Decellularized Umbilical Cord as a Scaffold to Support Healing of Full-Thickness Wounds.

The umbilical cord is a material that enhances regeneration and is devoid of age-related changes in the extracellular matrix (ECM). The aim of this work was to develop a biodegradable scaffold from a decellularized human umbilical cord (UC-scaffold) to heal full-thickness wounds. Decellularization was performed with 0.05% sodium dodecyl sulfate solution. The UC-scaffold was studied using morphological analysis methods. The composition of the UC-scaffold was studied using immunoblotting and Fourier transform infrared spectroscopy. The adhesion and proliferation of mesenchymal stromal cells were investigated using the LIVE/DEAD assay. The local reaction was determined by subcutaneous implantation in mice (n = 60). A model of a full-thickness skin wound in mice (n = 64) was used to assess the biological activity of the UC-scaffold. The proposed decellularization method showed its effectiveness in the umbilical cord, as it removed cells and retained a porous structure, type I and type IV collagen, TGF-ß3, VEGF, and fibronectin in the ECM. The biodegradation of the UC-scaffold in the presence of collagenase, its stability during incubation in hyaluronidase solution, and its ability to swell by 1617 ± 120% were demonstrated. Subcutaneous scaffold implantation in mice showed gradual resorption of the product in vivo without the formation of a dense connective tissue capsule. Epithelialization of the wound occurred completely in contrast to the controls. All of these data suggest a potential for the use of the UC-scaffold.

Evaluation of Approaches for the Assessment of HER2 Expression in Breast Cancer by Radionuclide Imaging Using the Scaffold Protein [99mTc]Tc-ADAPT6.

Due to its small size and high affinity binding, the engineered scaffold protein ADAPT6 is a promising targeting probe for radionuclide imaging of human epidermal growth factor receptor type 2 (HER2). In a Phase I clinical trial, [99mTc]Tc-ADAPT6 demonstrated safety, tolerability and capacity to visualize HER2 expression in primary breast cancer. In this study, we aimed to select the optimal parameters for distinguishing between breast cancers with high and low expression of HER2 using [99mTc]Tc-ADAPT6 in a planned Phase II study. HER2 expression was evaluated in primary tumours and metastatic axillary lymph nodes (mALNs). SPECT/CT imaging of twenty treatment-naive breast cancer patients was performed 2 h after injection of [99mTc]Tc-ADAPT6. The imaging data were compared with the data concerning HER2 expression obtained by immunohistochemical evaluation of samples obtained by core biopsy. Maximum Standard Uptake Values (SUVmax) afforded the best performance for both primary tumours and mALNs (areas under the receiver operating characteristic curve (ROC AUC) of 1.0 and 0.97, respectively). Lesion-to-spleen ratios provided somewhat lower performance. However, the ROC AUCs were still over 0.90 for both primary tumours and mALNs. Thus, lesion-to-spleen ratios should be further evaluated to find if these could be applied to imaging using stand-alone SPECT cameras that do not permit SUV calculations.

Humic Polyelectrolytes Facilitate Rapid Microwave Synthesis of Silver Nanoparticles Suitable for Wound-Healing Applications.

This article describes the one-pot microwave synthesis of silver nanoparticles (AgNPs) assisted with natural polyelectrolytes-humic substances (HS). The humic polyelectrolytes served both as chemical reductants for silver ions and as end-capping agents for AgNPs. Three commercially available sodium humates extracted from lignites and leonardite and one sodium fulvate isolated from natural brown water seeped through peat deposits were used in this study. The dynamics of the growth rate of AgNPs was characterised by UV-VIS spectroscopy by measuring the intensity of surface plasmon resonance at 420 nm. Transmission electron microscopy was used to characterise the size and morphology of AgNPs. Dynamic light scattering was used to determine size distributions of the synthesised AgNPs in the solutions. It was established that both conventional and microwave syntheses assisted with the coal humates produced small-size AgNPs in the range from 4 to 14 nm, with the maximum share of particles with sizes of (6 ± 2) nm by TEM estimates. The peat fulvate yielded much larger NPs with sizes from 10 to 50 nm by TEM estimates. DLS measurements revealed multimodal distributions of AgNPs stabilised with HS, which included both single NPs with the sizes from 5 to 15 nm, as well as their dominating aggregates with sizes from 20 to 200 nm and a smaller portion of extra-large aggregates up to 1000 nm. The given aggregates were loosely bound by humic polyelectrolyte, which prevented the coalescence of AgNPs into larger particles, as can be seen in the TEM images. The significant acceleration in the reaction time-a factor of 60 to 70-was achieved with the use of MW irradiation: from 240 min down to 210-240 s. The coal humate stabilised AgNPs showed antimicrobial properties in relation to S. aureus. A conclusion was made regarding the substantial advantages of microwave synthesis in the context of time and scaling up for the large-scale production of AgNP-HS preparations with antimicrobial properties suitable for external wound-healing applications.

