Non-invasive early detection of cervical carcinogenesis through the olfactory response of Caenorhabditis elegans

Abstract

Background: Human papillomavirus (HPV) is the primary cause of cervical cancer, often through the development of cervical intraepithelial neoplasia (CIN). Persistent infection with high-risk HPV types can lead to severe dysplasia and invasive cancer. Early detection of progressive cervical carcinogenesis is crucial for improving outcomes. By targeting CIN, a pre-cancerous stage, therapeutic interventions are most effective and least invasive, offering the potential to reduce the incidence of invasive cervical cancer drastically. Early cervical carcinogenesis detection is hindered by inadequate screening coverage due to barriers, inaccurate screening methods, and patient compliance issues. Optimizing the timing, frequency, and technology availability of screening in resource-limited settings also poses significant challenges. Aim: This study evaluates the efficacy of Nematode-NOSE (N-NOSE), a novel Caenorhabditis elegans-based olfactory cancer screening tool, in detecting early cervical carcinogenesis. Methods: Urine specimens from 74 patients with cervical cancer and 245 patients with CIN were analyzed, with institutional review board approval from the National Hospital Organization Shikoku Cancer Center Hospital, Ehime, Japan. Results: We discovered that urine samples from CIN patients were successfully predicted to be positive using the N-NOSE test, with a sensitivity of 73% (180/245 patients) and 100% (9/9 patients) for CIN and pre-cancerous CIN3, respectively. Conclusion: N-NOSE demonstrates high sensitivity in detecting both early-stage CIN and invasive cervical cancer, suggesting its potential as a non-invasive, urine-based screening tool for early detection. This advancement also holds the promise of significantly improving preventive healthcare by enabling timely identification and intervention, leading to more efficient treatment modalities that effectively halt the progression of cervical carcinogenesis. Relevance for patients: The N-NOSE test offers a non-invasive, urine-based method for early detection of cervical cancer and pre-cancerous changes, enabling timely intervention and potentially enhancing patient outcomes.

Keywords

Caenorhabditis elegans / Cervical carcinogenesis / N-NOSE / Olfactory response

Funding

None.

Conflict of interest

The authors are employees of HIROTSU BIO SCIENCE Inc., which developed and commercialized the N-NOSE technology described in this paper. T.H. is the CEO and Founder. The authors declare that these affiliations have not influenced the design, conduct, interpretation, or reporting of the research presented in this study.

References

[1] World Health Organization. Cervical Cancer; 2024. Available from: https://www.who.int/news-room/fact-sheets/detail/cervical-cancer [Last accessed on 2025 Feb 01].

[2] Jenkins D. A review of cross-protection against oncogenic HPV by an HPV-16/18 AS04-adjuvanted cervical cancer vaccine: Importance of virological and clinical endpoints and implications for mass vaccination in cervical cancer prevention. Gynecol Oncol. 2008; 110(3):S18-S25. doi: 10.1016/j.ygyno.2008.06.027

[3] Ahmed HG, Bensumaidea SH, Alshammari FD, et al. Prevalence of human papillomavirus subtypes 16 and 18 among yemeni patients with cervical cancer. Asian Pac J Cancer Prev. 2017; 18(6):1543-1548. doi: 10.22034/APJCP.2017.18.6.1543

[4] Lu X, Lin Q, Lin M, et al. Multiple-integrations of HPV16 genome and altered transcription of viral oncogenes and cellular genes are associated with the development of cervical cancer. PLoS One. 2014; 9(7):e97588. doi: 10.1371/journal.pone.0097588

[5] Prati B, Marangoni B, Boccardo E. Human papillomavirus and genome instability: From productive infection to cancer. Clinics (Sao Paulo). 2018; 73(Suppl 1):e539s. doi: 10.6061/clinics/2018/e539s

