| 1. |
- Bragina, Olga, et al.
(författare)
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Phase I Trial of 99mTc-(HE)3-G3, a DARPin-Based Probe for Imaging of HER2 Expression in Breast Cancer
- 2022
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Ingår i: Journal of Nuclear Medicine. - : Society of Nuclear Medicine. - 0161-5505 .- 1535-5667 .- 2159-662X. ; 63:4, s. 528-535
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Tidskriftsartikel (refereegranskat)abstract
- 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.
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| 2. |
- Garousi, Javad, et al.
(författare)
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Experimental HER2-Targeted Therapy Using ADAPT6-ABD-mcDM1 in Mice Bearing SKOV3 Ovarian Cancer Xenografts : Efficacy and Selection of Companion Imaging Counterpart
- 2022
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Ingår i: Pharmaceutics. - : MDPI. - 1999-4923. ; 14:8
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Tidskriftsartikel (refereegranskat)abstract
- Overexpression of the human epidermal growth factor receptor 2 (HER2) in breast and gastric cancer is exploited for targeted therapy using monoclonal antibodies and antibody-drug conjugates. Small engineered scaffold proteins, such as the albumin binding domain (ABD) derived affinity proteins (ADAPTs), are a promising new format of targeting probes for development of drug conjugates with well-defined structure and tunable pharmacokinetics. Radiolabeled ADAPT6 has shown excellent tumor-targeting properties in clinical trials. Recently, we developed a drug conjugate based on the HER2-targeting ADAPT6 fused to an albumin binding domain (ABD) for increased bioavailability and conjugated to DM1 for cytotoxic action, designated as ADAPT6-ABD-mcDM1. In this study, we investigated the therapeutic efficacy of this conjugate in mice bearing HER2-expressing SKOV3 ovarian cancer xenografts. A secondary aim was to evaluate several formats of imaging probes for visualization of HER2 expression in tumors. Administration of ADAPT6-ABD-mcDM1 provided a significant delay of tumor growth and increased the median survival of the mice, in comparison with both a non-targeting homologous construct (ADAPT(Neg)-ABD-mcDM1) and the vehicle-treated groups, without inducing toxicity to liver or kidneys. Moreover, the evaluation of imaging probes showed that small scaffold proteins, such as Tc-99m(CO)(3)-ADAPT6 or the affibody molecule Tc-99m-Z(HER2:41071), are well suited as diagnostic companions for potential stratification of patients for ADAPT6-ABD-mcDM1-based therapy.
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| 3. |
- Larkina, Mariia, et al.
(författare)
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Comparative Preclinical Evaluation of Peptide-Based Chelators for the Labeling of DARPin G3 with 99mTc for Radionuclide Imaging of HER2 Expression in Cancer
- 2022
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Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 23:21, s. 13443-
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Tidskriftsartikel (refereegranskat)abstract
- Non-invasive radionuclide imaging of human epidermal growth factor receptor type 2 (HER2) expression in breast, gastroesophageal, and ovarian cancers may stratify patients for treatment using HER2-targeted therapeutics. Designed ankyrin repeat proteins (DARPins) are a promising type of targeting probe for radionuclide imaging. In clinical studies, the DARPin [Tc-99m]Tc-(HE)(3)-G3 labeled using a peptide-based chelator His-Glu-His-Glu-His-Glu ((HE)(3)), provided clear imaging of HER2 expressing breast cancer 2-4 h after injection. The goal of this study was to evaluate if the use of cysteine-containing peptide-based chelators Glu-Glu-Glu-Cys (E3C), Gly-Gly-Gly-Cys (G(3)C), and Gly-Gly-Gly-Ser-Cys connected via a (Gly-Gly-Gly-Ser)(3)-linker (designated as G3-(G(3)S)(3)C) would further improve the contrast of imaging using Tc-99m-labeled derivatives of G3. The labeling of the new variants of G3 provided a radiochemical yield of over 95%. Labeled G3 variants bound specifically to human HER2-expressing cancer cell lines with affinities in the range of 1.9-5 nM. Biodistribution of [Tc-99m]Tc-G3-G(3)C, [Tc-99m]Tc-G3-(G(3)S)(3)C, and [Tc-99m]Tc-G3-E3C in mice was compared with the biodistribution of [Tc-99m]Tc-(HE)(3)-G3. It was found that the novel variants provide specific accumulation in HER2-expressing human xenografts and enable discrimination between tumors with high and low HER2 expression. However, [Tc-99m]Tc-(HE)(3)-G3 provided better contrast between tumors and the most frequent metastatic sites of HER2-expressing cancers and is therefore more suitable for clinical applications.