Phase I clinical evaluation of 99mTc-labeled Affibody molecule for imaging HER2 expression in breast cancer.

The determination of tumor human epidermal growth factor receptor type 2 (HER2) status is of increasing importance with the recent approval of more efficacious HER2-targeted treatments. There is a lack of suitable methods for clinical in vivo HER2 expression assessment. Affibody molecules are small affinity proteins ideal for imaging detection of receptors, which are engineered using a small (molecular weight 6.5 kDa) nonimmunoglobulin scaffold. Labeling of Affibody molecules with positron emitters enabled the development of sensitive and specific agents for molecular imaging. The development of probes for SPECT would permit the use of Affibody-based imaging in regions where PET is not available. In this first-in-human study, we evaluated the safety, biodistribution, and dosimetry of the 99mTc-ZHER2:41071 Affibody molecule developed for SPECT/CT imaging of HER2 expression. Methods: Thirty-one patients with primary breast cancer were enrolled and divided into three cohorts (injected with 500, 1000, or 1500 µg ZHER2:41071) comprising at least five patients with high (positive) HER2 tumor expression (IHC score 3+ or 2+ and ISH positive) and five patients with low (IHC score 2+ or 1+ and ISH negative) or absent HER2 tumor expression. Patients were injected with 451 ± 71 MBq 99mTc-ZHER2:4107. Planar scintigraphy was performed after 2, 4, 6 and 24 h, and SPECT/CT imaging followed planar imaging 2, 4 and 6 h after injection. Results: Injections of 99mTc-ZHER2:41071 were well tolerated and not associated with adverse events. Normal organs with the highest accumulation were the kidney and liver. The effective dose was 0.019 ± 0.004 mSv/MBq. Injection of 1000 µg provided the best standard discrimination between HER2-positive and HER2-low or HER2-negative tumors 2 h after injection (SUVmax 16.9 ± 7.6 vs. 3.6 ± 1.4, p < 0.005). The 99mTc-ZHER2:41071 uptake in HER2-positive lymph node metastases (SUVmax 6.9 ± 2.4, n = 5) was significantly (p < 0.05) higher than that in HER2-low/negative lymph nodes (SUVmax 3.5 ± 1.2, n = 4). 99mTc-ZHER2:41071 visualized hepatic metastases in a patient with liver involvement. Conclusions: Injections of 99mTc-ZHER2:41071 appear safe and exhibit favorable dosimetry. The protein dose of 1000 µg provides the best discrimination between HER2-positive and HER2-low/negative expression of HER2 according to the definition used for current HER2-targeting drugs.

Direct Intra-Patient Comparison of Scaffold Protein-Based Tracers, [99mTc]Tc-ADAPT6 and [99mTc]Tc-(HE)3-G3, for Imaging of HER2-Positive Breast Cancer.

