FLT-PET to evaluate sequencing of chemotherapies to enhance efficacy of treatment in triple negative breast cancers

Objectives: The purpose of this study is to image changes in proliferation with 3’-Deoxy-3’-[¹⁸F]fluorothymidine ([¹⁸F]-FLT) positron emission tomography (PET) to guide sequencing of combination chemotherapies [paclitaxel (PTX), doxorubicin (DRB)] in triple negative breast cancer (TNBC). Currently, sequencing of these therapies is varied between clinical locations. [¹⁸F]-FLT PET imaging has been shown to be a noninvasive, highly sensitive approach to identify changes in cell proliferation in both preclinical and clinical breast cancer and allows quantification of early signals of drug response prior to tumor size changes. Therefore, our goal is to use imaging to guide sequencing of therapies based on the alterations in cellular proliferation and heterogeneity of cellular response.

Methods: MDA-MB-231-FUCCI [GFP fluorescence (S/G2/M; 480/520nm); RFP fluorescence (G1; 560/620nm)] TNBC cells (2×10⁶) were subcutaneously injected into nude athymic mice (N=12) and randomly assigned into three groups: PTX(10mg/kg)->DRB(10mg/kg), DRB->PTX (same dosage), and saline control. [¹⁸F]-FLT PET/CT was acquired 60min post 100uL/uCi intravenous injection at day 0 (baseline), 3 and 6. Treatment occurred on day 0 and 3. GFP and RFP fluorescence were measured in vivo on day -1 and 6. In vivo cell proliferation was quantified via normalized [¹⁸F]-FLT standard uptake value (SUV) and GFP. In vivo cell viability was determined by RFP. Alterations in tumor heterogeneity were quantified by comparing [¹⁸F]-FLT SUV histograms. Statistical significance was evaluated with ANOVA and Kolmogorov-Smirnov test. Ongoing studies are measuring long-term response characteristics of variations in chemotherapy sequencing in preclinical TNBC models.

Results: [¹⁸F]-FLT PET and fluorescence imaging showed that PTX->DRB treatment significantly decreased cancer cell viability, cell proliferation and tumor heterogeneity comparing day 6 to baseline. RFP signal of control increased 14%, while DRB->PTX decreased 36%, and PTX->DRB decreased 50% (p=0.05). GFP fluorescence imaging showed DRB->PTX increased by 3-fold (p=0.0037) compared to control, while PTX->DRB was similar to control (p>0.05). [¹⁸F]-FLT PET SUV indicated that control had a 50% increase, DRB->PTX showed no increase, while PTX->DRB showed about 40% decrease (p=0.03). Tumors treated with saline and DRB->PTX showed significantly broadening of SUV values (p<0.0001 and p=0.0001, respectively) indicating increase in tumor heterogeneity. Tumor treated with PTX->DRB showed more homogenous distributions (p>0.05). Biological changes occurred prior to significant changes in tumor size between treatment groups.

Conclusions: FLT-PET may provide a role in directing sequencing of chemotherapy in TNBC. Through quantifying changes in proliferation, preliminary evidence reveals that administrating PTX prior to DRB significantly improves tumor treatment in this TNBC model by decreasing tumor cell viability, proliferation and decreasing tumor heterogeneity. Imaging allows us to visualize and quantitate cellular changes and alterations in tumor heterogeneity prior to tumor size changes and may help guide clinical decision-making for treatment of TNBC patients. Acknowledgement: We thank the American Cancer Society Grant (RSG-18-006-01-CCE) for support and the preclinical imaging shared facility, supported by NIH grant P30CA013148.