Radionuclides transform chemotherapeutics into phototherapeutics for precise treatment of disseminated cancer

Abstract

Most cancer patients succumb to disseminated disease because conventional systemic therapies lack spatiotemporal control of their toxic effects in vivo, particularly in a complicated milieu such as bone marrow where progenitor stem cells reside. Here, we demonstrate the treatment of disseminated cancer by photoactivatable drugs using radiopharmaceuticals. An orthogonal-targeting strategy and a contact-facilitated nanomicelle technology enabled highly selective delivery and co-localization of titanocene and radiolabelled fluorodeoxyglucose in disseminated multiple myeloma cells. Selective ablation of the cancer cells was achieved without significant off-target toxicity to the resident stem cells. Genomic, proteomic and multimodal imaging analyses revealed that the downregulation of CD49d, one of the dimeric protein targets of the nanomicelles, caused therapy resistance in small clusters of cancer cells. Similar treatment of a highly metastatic breast cancer model using human serum albumin-titanocene formulation significantly inhibited cancer growth. This strategy expands the use of phototherapy for treating previously inaccessible metastatic disease.

Conflict of interest statement

The authors declare no competing financial interests.

Figures

Fig. 1

Orthogonal cancer targeting strategy using nanomicelles. a Schematic of the process of photoactivation of Titanocene in disseminated cancer cells in the bone marrow microenvironment. The various phases are numbered: 1. Administration of targeted NM-TC; 2. The targeted NM enter the bone marrow from the vasculature and bind to α4β1 receptor on the cancer cells and subsequently deliver the drug to the cell; 3. Administration of radiopharmaceuticals (18FDG), which is typically 1.5–2 h after phase (1); 4. 18FDG enters the cancer cells through the overexpressed Glut transporters on cancer cells; 5. Once the drug and radiopharmaceutical are co-localized in the cancer cells, the former is photoactivated by the latter through CR leading to cell death (6). Notice that since the other vital cells in the bone marrow, such as stem cells and stromal cells, do not express the combination of α4β1 and glut receptors essential for the treatment to work, they would largely remain unaffected causing minimal off-target toxicity. b Schematic of phospholipid NM with VLA-4 homing ligands. c TEM image of micelles alone. Scale bar, 100 nm. Inset: single micelle. Scale bar, 10 nm. d Schematic of phospholipid NM encapsulating TC with VLA-4 homing ligands. e TEM image of micelle incorporated with TC in the membrane. Scale bar, 100 nm. Inset: single NM-TC. Scale bar, 10 nm

Fig. 2

Monitoring nanomicelles biodistribution and spread of multiple myeloma in vivo. a Pharmacokinetics of NM-TC in rats using coupled plasma optical emission spectrometry. Half-life is 123.4 min. b Comparison of biodistribution in mice of targeted NM-TC and pristine TC in vivo showing highest uptake and retention in bones and spleen, characteristic of multiple myeloma, 2 h post injection. 18FDG-PET images showing increased uptake of 18FDG in mouse forelimbs, spine, and hind limbs of mice with multiple myeloma (c, e, g) compared to naive mice (d, f, h, i). Comparison of standard uptake values (SUV) of 18FDG in multiple myeloma vs. naive mice in various bones. Values are means ± s.e.m. *P < 0.05, **P < 0.01. n = 5 mice for each of the pharmacokinetics study in rats; and biodistribution study in mice

Fig. 3

Response of multiple myeloma to CRIT. a Timeline of treatment. b Bioluminescence imaging of representative multiple myeloma-bearing mice in different treatment groups—untreated, 18FDG, NM controls and CRIT. All images are dorsal images and on the same scale. The images of control groups appear saturated on week 6 in comparison to CRIT. c Change in bioluminescence intensity as a result of treatment compared to untreated control. The intensity consistently remains lower than untreated controls during the treatment and beyond. d Comparison of survival of different treatment groups showing a twofold increase in survival in treated mice compared to control groups. **P < 0.01. e18FDG-PET images of MM mice before and after treatment showing lower tumour burden in the latter. F: frontal view, S: sagittal view. Boxes denote tumour region. f SUV values of the treatment group were lower than untreated controls. **P < 0.01. n = 15 mice for CRIT, n = 10 mice for untreated control and n = 5 mice for NM-TC alone and 18FDG alone treated mice

Fig. 4

CRIT selects for CD49d cells in MM model. a Bioluminescence intensity plot showing resistant nature of MM cells extracted from treated cohort (MM1CRIT-RES) upon rechallenging with CRIT in fresh mice. b No difference in GLUT1 mRNA expression was observed between parental MM1.S cells and resistant MM1.SCRIT-RES cells as assessed by qRT-PCR. ns not significant. c–h No difference in expression of CD29 was observed between MM1.S cells (c) or MM1.SCRIT-RES cells (d) following treatment with CRIT in vivo. MM1.S stopped responding to CRIT by downregulating expression of VLA-4 subunit CD49d (Resistance = 28.12% CD49d+) (f) relative to parental cells injected into mice at the beginning of the experiment (parental = 99.92% CD49d+) (e), resulting in reduced binding of the VLA-4-targeting ligand LLP2A on resistant cells (h) (LLP2A+ = 6.6%) compared to parental MM1.S (g) (LLP2A+= 84.15%). i No significant difference in colony-forming units of progenitor stem cells was observed between untreated, control and treated mice. j To determine if CRIT reduced engraftment of haematologic cells in vivo, we assessed BM repopulation following CRIT treatment. Bone marrow from treated mice or PBS-treated controls were mixed with congenic B6.CD45.1/2 at a ratio of 1:1 before infusion of 1 × 106 total BM cells into lethally irradiated (TBI 1100 cGy) B6.CD45.1 recipients. k Percentage of cells derived from treated donor BM (CD45.2) were calculated as a percentage of total donor BM (CD45.2 + CD45.1/2). BM from CRIT-treated mice effectively repopulated recipients (n = 5 per group)

Fig. 5

Dissemination of metastatic breast cancer cells and biodistribution of TC in mice. a In vivo BLI non-tumour-bearing C57BL/6J mice. b In vivo BLI of metastatic breast cancer 10 days post intracardiac injection of PyMT-BO1 GFP/Luc in C57BL/6J mice. c Ex vivo BLI of metastatic tumour burden 10 days post tumour initiation. The left panel are tissues obtained from a mouse and the right panel are tissues obtained from b mouse. d Inductively coupled plasma mass spectrometry analysis of Ti content in blood samples and the lower limbs, where tumour burden is high. The Ti content was background-corrected from untreated mice; *P < 0.05. Studies were performed with n = 5 mice per each group

Fig. 6

Representative BLI of PyMT-BO1 GFP/Luc metastatic breast cancer cells in C57B6. a Untreated C57B6 mouse bearing highly metastatic PyMT-BO1 cancer. Accumulation in the lower limbs were predominant. b Mouse treated with 30 mg kg−1 of HSA-TC nanoparticles. c Mouse treated with 800 µCi of 18FDG. d Mouse treated with a combination of HSA-TC and 800 µCi 18FDG. e Quantification of whole-body luminescence in CRIT-treated mice compared to untreated, HSA-TC treated or 18FDG-treated controls (*P values are 0.038, 0.23 and 0.017 for CRIT, HSA-TC alone and 18FDG alone, respectively). BLI and data analysis were performed on day 9 after initiation of PyMT-BO1 metastasis in mice. Studies were performed with n = 5 mice per each group