Previous Phase I clinical evaluations of the radiolabelled scaffold proteins [99mTc]Tc-ADAPT6 and DARPin [99mTc]Tc-(HE)3-G3 in breast cancer patients have demonstrated their safety and indicated their capability to discriminate between HER2-positive and HER2-negative tumours. The objective of this study was to compare the imaging of HER2-positive tumours in the same patients using [99mTc]Tc-ADAPT6 and [99mTc]Tc-(HE)3-G3. Eleven treatment-naïve female patients (26-65 years) with HER2-positive primary and metastatic breast cancer were included in the study. Each patient was intravenously injected with [99mTc]Tc-ADAPT6, followed by an [99mTc]Tc-(HE)3-G3 injection 3-4 days later and chest SPECT/CT was performed. All primary tumours were clearly visualized using both tracers. The uptake of [99mTc]Tc-ADAPT6 in primary tumours (SUVmax = 4.7 ± 2.1) was significantly higher (p < 0.005) than the uptake of [99mTc]Tc-(HE)3-G3 (SUVmax = 3.5 ± 1.7). There was no significant difference in primary tumour-to-contralateral site values for [99mTc]Tc-ADAPT6 (15.2 ± 7.4) and [99mTc]Tc-(HE)3-G3 (19.6 ± 12.4). All known lymph node metastases were visualized using both tracers. The uptake of [99mTc]Tc-ADAPT6 in all extrahepatic soft tissue lesions was significantly (p < 0.0004) higher than the uptake of [99mTc]Tc-(HE)3-G3. In conclusion, [99mTc]Tc-ADAPT6 and [99mTc]Tc-(HE)3-G3 are suitable for the visualization of HER2-positive breast cancer. At the selected time points, [99mTc]Tc-ADAPT6 has a significantly higher uptake in soft tissue lesions, which might be an advantage for the visualization of small metastases.

The GRPR Antagonist [99mTc]Tc-maSSS-PEG2-RM26 towards Phase I Clinical Trial: Kit Preparation, Characterization and Toxicity.

Gastrin-releasing peptide receptors (GRPRs) are overexpressed in the majority of primary prostate tumors and in prostatic lymph node and bone metastases. Several GRPR antagonists were developed for SPECT and PET imaging of prostate cancer. We previously reported a preclinical evaluation of the GRPR antagonist [99mTc]Tc-maSSS-PEG2-RM26 (based on [D-Phe6, Sta13, Leu14-NH2]BBN(6-14)) which bound to GRPR with high affinity and had a favorable biodistribution profile in tumor-bearing animal models. In this study, we aimed to prepare and test kits for prospective use in an early-phase clinical study. The kits were prepared to allow for a one-pot single-step radiolabeling with technetium-99m pertechnetate. The kit vials were tested for sterility and labeling efficacy. The radiolabeled by using the kit GRPR antagonist was evaluated in vitro for binding specificity to GRPR on PC-3 cells (GRPR-positive). In vivo, the toxicity of the kit constituents was evaluated in rats. The labeling efficacy of the kits stored at 4 °C was monitored for 18 months. The biological properties of [99mTc]Tc-maSSS-PEG2-RM26, which were obtained after this period, were examined both in vitro and in vivo. The one-pot (gluconic acid, ethylenediaminetetraacetic acid, stannous chloride, and maSSS-PEG2-RM26) single-step radiolabeling with technetium-99m was successful with high radiochemical yields (>97%) and high molar activities (16-24 MBq/nmol). The radiolabeled peptide maintained its binding properties to GRPR. The kit constituents were sterile and non-toxic when tested in living subjects. In conclusion, the prepared kit is considered safe in animal models and can be further evaluated for use in clinics.

Phase I Trial of [99mTc]Tc-maSSS-PEG2-RM26, a Bombesin Analogue Antagonistic to Gastrin-Releasing Peptide Receptors (GRPRs), for SPECT Imaging of GRPR Expression in Malignant Tumors.

The gastrin-releasing peptide receptor (GRPR) is overexpressed in prostate cancer (PCa) and in hormone-driven breast cancer (BCa). The aim of this phase I clinical trial was to evaluate safety, biodistribution, and dosimetry after the administration of the recently developed GRPR-targeting antagonistic bombesin analogue [99mTc]Tc-maSSS-PEG2-RM26 in PCa and BCa patients. Planar and whole-body SPECT/CT imaging was performed in six PCa patients and seven BCa patients 2, 4, 6, and 24 h post the intravenous administration of 40 µg of [99mTc]Tc-maSSS-PEG2-RM26 (600-700 MBq). No adverse events or pathological changes were observed. The rapid blood clearance of [99mTc]Tc-maSSS-PEG2-RM26 was observed with predominantly hepatobiliary excretion. The effective doses were 0.0053 ± 0.0007 for male patients and 0.008 ± 0.003 mSv/MBq for female patients. The accumulation of [99mTc]Tc-maSSS-PEG2-RM26 in tumors was observed in four out of six PCa and in seven out of seven BCa patients. In four BCa patients, a high uptake of the agent into the axillary lymph nodes was detected. Immunohistochemistry revealed positive GRPR expression in 60% of primary PCa, 71.4% of BCa tumors, and 50% of examined BCa lymph nodes. In conclusion, a single administration of [99mTc]Tc-maSSS-PEG2-RM26 was safe and well tolerated. [99mTc]Tc-maSSS-PEG2-RM26 SPECT may be useful for tumor detection in PCa and BCa patients, pending further studies.

