Background & Aims

Anastrozole, an aromatase inhibitor, constitutes the primary pharmacological approach for women diagnosed with hormone receptor-positive breast cancer, being prescribed by a minimum of five years post-cancer diagnosis [1, 2]. However, 30-60% of patients report pain, mainly musculoskeletal pain symptoms, characterized as symmetrical joint pain, morning stiffness, myalgia, and decreased strength [3, 4]. Understanding the mechanisms involved in painful symptoms related to anastrozole use is essential to manage them better. In this context, we investigated the participation of the Transient Receptor Potential Vanilloid 4 (TRPV4), a potential mediator of several nociceptive mechanisms [5, 6], in anastrozole-induced musculoskeletal pain in mice. Additionally, we explored the potential sensibilization of TRPV4 channel by signalling pathways downstream, PLC, PKC, and PKC? from kinin B2 (B2R) and B1 (B1R) receptors activation in anastrozole-induced pain.

Methods

Anastrozole (0.2 mg/kg) or vehicle (control group, 10 mL/kg, p.o.; 0.5% CMC) were administered by oral route (p.o.) in C57BL/6J male mice (IACUC #3026220520/2020 and #4535190722/2022). At 2 h after the anastrozole or vehicle administration, the mechanical allodynia and muscle strength were evaluated using the von Frey test and Grip test, respectively. The involvement of the TRPV4 channel was assessed using its antagonist HC067047 (10 mg/kg, intraperitoneal, [i.p.]) and agonists, 4?-PDD (3 nmol/paw) or hypotonic solution (deionized water, 20 ?L/paw). The interaction between the kinin receptors and the TRPV4 channel was evaluated with the local administration of sub-nociceptive doses of the kinin B2R, Bradykinin (1 nmol/paw) or B1R, DABk (3 nmol/paw) agonists and the effect of TRPV4 antagonist. PLC/PKC and PKC? inhibitors were used to evaluate the signalling pathways downstream from B2R and B1R activation and their effect on TRPV4 sensitization.

Results

TRPV4 antagonist, HC067047, reduced the anastrozole-induced mechanical allodynia and muscle strength loss of mice. The local administration of sub-nociceptive doses of the TRPV4 (4?-PDD or hypotonic solution) enhanced the anastrozole-induced pain behaviours, and HC067047 reduced this sensitizing effect induced by TRPV4 agonist in animals previously treated with anastrozole. Kinin B2R (Bradykinin) or B1R (DABk) agonists increased the anastrozole-induced pain behaviours. The sensitizing effects induced by local injection of kinin B2R and B1R agonists in animals previously treated with anastrozole were reduced by pre-treatment with TRPV4 antagonist. Furthermore, inhibition of PLC, PKC or PKC? attenuated the enhancement of mechanical allodynia and muscle strength loss induced by B2R and B1R agonists in previously anastrozole-treated mice.

Conclusions

Here, we demonstrated for the first time that the painful symptoms induced by anastrozole are TRPV4 channel-dependent in mice. Furthermore, the effect of the TRPV4 antagonist in reducing the sensitizing effect on painful symptoms caused by kinin agonists in mice previously treated with anastrozole showed the existence of a crosstalk between the TRPV4 channels and the kinin B2R and B1R. Since intracellular signalling pathways play vital roles in pain behaviours and can be activated downstream of kinin receptor activation, our results suggest that kinin B2R and B1R activation can lead to TRPV4 sensitization via the PLC, PKC and PKC? signalling pathways, supporting kinin receptor-dependent signalling pathways as a regulator of TRPV4 activity in the anastrozole-induced pain model. Thus, regulating this signalling pathway makes them potential therapeutic targets for alleviating the pain symptoms reported by aromatase inhibitor users, such as anastrozole.

References

1. Waks AG, Winer EP (2019) Breast Cancer Treatment. JAMA. 321:288. https://doi.org/10.1001/jama.2018.19323.
2. Tenti S, Correale P, Cheleschi S et al (2020) Aromatase Inhibitors-Induced Musculoskeletal Disorders: current knowledge on clinical and molecular aspects. Int J Mol Sci 21:5625. https://doi. org/10.3390/ijms21165625.
3. Crew KD, Greenlee H, Capodice J et al (2007) Prevalence of joint symptoms in Postmenopausal Women taking aromatase inhibitors for early-stage breast Cancer. J Clin Oncol 25:3877–3883. https://doi.org/10.1200/JCO.2007.10.7573.
4. Laroche F, Coste J, Medkour T et al (2014) Classification of and risk factors for Estrogen Deprivation Pain Syndromes related to aromatase inhibitor treatments in women with breast Cancer: a prospective Multicenter Cohort Study. J Pain 15:293–303. https://doi.org/10.1016/j.jpain.2013.11.004.
5. Kumar H, Lee SH, Kim KT et al (2018) TRPV4: a Sensor for Homeostasis and pathological events in the CNS. Mol Neurobiol 55:8695–8708. https://doi.org/10.1007/s12035-018-0998-8.
6. Grace MS, Bonvini SJ, Belvisi MG, McIntyre P (2017) Modulation of the TRPV4 ion channel as a therapeutic target for disease. Pharmacol Ther 177:9–22. https://doi.org/10.1016/j. pharmthera.2017.02.019.

Presenting Author

Gabriela Becker

Poster Authors

Gabriela Becker

PhD student

Federal University of Santa Maria

Lead Author

Maria Fialho

Federal University of Santa Maria, Santa Maria, RS, Brazil

Lead Author

Evelyne Silva Brum

PhD

Federal University of Santa Maria

Lead Author

Sara Marchesan Oliveira

PhD

Federal University of Santa Maria

Lead Author

Topics

  • Models: Chronic Pain - Neuropathic