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| 4. |
- Liu, Yongsheng, et al.
(författare)
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Biologic Evaluation of a Heterodimeric HER2-Albumin Targeted Affibody Molecule Produced by Chemo-Enzymatic Peptide Synthesis
- 2022
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Ingår i: Pharmaceutics. - : MDPI. - 1999-4923. ; 14:11
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Tidskriftsartikel (refereegranskat)abstract
- Targeted molecular radiation therapy is a promising emerging treatment modality in oncology, and peptide synthesis may shorten the time to reach the clinical stage. In this study, we have explored Chemo-Enzymatic Peptide Synthesis, or CEPS, as a new means of producing a therapeutic HER2 targeted Affibody((R)) molecule, comprising a C-terminal albumin binding domain (ABD) for half-life extension and a total length of 108 amino acids. In addition, a DOTA moiety could be incorporated at N-terminus directly during the synthesis step and subsequently utilized for site-specific radiolabeling with the therapeutic radionuclide Lu-177. Retained thermodynamic stability as well as retained binding to both HER2 and albumin was verified. Furthermore, HER2 binding specificity of the radiolabeled Affibody molecule was confirmed by an in vitro saturation assay showing a significantly higher cell-bound activity of SKOV-3 (high HER2 expression) compared with BxPC3 (low HER2 expression), both in the presence and absence of HSA. In vivo evaluation in mice bearing HER2 expressing xenografts also showed specific tumor targeting as well as extended time in circulation and reduced kidney uptake compared with a HER2 targeted Affibody molecule without the ABD moiety. To conclude, we have demonstrated that CEPS can be used for production of Affibody-fusion molecules with retained in vitro and in vivo functionality.
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| 5. |
- Liu, Yongsheng, et al.
(författare)
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Experimental Therapy of HER2-Expressing Xenografts Using the Second-Generation HER2-Targeting Affibody Molecule 188Re-ZHER2:41071
- 2022
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Ingår i: Pharmaceutics. - : MDPI. - 1999-4923. ; 14:5
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Tidskriftsartikel (refereegranskat)abstract
- HER2-targeted radionuclide therapy might be helpful for the treatment of breast, gastric, and ovarian cancers which have developed resistance to antibody and antibody-drug conjugate-based therapies despite preserved high HER2-expression. Affibody molecules are small targeting proteins based on a non-immunoglobulin scaffold. The goal of this study was to test in an animal model a hypothesis that the second-generation HER2-targeting Affibody molecule 188Re-ZHER2:41071 might be useful for treatment of HER2-expressing malignant tumors. ZHER2:41071 was efficiently labeled with a beta-emitting radionuclide rhenium-188 (188Re). 188Re-ZHER2:41071 demonstrated preserved specificity and high affinity (KD = 5 ± 3 pM) of binding to HER2-expressing cells. In vivo studies demonstrated rapid washout of 188Re from kidneys. The uptake in HER2-expressing SKOV-3 xenografts was HER2-specific and significantly exceeded the renal uptake 4 h after injection and later. The median survival of mice, which were treated by three injections of 16 MBq 188Re-ZHER2:41071 was 68 days, which was significantly longer (<0.0001 in the log-rank Mantel-Cox test) than survival of mice in the control groups treated with vehicle (29 days) or unlabeled ZHER2:41071 (27.5 days). In conclusion, the experimental radionuclide therapy using 188Re-ZHER2:41071 enabled enhancement of survival of mice with human tumors without toxicity to the kidneys, which is the critical organ.