A concept of "materials" diffraction and imaging beamline for SKIF: Siberian circular photon source.

Over the next decade, the extremely brilliant fourth generation synchrotron radiation sources are set to become a key driving force in materials characterization and technology development. In this study, we present a conceptual design of a versatile "Materia" diffraction and imaging beamline for a low-emittance synchrotron radiation facility. The beamline was optimized for operation with three main principal delivery regimes: parallel collimated beam ∼1 mm beam size, micro-focus regime with ∼10 µm beam spot size on the sample, and nano-focus regime with <100 nm focus. All regimes will operate in the photon energy range of 10-30 keV with the key feature of the beamline being fast switching between them, as well as between the various realizations of diffraction and imaging operation modes while maintaining the target beam position at the sample, and with both spectrally narrow and spectrally broad beams up to the energy band ΔE/E of 5 × 10-2. The manuscript presents the details of the principal characteristics selected for the insertion device and beamline optics, the materials characterization techniques, including the simulations of thermal load impact on the critical beamline optics components. Significant efforts were made to design the monochromators to mitigate the very high beam power load produced by a superconducting undulator source. The manuscript will be of interest to research groups involved in the design of new synchrotron beamlines.

Performing a radical trachelectomy with uterine transposition in a patient with stage IB2 cervical cancer: A case report.

We describe a case of a woman with invasive IB2 cervical cancer who desired to maintain fertility and required complex treatment. The suggested surgical approach with uterine transposition improves the existing radical trachelectomy procedure. Oncologic outcomes are encouraging, and no perioperative complications were noted. This report may represent a "milestone" in fertility-sparing surgeries, supporting the feasibility and safety of the opted method in stage IB2 cervical cancer with tumors about or smaller than 2 cm.

Phase I Clinical Trial Using [99mTc]Tc-1-thio-D-glucose for Diagnosis of Lymphoma Patients.

Similar to [18F]-FDG, [99mTc]Tc-1-thio-D-glucose ([99mTc]Tc-TG) also binds to GLUT receptors. The aim of this Phase I study was to evaluate the safety, biodistribution and dosimetry of [99mTc]Tc-TG. Twelve lymphoma patients were injected with 729 ± 102 MBq [99mTc]Tc-TG. Whole-body planar imaging was performed in 10 patients at 2, 4, 6 and 24 h after injection. In all 12 patients, SPECT/CT (at 2 h) and SPECT (at 4 and 6 h) imaging was performed. Vital signs and possible side effects were monitored during imaging and up to 7 days after injection. [99mTc]Tc-TG injections were well-tolerated and no side effects or alterations in blood and urine analyses data were observed. The highest absorbed dose was in the kidneys and urinary bladder wall, followed by the adrenals, prostate, bone marrow, lungs, myocardium, ovaries, uterus, liver and gall bladder wall. [99mTc]Tc-TG SPECT/CT revealed foci of high activity uptake in the lymph nodes of all nine patients with known nodal lesions. Extranodal lesions were detected in all nine cases. In one patient, a lesion in the humerus head, which was not detected by CT, was visualized using [99mTc]Tc-TG. Potentially, [99mTc]Tc-TG can be considered as an additional diagnostic method for imaging GLUT receptors in lymphoma patients.

Phase I Trial of 99mTc-(HE)3-G3, a DARPin-Based Probe for Imaging of HER2 Expression in Breast Cancer.

