTC platform was designed and clinically validated.
Conclusions: ‘OncoDiscover’ liquid biopsy technology for CTC enumeration is poised to revolutionize the liquid biopsy due to its high sensitivity and affordability (~ $120) and shall resolve a major unmet medical need in impoverished world.
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https://ascopubs.org/doi/abs/10.1200/JCO.2019.37.15_suppl.e14516
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Journal of Clinical Oncology
Volume 37, No. 15 suppl, ASCO 2019Burhanuddin Qayyumi, Atul Bharde, Gourishankar Aland, Alain D'Souza, Sreeja Jayant, Nitin Singh, Swati Tripathi, Reecha Badave, Narendra Kale, Balram Singh, Smriti Arora, Isha Gore, Arjun Singh, Aravindan Vasudevan, Kumar Prabhash, Jayant Khandare, Pankaj Chaturvedi
Department of Head and Neck Surgical Oncology, Tata Memorial Hospital, Mumbai, India; Homi Bhabha National Institute, Mumbai, India.
Department of Microbiology, Savitribai Phule Pune University, Pune, India.
Actorius Innovations and Research Pvt. Ltd., Pune, India.
Actorius Innovations and Research Pvt. Ltd., Pune, India; Actorius Innovations and Research Co, Simi Valley, CA, USA; OneCell Diagnostics Inc, Cupertino, CA, USA; OneCell Diagnostics Pvt. Ltd., Pune, India. Electronic address: jayant@actorius.co.
Department of Head and Neck Surgical Oncology, Tata Memorial Hospital, Mumbai, India; Homi Bhabha National Institute, Mumbai, India. Electronic address: chaturvedi.pankaj@gmail.com.
Objective: The aim of this study was to investigate the presence of circulating tumor cells (CTCs) and their correlation with prognostic factors and clinical outcomes in treatment-naive patients with oral squamous cell carcinoma.
Study design: CTCs were isolated using OncoDiscover technique from presurgically obtained peripheral blood of 152 patients with treatment naïve oral squamous cell carcinoma. Sensitivity analysis was performed by including 40 healthy controls. CTCs cutoff values for clinicopathologic factors were obtained from receiver operating characteristic curves. Multivariate models determined the significance of CTC as independent variables. Kaplan-Meier analysis differentiated in overall survival between CTC values corresponding to the stage.
Results: Sensitivity, specificity, and accuracy of CTC detection were 94.32%, 98%, and 95.17%, respectively. Platform differentiated true positives at >3.5 CTCs (P < .00001). CTCs above 20.5 were suggestive of nodal metastasis (P < .0001) with a linear trend for detecting occult metastasis (P = .061). Early and advanced stages could be differentiated by >13.5 CTCs (P < .0001). Elevated CTCs were significantly associated with extranodal extension (>21.45 CTCs, P = .025), perineural invasion (>19.35 CTCs, P = .049), and depth of invasion (>12.5 CTCs, P = .0038). Median survival was reduced by 19 months when CTCs were >13.
Conclusions: Preoperative CTC levels demonstrated a strong correlation with adverse clinicopathology factors and suggested its role as a sensitive prognostic marker to predict survival outcome and disease progress.
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https://pubmed.ncbi.nlm.nih.gov/35595620/
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Journal of Clinical Oncology
Volume 37, No. 15 suppl, ASCO 2019Balram Singh, Smriti Arora, Alain D’Souza, Narendra Kale, Gourishankar Aland, Atul Bharde, Mohiuddin Quadir, Marcelo Calderón, Pankaj Chaturvedi and Jayant Khandare
Actorius Innovations and Research Pvt. Ltd, Pune, India
Department of Coatings and Polymeric Materials, North Dakota State University, Fargo, ND, USA
POLYMAT, Applied Chemistry Department, Faculty of Chemistry, University of the Basque Country UPV/EHU, Paseo Manuel de Lardizabal 3, 20018 Donostia-San Sebastián, Spain
IKERBASQUE, Basque Foundation for Science, 48009 Bilbao, Spain
Department of Head and Neck Surgical Oncology, Tata Memorial Hospital, Mumbai, India
School of Pharmacy, Dr Vishwanath Karad Maharashtra Institute of Technology-World Peace University, Kothrud, Pune 411038, India
School of Consciousness, Dr Vishwanath Karad Maharashtra Institute of Technology-World Peace University, Kothrud, Pune 411038, India
Abstract: Advanced materials and chemo-specific designs at the nano/micrometer-scale have ensured revolutionary progress in next-generation clinically relevant technologies. For example, isolating a rare population of cells, like circulating tumor cells (CTCs) from the blood amongst billions of other blood cells, is one of the most complex scientific challenges in cancer diagnostics. The chemical tunability for achieving this degree of exceptional specificity for extra-cellular biomarker interactions demands the utility of advanced entities and multistep reactions both in solution and in the insoluble state. Thus, this review delineates the chemo-specific substrates, chemical methods, and structure–activity relationships (SARs) of chemical platforms used for isolation and enumeration of CTCs in advancing the relevance of liquid biopsy in cancer diagnostics and disease management. We highlight the synthesis of cell-specific, tumor biomarker-based, chemo-specific substrates utilizing functionalized linkers through chemistry-based conjugation strategies. The capacity of these nano/micro substrates to enhance the cell interaction specificity and efficiency with the targeted tumor cells is detailed. Furthermore, this review accounts for the importance of CTC capture and other downstream processes involving genotypic and phenotypic CTC analysis in real-time for the detection of the early onset of metastases progression and chemotherapy treatment response, and for monitoring progression free-survival (PFS), disease-free survival (DFS), and eventually overall survival (OS) in cancer patients.
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https://pubs.rsc.org/en/content/articlelanding/2021/tb/d0tb02574g