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| 6. |
- Liu, Yongsheng, et al.
(författare)
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Preclinical Evaluation of a New Format of Ga-68- and In-111-Labeled Affibody Molecule Z(IGF-1R:4551) for the Visualization of IGF-1R Expression in Malignant Tumors Using PET and SPECT
- 2022
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Ingår i: Pharmaceutics. - : MDPI AG. - 1999-4923. ; 14:7
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Tidskriftsartikel (refereegranskat)abstract
- The Insulin-like growth factor-1 receptor (IGF-1R) is a molecular target for several monoclonal antibodies undergoing clinical evaluation as anticancer therapeutics. The non-invasive detection of IGF-1R expression in tumors might enable stratification of patients for specific treatment and improve the outcome of both clinical trials and routine treatment. The affibody molecule Z(IGF-1R:4551) binds specifically to IGF-1R with subnanomolar affinity. The goal of this study was to evaluate the Ga-68 and In-111-labeled affibody construct NODAGA-(HE)(3)-Z(IGF-1R:4551) for the imaging of IGF-1R expression, using PET and SPECT. The labeling was efficient and provided stable coupling of both radionuclides. The two imaging probes, [Ga-68]Ga-NODAGA-(HE)(3)-Z(IGF-1R:4551) and [In-111]In-NODAGA-(HE)(3)-Z(IGF-1R:4551), demonstrated specific binding to IGF-1R-expressing human cancer cell lines in vitro and to IGF-1R-expressing xenografts in mice. Preclinical PET and SPECT/CT imaging demonstrated visualization of IGF-1R-expressing xenografts already one hour after injection. The tumor-to-blood ratios at 3 h after injection were 7.8 +/- 0.2 and 8.0 +/- 0.6 for [Ga-68]Ga-NODAGA-(HE)(3)-Z(IGF-1R:4551) and [In-111]In-NODAGA-(HE)(3)-Z(IGF-1R:4551), respectively. In conclusion, a molecular design of the Z(IGF-1R:4551) affibody molecule, including placement of a (HE)(3)-tag on the N-terminus and site-specific coupling of a NODAGA chelator on the C-terminus, provides a tracer with improved imaging properties for visualization of IGF-1R in malignant tumors, using PET and SPECT.
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| 7. |
- Oroujeni, Maryam, PhD, 1982-, et al.
(författare)
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Affibody-Mediated PNA-Based Pretargeted Cotreatment Improves Survival of Trastuzumab-Treated Mice Bearing HER2-Expressing Xenografts
- 2022
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Ingår i: Journal of Nuclear Medicine. - : Society of Nuclear Medicine. - 0161-5505 .- 1535-5667 .- 2159-662X. ; 63:7, s. 1046-1051
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Tidskriftsartikel (refereegranskat)abstract
- Treatment of patients with human epidermal growth factor receptor 2 (HER2)-expressing tumors using the monoclonal antibody trastuzumab increases survival. The Affibody-based peptide nucleic acid (PNA)-mediated pretargeted radionuclide therapy has demonstrated efficacy against HER2-expressing xenografts in mice. Structural studies suggest that Affibody molecules and trastuzumab bind to different epitopes on HER2. The aim of this study was to test the hypothesis that a combination of PNA-mediated pretargeted radionuclide therapy and trastuzumab treatment of HER2-expressing xenografts can extend survival compared with monotherapies. Methods: Mutual interference of the primary pretargeting probe Z(HER2:342)-SR-HP1 and trastuzumab in binding to HER2-expressing cell lines was investigated in vitro. Experimental therapy evaluated the survival of mice bearing HER2-expressing SKOV-3 xenografts after treatment with vehicle, trastuzumab only, pretargeting using Affibody-PNA chimera Z(HER2:342)-SR-HP1 and complementary probe Lu-177-HP2, and combination of trastuzumab and pretargeting. The ethical permit limited the study to 90 d. The animals'weightsweremonitored during the study. After study termination, samples of liver and kidneys were evaluated by a veterinary pathologist for toxicity signs. Results: The presence of a large