Radionuclide molecular imaging of human epidermal growth factor receptor type 2 (HER2) expression may enable a noninvasive discrimination between HER2-positive and HER2-negative breast cancers for stratification of patients for HER2-targeted treatments. DARPin (designed ankyrin repeat proteins) G3 is a small (molecular weight, 14 kDa) scaffold protein with picomolar affinity to HER2. The aim of this first-in-humans study was to evaluate the safety, biodistribution, and dosimetry of 99mTc-(HE)3-G3. Methods: Three cohorts of patients with primary breast cancer (each including at least 4 patients with HER2-negative and 5 patients with HER2-positive tumors) were injected with 1,000, 2,000, or 3,000 µg of 99mTc-(HE)3-G3 (287 ± 170 MBq). Whole-body planar imaging followed by SPECT was performed at 2, 4, 6, and 24 h after injection. Vital signs and possible side effects were monitored during imaging and up to 7 d after injection. Results: All injections were well tolerated. No side effects were observed. The results of blood and urine analyses did not differ before and after studies. 99mTc-(HE)3-G3 cleared rapidly from the blood. The highest uptake was detected in the kidneys and liver followed by the lungs, breasts, and small intestinal content. The hepatic uptake after injection of 2,000 or 3,000 µg was significantly (P < 0.05) lower than the uptake after injection of 1,000 µg. Effective doses did not differ significantly between cohorts (average, 0.011 ± 0.004 mSv/MBq). Tumor-to-contralateral site ratios for HER-positive tumors were significantly (P < 0.05) higher than for HER2-negative at 2 and 4 h after injection. Conclusion: Imaging of HER2 expression using 99mTc-(HE)3-G3 is safe and well tolerated and provides a low absorbed dose burden on patients. This imaging enables discernment of HER2-positive and HER2-negative breast cancer. Phase I study data justify further clinical development of 99mTc-(HE)3-G3.

Preclinical Evaluation of 99mTc-ZHER2:41071, a Second-Generation Affibody-Based HER2-Visualizing Imaging Probe with a Low Renal Uptake.

Radionuclide imaging of HER2 expression in tumours may enable stratification of patients with breast, ovarian, and gastroesophageal cancers for HER2-targeting therapies. A first-generation HER2-binding affibody molecule [99mTc]Tc-ZHER2:V2 demonstrated favorable imaging properties in preclinical studies. Thereafter, the affibody scaffold has been extensively modified, which increased its melting point, improved storage stability, and increased hydrophilicity of the surface. In this study, a second-generation affibody molecule (designated ZHER2:41071) with a new improved scaffold has been prepared and characterized. HER2-binding, biodistribution, and tumour-targeting properties of [99mTc]Tc-labelled ZHER2:41071 were investigated. These properties were compared with properties of the first-generation affibody molecules, [99mTc]Tc-ZHER2:V2 and [99mTc]Tc-ZHER2:2395. [99mTc]Tc-ZHER2:41071 bound specifically to HER2 expressing cells with an affinity of 58 ± 2 pM. The renal uptake for [99mTc]Tc-ZHER2:41071 and [99mTc]Tc-ZHER2:V2 was 25-30 fold lower when compared with [99mTc]Tc-ZHER2:2395. The uptake in tumour and kidney for [99mTc]Tc-ZHER2:41071 and [99mTc]Tc-ZHER2:V2 in SKOV-3 xenografts was similar. In conclusion, an extensive re-engineering of the scaffold did not compromise imaging properties of the affibody molecule labelled with 99mTc using a GGGC chelator. The new probe, [99mTc]Tc-ZHER2:41071 provided the best tumour-to-blood ratio compared to HER2-imaging probes for single photon emission computed tomography (SPECT) described in the literature so far. [99mTc]Tc-ZHER2:41071 is a promising candidate for further clinical translation studies.

Preclinical Evaluation of 99mTc-Labeled GRPR Antagonists maSSS/SES-PEG2-RM26 for Imaging of Prostate Cancer.