[6] Koshiol J, Lindsay L, Pimenta JM, Poole C, Jenkins D, Smith JS. Persistent Human papillomavirus infection and cervical neoplasia: A systematic review and meta-analysis. Am J Epidemiol. 2008; 168(2):123-137. doi: 10.1093/aje/kwn036

[7] Kusakabe M, Taguchi A, Sone K, Mori M, Osuga Y. Carcinogenesis and management of human papillomavirus-associated cervical cancer. Int J Clin Oncol. 2023; 28(8):965-974. doi: 10.1007/s10147-023-02337-7

[8] Schiffman M, Castle PE, Jeronimo J, Rodriguez AC, Wacholder S. Human papillomavirus and cervical cancer. Lancet. 2007; 370(9590):890-907. doi: 10.1016/S0140-6736(07)61416-0

[9] Alshammari AH, Ishii H, Hirotsu T, Hatakeyama H, Morishita M, Di Luccio E. Bridging the gap in cervical cancer screening for underserved communities: MCED and the promise of future technologies. Front Oncol. 2024;14:1407008. doi: 10.3389/fonc.2024.1407008

[10] Bobdey S, Sathwara J, Jain A, Balasubramaniam G. Burden of cervical cancer and role of screening in India. Indian J Med Paediatr Oncol. 2016; 37(4):278-285. doi: 10.4103/0971-5851.195751

[11] Gupta S, Nagtode N, Chandra V, Gomase K. From diagnosis to treatment: Exploring the latest management trends in cervical intraepithelial neoplasia. Cureus. 2023; 15(12):e50291. doi: 10.7759/cureus.50291

[12] Wu M, You Z. Selective applications of excisional surgical treatments of cervical precancers. Gynecol Obstet Clin Med. 2023; 3(4):198-202. doi: 10.1016/j.gocm.2023.11.003

[13] Jendrny P, Twele F, Meller S, Osterhaus ADME, Schalke E, Volk HA. Canine olfactory detection and its relevance to medical detection. BMC Infect Dis. 2021;21:1-15. doi: 10.1186/s12879-021-06523-8

[14] McCulloch M, Jezierski T, Broffman M, Hubbard A, Turner K, Janecki T. Diagnostic accuracy of canine scent detection in early-and late-stage lung and breast cancers. Integr Cancer Ther. 2006; 5(1):30-39. doi: 10.1177/1534735405285096

[15] Horvath G, Järverud GAK, Järverud S, Horváth I. Human ovarian carcinomas detected by specific odor. Integr Cancer Ther. 2008; 7(2):76-80. doi: 10.1177/153473540831905

[16] Sonoda H, Kohnoe S, Yamazato T, et al. Colorectal cancer screening with odour material by canine scent detection. Gut. 2011; 60(6):814-819. doi: 10.1136/gut.2011.242586

[17] White DA. Long-range potential scattering by Enss's method in two hilbert spaces. Trans Am Math Soc. 1986; 295(1):1-33. doi: 10.2307/2000143

[18] Milanetti E, Gosti G, De Flaviis L, et al. Investigation of the binding between olfactory receptors and odorant molecules in C. elegans organism. Biophys Chem. 2019;255:106264. doi: 10.1016/j.bpc.2019.106264

[19] Hirotsu T, Sonoda H, Uozumi T, et al. A highly accurate inclusive cancer screening test using Caenorhabditis elegans scent detection. PLoS One. 2015; 10(3):e0118699. doi: 10.1371/journal.pone.0118699

[20] Ueda Y, Kawamoto K, Konno M, et al. Application of C. elegans cancer screening test for the detection of pancreatic tumor in genetically engineered mice. Oncotarget. 2019; 10(52):5412. doi: 10.18632/oncotarget.27124

[21] Lanza E, Di Rocco M, Schwartz S, et al.C. elegans-based chemosensation strategy for the early detection of cancer metabolites in urine samples. Sci Rep. 2021; 11(1):17133. doi: 10.1038/s41598-021-96613-z

