**1. Introduction**

Despite widespread awareness programs and improvements in screening, cervical cancer continues to be a major cause of morbidity and mortality amongst women in lowand middle-income states [1]. Some of the challenges related to the management include

**Citation:** Mokoala, K.M.G.; Lawal, I.O.; Maserumule, L.C.; Hlongwa, K.N.; Ndlovu, H.; Reed, J.; Bida, M.; Maes, A.; van de Wiele, C.; Mahapane, J.; et al. A Prospective Investigation of Tumor Hypoxia Imaging with 68Ga-Nitroimidazole PET/CT in Patients with Carcinoma of the Cervix Uteri and Comparison with 18F-FDG PET/CT: Correlation with Immunohistochemistry. *J. Clin. Med.* **2022**, *11*, 962. https://doi.org/ 10.3390/jcm11040962

Academic Editors: Arnoldo Piccardo and Francesco Fiz

Received: 19 December 2021 Accepted: 10 February 2022 Published: 12 February 2022

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late presentation, access to treatment and poor response to therapy [2,3]. Hypoxia is a common phenomenon linked to resistance to most forms of therapy in various malignancies including cervical cancer [4–7]. Hypoxia refers to a state of sub-physiological levels of tissue oxygen that develops due to excessive tumor growth which outgrows its blood supply and the inability of the new impaired blood vessels to keep up with the demand [8,9]. The presence or absence of hypoxia has therapeutic and prognostic implications.

Strategies to overcome hypoxia have been the subject of investigations over the years as treatment becomes more personalized. Several methods have been implemented to try and overcome hypoxia including intensity modulated radiation therapy (IMRT) which is closely linked to dose-painting, and hypoxia sensitizing drugs [10,11]. Due to the high costs and side effects of some of these therapies, it is vital to select patients that will likely benefit from these additional therapies. Therefore, detection of hypoxia in cancer lesions may help aid treatment planning.

The Eppendorf probe is considered the gold standard for hypoxia detection, however there are many reasons that have seen it fall out of favor. The analysis of genes and molecular markers that are upregulated in hypoxic states is another strategy to assess for hypoxia. These markers include hypoxia inducible factor 1 alpha (HIF-1α), vascular endothelial growth factor (VEGF) and carbonic anhydrase IX (CAIX), to name a few [12]. Other non-invasive methods have been sought and by far positron emission tomography (PET) imaging has been the most widely investigated. Fluorinated nitroimidazole derivates, specifically 18F-FMISO, were among the first tracers to be investigated for hypoxia imaging and are by far the most robust [13]. Due to its inherent limitations (slow tracer kinetics and non-specific washout) second and third generation nitroimidazole compounds such as 18F-FAZA, 18F-FETNIM and 18F-HX4 were developed [14–16]. The 60/64Cu-ATSM radionuclides were also developed and investigated as hypoxia markers [17]. The feasibility of all these tracers for hypoxia imaging in cervical cancers has been demonstrated, however some of them are not widely available. While imaging with the 18F-FDG plays a major role in oncology for staging and therapy planning, several studies have demonstrated that its role in imaging hypoxia is limited.

The chemistry of 68Ga makes for easy labelling with several peptides and molecules. With the increase in availability of the 68Ga generator this makes the 68Ga-labelled nitroimidazole derivatives attractive because 68Ga is available from a generator with a shelf life of almost a year [18]. The pre-clinical work on these tracers have demonstrated the feasibility of imaging hypoxia [19–23]. In these studies, 68Ga-Nitroimidazole compounds were more hydrophilic than the 18F-labelled radiotracers and were selectively taken up in hypoxic areas [24]. The aim of this paper was to investigate the feasibility of PET imaging of hypoxia with 68Ga-Nitroimidazole in cervical cancer lesions and to correlate imaging findings to findings on 18F-FDG PET/CT as well as immunohistochemical staining for HIF-1α.

#### **2. Materials and Methods**

#### *2.1. Patients*

Twenty women with histologically confirmed locally advanced cervical cancer were prospectively recruited into the study. The patients were enrolled consecutively from January 2020 to November 2021. The 18F-FDG PET/CT was performed as part of their workup for initial staging to plan therapy. These patients were recruited as part of an ongoing study on hypoxia imaging. Informed consent was obtained from the patients for the scan as well as to access their hospital records. In the patients that were recruited, the hemoglobin levels at the time of imaging were determined and recorded. The study was approved by the Human Research Ethics Committee of the University of Pretoria (protocol number: 691/2019). All procedures were performed in accordance with the ethical standards of the institutional research committee in alignment with the 1964 Helsinki declaration and its latter amendments.