BACKGROUND: Gastrin-releasing peptide receptor (GRPR) is an important target for imaging of prostate cancer. The wide availability of single-photon emission computed tomography/computed tomography (SPECT/CT) and the generator-produced 99mTc can be utilized to facilitate the use of GRPR-targeting radiotracers for diagnostics of prostate cancers. METHODS: Synthetically produced mercaptoacetyl-Ser-Ser-Ser (maSSS)-PEG2-RM26 and mercaptoacetyl-Ser-Glu-Ser (maSES)-PEG2-RM26 (RM26 = d-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2) were radiolabeled with 99mTc and characterized in vitro using PC-3 cells and in vivo, using NMRI or PC-3 tumor bearing mice. SPECT/CT imaging and dosimetry calculations were performed for [99mTc]Tc-maSSS-PEG2-RM26. RESULTS: Peptides were radiolabeled with high yields (>98%), demonstrating GRPR specific binding and slow internalization in PC-3 cells. [99mTc]Tc-maSSS-PEG2-RM26 outperformed [99mTc]Tc-maSES-PEG2-RM26 in terms of GRPR affinity, with a lower dissociation constant (61 pM vs 849 pM) and demonstrating higher tumor uptake. [99mTc]Tc-maSSS-PEG2-RM26 had tumor-to-blood, tumor-to-muscle, and tumor-to-bone ratios of 97 ± 56, 188 ± 32, and 177 ± 79, respectively. SPECT/CT images of [99mTc]Tc-maSSS-PEG2-RM26 clearly visualized the GRPR-overexpressing tumors. The dosimetry estimated for [99mTc]Tc-maSSS-PEG2-RM26 showed the highest absorbed dose in the small intestine (1.65 × 10-3 mGy/MBq), and the effective dose is 3.49 × 10-3 mSv/MBq. CONCLUSION: The GRPR antagonist maSSS-PEG2-RM26 is a promising GRPR-targeting agent that can be radiolabeled through a single-step with the generator-produced 99mTc and used for imaging of GRPR-expressing prostate cancer.

Fertility-Sparing Surgery Using Knitted TiNi Mesh Implants and Sentinel Lymph Nodes: A 10-Year Experience.

OBJECTIVES: The aim of this cohort study is to improve the procedure of fertility-sparing surgery and to assess oncological and reproductive follow-up outcomes after radical trachelectomy (RT) for cervical cancer (T1a2-1bNxM0). METHODS: We have suggested the method combining sentinel lymph nodes (SLNs) and cervicoisthmic cerclage using a superelastic knitted TiNi mesh (KTNM) implant to facilitate the primary biomechanical/retention function of the uterus. Sixty-eight consented patients, who underwent fertility-sparing surgery using both transabdominal and laparoscopic route from 2009 through 2019, were recruited in the study and prospectively followed for a mean of 69 months. RESULTS: There were no intraoperative or postoperative complications. No cervical stenoses or mesh failures were noted in all cases. The 5-year overall and recurrence-free survival rates were 100% and 97%, respectively. Two patients indicated recurrence, it occurred in 3 and 36 months. There were 19 (28%) spontaneous pregnancies, 6 resulted in full-term delivery, whereas 2 and 11 ended in miscarriage and early abortion, respectively. CONCLUSIONS: This sparing-surgery technique is turned out to be feasible and efficient as allows to achieve well oncologic and fertility outcomes, mimicking the effect of the cervix. It complements existing surgical approaches and may provide further insight into how to overcome challenges even in aggravated cases or previously failed procedures.

Phase I Study of 99mTc-ADAPT6, a Scaffold Protein-Based Probe for Visualization of HER2 Expression in Breast Cancer.