[22] Inaba S, Shimozono N, Yabuki H, et al. Accuracy evaluation of the C. elegans cancer test (N-NOSE) using a new combined method. Cancer Treat Res Commun. 2021;27:100370. doi: 10.1016/j.ctarc.2021.100370

[23] Kusumoto H, Tashiro K, Shimaoka S, et al. Efficiency of gastrointestinal cancer detection by nematode-nose (N-NOSE). In Vivo. 2020; 34(1):73-80. doi: 10.21873/invivo.11747

[24] Hatakeyama H, Morishita M, Alshammari AH, et al. A non-invasive screening method using caenorhabditis elegans for early detection of multiple cancer types: A prospective clinical study. Biochem Biophys Rep. 2024;39:101778. doi: 10.1016/j.bbrep.2024.101778

[25] Solomon D, Davey D, Kurman R, et al. The 2001 Bethesda system: Terminology for reporting results of cervical cytology. JAMA. 2002; 287(16):2114-2119. doi: 10.1001/jama.287.16.2114

[26] Bertero L, Massa F, Metovic J, et al. Eighth edition of the UICC classification of malignant tumours: An overview of the changes in the pathological tnm classification criteria-what has changed and why? Virchows Arch. 2018; 472:519-531. doi: 10.1007/s00428-017-2276-y

[27] Yoshida K, Hirotsu T, Tagawa T, et al. Odour Concentration-dependent olfactory preference change in C. elegans. Nat Commun. 2012; 3(1):739. doi: 10.1038/ncomms1750

[28] Bargmann CI, Hartwieg E, Horvitz HR. Odorant-selective genes and neurons mediate olfaction in C. elegans. Cell. 1993; 74(3):515-527. doi: 10.1016/0092-8674(93)80053-H

[29] Nygård M, Daltveit AK, Thoresen SØ, Nygård JF. Effect of an antepartum pap smear on the coverage of a cervical cancer screening programme: A population-based prospective study. BMC Health Serv Res. 2007;7:1-8. doi: 10.1186/1472-6963-7-10

[30] Chidyaonga-Maseko F, Chirwa ML, Muula AS. Underutilization of cervical cancer prevention services in low and middle income countries: A review of contributing factors. Pan Afr Med J. 2015; 21(1):231. doi: 10.11604/pamj.2015.21.231.6350

[31] Sato Y, Futamura M, Tanaka Y, et al. Clinical possibility of caenorhabditis elegans as a novel evaluation tool for esophageal cancer patients receiving chemotherapy: A prospective study. Cancers (Basel). 2023; 15(15):3870. doi: 10.3390/cancers15153870

[32] Asai A, Konno M, Ozaki M, et al. Scent test using Caenorhabditis elegans to screen for early-stage pancreatic cancer. Oncotarget. 2021; 12(17):1687. doi: 10.18632/oncotarget.28035

[33] Ungkulpasvich U, Hatakeyama H, Hirotsu T, Di Luccio E. Pancreatic cancer and detection methods. Biomedicines. 2023; 11(9):2557. doi: 10.3390/biomedicines11092557

[34] Rengarajan S, Hallem EA. Olfactory circuits and behaviors of nematodes. Curr Opin Neurobiol. 2016; 41:136-148. doi: 10.1016/j.conb.2016.09.002

[35] Varshney LR, Chen BL, Paniagua E, Hall DH, Chklovskii DB. Structural properties of the Caenorhabditis elegans neuronal network. PLoS Comput Biol. 2011; 7(2):e1001066. doi: 10.1371/journal.pcbi.1001066

[36] Bian J, Zhang H, Meng S, Liu Y. Chemotaxis of toward volatile organic compounds from induced by amino acids. J Nematol. 2018; 50(1):3-8. doi: 10.21307/jofnem-2018-003

[37] Thompson M, Sarabia Feria N, Yoshioka A, et al. A Caenorhabditis elegans behavioral assay distinguishes early stage prostate cancer patient urine from controls. Biol Open. 2021; 10(3):bio057398. doi: 10.1242/bio.057398