molar excess of trastuzumab had no influence on the affinity of Z(HER2:342)-SR-HP1 binding to HER2-expressing cells in vitro. The affinity of trastuzumab was not affected by a large excess of Z(HER2:342)-SR-HP1. Themedian survival of mice treated with trastuzumab (75.5 d) was significantly longer than the survival of mice treated with a vehicle (59.5 d). Median survival of mice treated with pretargeting was not reached by day 90. Six mice of 10 in this group survived, and 2 had complete remission. All mice in the combination treatment group survived, and tumors in 7 mice had disappeared at study termination. There was no significant difference between animal weights in the different treatment groups. No significant pathologic alterations were detected in livers and kidneys of treated animals. Conclusion: Treatment of mice bearing HER2-expressing xenografts with the combination of trastuzumab and Affibody-mediated PNA-based radionuclide pretargeting significantly increased survival compared with monotherapies. Cotreatment was not toxic for normal tissues.
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| 8. |
- Rinne, Sara Sophie
(författare)
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Affibody-Based Molecular Imaging and Targeted Therapy of HER3-Expressing Cancer
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The human epidermal growth factor receptor type 3 (HER3) is overexpressed in different types of cancer and is a known contributor to disease progression and resistance to cancer therapy. This thesis is based on five original articles, which aimed to improve the diagnostic and therapeutic potential of affibody-based agents for management of HER3-expressing cancers. Papers I-III focused on the development and optimization of radiolabeled affibody molecules for radionuclide molecular imaging of HER3 expression. In particular, they investigated the influence of different radiometal/chelator complexes and hydrophilicity on the biodistribution and imaging properties of the HER3-targeting affibody molecule ZHER3. Paper IV compared the optimized ZHER3-based radiotracer with antibody and antibody-fragment based radiotracers for PET imaging of HER3 expression. In Paper V, a preclinical therapy study was conducted to investigate the efficacy of different monomeric and dimeric HER3-targeting affibody constructs for treatment of HER3-expressing cancer.It was shown that by optimizing the radiometal/chelator complex and incorporation of a hydrophilic (HE)3-tag the imaging properties of ZHER3-based radiotracers could be improved (Papers I-III). Generally, replacing a positively charged radiometal/chelator complex with a neutral or negatively charged complex improved the image contrast by reducing the normal organ uptake, especially in the liver. Further, it was demonstrated that the optimized affibody-based tracer [68Ga]Ga-(HE)3-ZHER3-NODAGA could provide higher contrast PET images of HER3 expression than the 89Zr-labeled antibody seribantumab and a seribantumab-derived F(ab’)2 fragment (Paper IV). The therapy study showed that the arrangement of the molecular building blocks affected the therapeutic efficacy of ZHER3-based affibody constructs. The monomeric and dimeric ABD-conjugated affibody constructs 3A and 3A3 showed the best therapeutic efficacy among the tested constructs and were able to delay tumor growth and prolong survival with the same efficacy as the therapeutic HER3-targeting antibody seribantumab (Paper V).In conclusion, the results described in this thesis show that HER3-targeting affibody-based agents could be well-suited for molecular imaging of HER3 expression and HER3-targeted therapy in cancer. Careful optimization of the molecular design could improve the imaging properties and therapeutic efficacy of HER3-targeting affibody molecules. Most importantly, it was demonstrated that HER3-targeting affibody molecules could provide superior diagnostic images and similar therapeutic effect than more traditional approaches for management of HER3-expressing cancer.