Radionuclide molecular imaging of human epidermal growth factor receptor type 2 (HER2) expression may help to stratify breast and gastroesophageal cancer patients for HER2-targeting therapies. Albumin-binding domain-derived affinity proteins (ADAPTs) are a new type of small (46-59 amino acids) protein useful as probes for molecular imaging. The aim of this first-in-humans study was to evaluate the biodistribution, dosimetry, and safety of the HER2-specific 99mTc-ADAPT6. Methods: Twenty-nine patients with primary breast cancer were included. In 22 patients with HER2-positive (n = 11) or HER2-negative (n = 11) histopathology, an intravenous injection of 385 ± 125 MBq of 99mTc-ADAPT6 was performed, randomized to an injected protein mass of either 500 µg (n = 11) or 1,000 µg (n = 11). Planar scintigraphy followed by SPECT imaging was performed after 2, 4, 6, and 24 h. An additional cohort (n = 7) was injected with 165 ± 29 MBq (injected protein mass, 250 µg), and imaging was performed after 2 h only. Results: Injections of 99mTc-ADAPT6 were well tolerated at all mass levels and not associated with adverse effects. 99mTc-ADAPT6 cleared rapidly from the blood and most other tissues. The normal organs with the highest accumulation were the kidney, liver, and lung. Effective doses were 0.009 ± 0.002 and 0.010 ± 0.003 mSv/MBq for injected protein masses of 500 and 1,000 µg, respectively. Injection of 500 µg resulted in excellent discrimination between HER2-positive and HER2-negative tumors as early as 2 h after injection (tumor-to-contralateral breast ratio, 37 ± 19 vs. 5 ± 2; P < 0.01). The tumor-to-contralateral breast ratios for HER2-positive tumors were significantly (P < 0.05) higher for an injected mass of 500 µg than for either 250 or 1,000 µg. Conclusion: Injections of 99mTc-ADAPT6 are safe and associated with low absorbed and effective doses. A protein dose of 500 µg is preferable for discrimination between tumors with high and low expression of HER2. Further studies are justified to evaluate whether 99mTc-ADAPT6 can be used as an imaging probe to stratify patients for HER2-targeting therapy in areas where PET imaging is not readily available.

Non-Invasive Diagnosis of Malignancies Based on the Analysis of Markers in Exhaled Air.

Novel non-invasive methods for the diagnosis of malignancies should be effective for early diagnosis, reproducible, inexpensive, and independent from the human factor. Our aim was to establish the applicability of the non-invasive method, based on the analysis of air exhaled by patients who are at different stages of oropharyngeal, larynx and lung cancer. The diagnostic device includes semiconductor sensors capable of measuring the concentrations of gas components in exhaled air, with the high sensitivity of 1 ppm. The neural network uses signals from these sensors to perform classification and identify cancer patients. Prior to the diagnostic procedure of the non-invasive method, we clarified the extent and stage of the tumor according to current international standards and recommendations for the diagnosis of malignancies. The statistical dataset for neural network training and method validation included samples from 121 patients with the most common tumor localizations (lungs, oropharyngeal region and larynx). The largest number of cases (21 patients) were lung cancer, while the number of patients with oropharyngeal or laryngeal cancer varied from 1 to 9, depending on tumor localization (oropharyngeal, tongue, oral cavity, larynx and mucosa of the lower jaw). In the case of lung cancer, the parameters of the diagnostic device are determined as follows: sensitivity-95.24%, specificity-76.19%. For oropharyngeal cancer and laryngeal cancer, these parameters were 67.74% and 87.1%, respectively. This non-invasive method could lead to relevant medicinal findings and provide an opportunity for clinical utility and patient benefit upon early diagnosis of malignancies.

Cancer Diagnosis by Neural Network Analysis of Data from Semiconductor Sensors.

"Electronic nose" technology, including technical and software tools to analyze gas mixtures, is promising regarding the diagnosis of malignant neoplasms. This paper presents the research results of breath samples analysis from 59 people, including patients with a confirmed diagnosis of respiratory tract cancer. The research was carried out using a gas analytical system including a sampling device with 14 metal oxide sensors and a computer for data analysis. After digitization and preprocessing, the data were analyzed by a neural network with perceptron architecture. As a result, the accuracy of determining oncological disease was 81.85%, the sensitivity was 90.73%, and the specificity was 61.39%.

Optimal composition and position of histidine-containing tags improves biodistribution of 99mTc-labeled DARPin G3.