[38] Archiv V, Burns JS, Manda G. Metabolic pathways of the warburg effect in health and disease: Perspectives of choice, chain or chance. Int J Mol Sci. 2017; 18(12):2755. doi: 10.3390/ijms18122755

[39] Liberti MV, Locasale JW. The Warburg effect: How does it benefit cancer cells? Trends Biochem Sci. 2016; 41(3):211-218. doi: 10.1016/j.tibs.2015.12.001

[40] Troemel ER, Kimmel BE, Bargmann CI. Reprogramming chemotaxis responses: Sensory neurons define olfactory preferences in C. elegans. Cell. 1997; 91(2):161-169. doi: 10.1016/S0092-8674(00)80399-2

[41] Emery JD. The challenges of early diagnosis of cancer in general practice. Med J Aust. 2015; 203(10):391-393. doi: 10.5694/mja15.00527

[42] Cleophas TJ, Droogendijk J,Van Ouwerkerk BM. Validating diagnostic tests, correct and incorrect methods, new developments. Curr Clin Pharmacol. 2008; 3(2):70-76. doi: 10.2174/157488408784293697

[43] Shin A, Choi KS, Jun JK, et al. Validity of fecal occult blood test in the national cancer screening program, Korea. PLoS One. 2013; 8(11):e79292. doi: 10.1371/journal.pone.0079292

[44] Miles A, McClements PL, Steele RJ, Redeker C, Sevdalis N, Wardle J. The psychological impact of a colorectal cancer diagnosis following a negative fecal occult blood test result. Cancer Epidemiol Biomarkers Prev. 2015; 24(7):1032-1038. doi: 10.1158/1055-9965.EPI-15-0004

[45] Shao SH, Allen B, Clement J, et al. Multi-cancer early detection test sensitivity for cancers with and without current population-level screening options. Tumori. 2023; 109(3):335-341. doi: 10.1177/03008916221133136

[46] Fenton JJ, Barton MB, Geiger AM, et al. Screening clinical breast examination: How often does it miss lethal breast cancer? J Natl Cancer Inst Monogr. 2005; 2005(35):67-71. doi: 10.1093/jncimonographs/lgi040

[47] Imperiale TF. Quantitative immunochemical fecal occult blood tests: Is it time to go back to the future? Ann Intern Med. 2007; 146(4):309-311. doi: 10.7326/0003-4819-146-4-200702200-00013

[48] Doria-Rose VP, Kamineni A, Barrett MJ, Ko CW, Weiss NS. Case-control studies of the efficacy of screening tests that seek to prevent cancer incidence: Results of an approach that utilizes administrative claims data that do not provide information regarding test indication. Am J Epidemiol. 2019; 188(4):703-708. doi: 10.1093/aje/kwy274

[49] Zauber AG. The Impact of screening on colorectal cancer mortality and incidence: Has it really made a difference? Dig Dis Sci. 2015;60:681-691. doi: 10.1007/s10620-015-3600-5

[50] Loud JT, Murphy J. Cancer Screening and Early Detection in the 21^stCentury. Netherlands: Elsevier; 2017. p. 121-128.

[51] Dai JY, Georg Luebeck E, Chang ET, et al. Strong association between reduction of late-stage cancers and reduction of cancer-specific mortality in meta-regression of randomized screening trials across multiple cancer types. J Med Screen. 2024;31:211-222. doi: 10.1177/09691413241256744

[52] Curry SJ, Byers T, Hewitt M. Potential of screening to reduce the burden of cancer. In: Fulfilling the Potential of Cancer Prevention and Early Detection. United States: National Academies Press; 2003.

[53] Perkins R, Mitchell E. Cervical cancer disparities. J Natl Med Assoc. 2023; 115(2):S19-S25. doi: 10.1016/j.jnma.2023.02.006