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| 9. |
- Rinne, Sara S., et al.
(författare)
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Targeting Tumor Cells Overexpressing the Human Epidermal Growth Factor Receptor 3 with Potent Drug Conjugates Based on Affibody Molecules
- 2022
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Ingår i: Biomedicines. - : MDPI AG. - 2227-9059. ; 10:6
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Tidskriftsartikel (refereegranskat)abstract
- Increasing evidence suggests that therapy targeting the human epidermal growth factor receptor 3 (HER3) could be a viable route for targeted cancer therapy. Here, we studied a novel drug conjugate, ZHER3-ABD-mcDM1, consisting of a HER3-targeting affibody molecule, coupled to the cytotoxic tubulin polymerization inhibitor DM1, and an albumin-binding domain for in vivo half-life extension. ZHER3-ABD-mcDM1 showed a strong affinity to the extracellular domain of HER3 (K-D 6 nM), and an even stronger affinity (KD 0.2 nM) to the HER3-overexpressing pancreatic carcinoma cell line, BxPC-3. The drug conjugate showed a potent cytotoxic effect on BxPC-3 cells with an IC50 value of 7 nM. Evaluation of a radiolabeled version, [99mTc]Tc-ZHER3-ABD-mcDM1, showed a relatively high rate of internalization, with a 27% internalized fraction after 8 h. Further in vivo evaluation showed that it could target BxPC-3 (pancreatic carcinoma) and DU145 (prostate carcinoma) xenografts in mice, with an uptake peaking at 6.3 +/- 0.4% IA/g at 6 h post-injection for the BxPC-3 xenografts. The general biodistribution showed uptake in the liver, lung, salivary gland, stomach, and small intestine, organs known to express murine ErbB3 naturally. The results from the study show that ZHER3-ABD-mcDM1 is a highly potent and selective drug conjugate with the ability to specifically target HER3 overexpressing cells. Further pre-clinical and clinical development is discussed.
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| 10. |
- Tolmachev, Vladimir, et al.
(författare)
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Direct In Vivo Comparison of Tc-99m-Labeled Scaffold Proteins, DARPin G3 and ADAPT6, for Visualization of HER2 Expression and Monitoring of Early Response for Trastuzumab Therapy
- 2022
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Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 23:23
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Tidskriftsartikel (refereegranskat)abstract
- Non-invasive radionuclide molecular visualization of human epidermal growth factor receptor type 2 (HER2) can provide stratification of patients for HER2-targeting therapy. This method can also enable monitoring of the response to such therapies, thereby making treatment personalized and more efficient. Clinical evaluation in a phase I study demonstrated that injections of two scaffold protein-based imaging probes, [Tc-99m]Tc-(HE)(3)-G3 and [Tc-99m]Tc-ADAPT6, are safe, well-tolerated and cause a low level of radioactivity in healthy tissue. The goal of this preclinical study was to select the best probe for stratification of patients and response monitoring. Biodistribution of both tracers was compared in mice bearing SKOV-3 xenografts with high HER2 expression or MDA-MB-468 xenografts with very low expression. Changes in accumulation of the probes in SKOV-3 tumors 24 h after injection of trastuzumab were evaluated. Both [Tc-99m]Tc-ADAPT6 and [Tc-99m]Tc-(HE)(3)-G3 permitted high contrast imaging of HER2-expressing tumors and a clear discrimination between tumors with high and low HER2 expression. However, [Tc-99m]Tc-ADAPT6 has better preconditions for higher sensitivity and specificity of stratification. On the other hand, [Tc-99m]Tc-(HE)(3)-G3 is capable of detecting the decrease of HER2 expression on response to trastuzumab therapy only 24 h after injection of the loading dose. This indicates that the [Tc-99m]Tc-(HE)(3)-G3 tracer would be better for monitoring early response to such treatment. The results of this study should be considered in planning of further clinical development of HER2 imaging probes.
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