Radionuclide molecular imaging of HER2 expression in disseminated cancer enables stratification of patients for HER2-targeted therapies. DARPin G3, a small (14 kDa) engineered scaffold protein, is a promising probe for imaging of HER2. We hypothesized that position (C- or N-terminus) and composition (hexahistidine or (HE)3) of histidine-containing tags would influence the biodistribution of [99mTc]Tc(CO)3-labeled DARPin G3. To test the hypothesis, G3 variants containing tags at N-terminus (H6-G3 and (HE)3-G3) or at C-terminus (G3-H6 and G3-(HE)3) were labeled with [99mTc]Tc(CO)3. Labeling yield, label stability, specificity and affinity of the binding to HER2, biodistribution and tumor targeting properties of these variants were compared side-by-side. There was no substantial influence of position and composition of the tags on binding of [99mTc]Tc(CO)3-labeled variants to HER2. The specificity of HER2 targeting in vivo was confirmed. The tumor uptake in BALB/c nu/nu mice bearing SKOV3 xenografts was similar for all variants. On the opposite, there was a strong influence of the tags on uptake in normal tissues. The tumor-to-liver ratio for [99mTc]Tc(CO)3-(HE)3-G3 was three-fold higher compared to the hexahistidine-tag containing variants. Overall, [99mTc]Tc(CO)3-(HE)3-G3 variant provided the highest tumor-to-lung, tumor-to-liver, tumor-to-bone and tumor-to-muscle ratios, which should improve sensitivity of HER2 imaging in these common metastatic sites.

Comparative Evaluation of Two DARPin Variants: Effect of Affinity, Size, and Label on Tumor Targeting Properties.

Designed ankyrin repeat proteins (DARPins) are small engineered scaffold proteins that can be selected for binding to desirable molecular targets. High affinity and small size of DARPins render them promising probes for radionuclide molecular imaging. However, detailed knowledge on many factors influencing their imaging properties is still lacking. We have evaluated two human epidermal growth factor 2 (HER2)-specific DARPins with different size and binding properties. DARPins 9_29-H6 and G3-H6 were radiolabeled with iodine-125 and tricarbonyl technetium-99m and evaluated in vitro. A side-by-side comparison of biodistribution and tumor targeting was performed. HER2-specific tumor accumulation of G3-H6 was demonstrated. A combination of smaller size and higher affinity resulted in a higher tumor uptake of G3-H6 in comparison to 9_29-H6. Technetium-99m labeled G3-H6 demonstrated a better biodistribution profile than 9_29-H6, with several-fold lower uptake in liver. Radioiodinated G3-H6 showed the best tumor-to-organ ratios. The combined effect of affinity, molecular weight, scaffold composition, and nonresidualizing properties of iodine label provided radioiodinated G3-H6 with high clinical potential for imaging of HER2.

Comparative Evaluation of Radioiodine and Technetium-Labeled DARPin 9_29 for Radionuclide Molecular Imaging of HER2 Expression in Malignant Tumors.

High expression of human epidermal growth factor receptor 2 (HER2) in breast and gastroesophageal carcinomas is a predictive biomarker for treatment using HER2-targeted therapeutics (antibodies trastuzumab and pertuzumab, antibody-drug conjugate trastuzumab DM1, and tyrosine kinase inhibitor lapatinib). Radionuclide molecular imaging of HER2 expression might permit stratification of patients for HER2-targeting therapies. In this study, we evaluated a new HER2-imaging probe based on the designed ankyrin repeat protein (DARPin) 9_29. DARPin 9_29 was labeled with iodine-125 by direct radioiodination and with [99mTc]Tc(CO)3 using the C-terminal hexahistidine tag. DARPin 9_29 preserved high specificity and affinity of binding to HER2-expressing cells after labeling. Uptake of [125I]I-DARPin 9_29 and [99mTc]Tc(CO)3-DARPin 9_29 in HER2-positive SKOV-3 xenografts in mice at 6 h after injection was 3.4 ± 0.7 %ID/g and 2.9 ± 0.7 %ID/g, respectively. This was significantly (p < 0.00005) higher than the uptake of the same probes in HER2-negative Ramos lymphoma xenografts, 0.22 ± 0.09 %ID/g and 0.30 ± 0.05 %ID/g, respectively. Retention of [125I]I-DARPin 9_29 in the lung, liver, spleen, and kidneys was appreciably lower compared with [99mTc]Tc(CO)3-DARPin 9_29, which resulted in significantly (p < 0.05) higher tumor-to-organ ratios. The biodistribution data were confirmed by SPECT/CT imaging. In conclusion, radioiodine is a preferable label for DARPin 9_29.