Vincristine Exposure and Sex-Specific Impact on Lower Urinary Tract Function

Background: vincristine, a childhood cancer drug, and lower urinary tract function

Vincristine (VCR) is a commonly-used chemotherapy agent for treating childhood cancer. Side effects of VCR and many other classes of chemotherapy agents include peripheral neuropathy, referred to as chemotherapy-induced peripheral neuropathy (CIPN). The risk of developing CIPN increases with higher doses, multiple therapy cycles, and combinations of neurotoxic chemotherapy. 

VCR-induced peripheral neuropathy (VIPN) can cause numbness, tingling, hand and foot pain, muscle weakness, gait abnormalities, and constipation due to affected sensorimotor and autonomic nerves. 

The impact of VCR on normal lower urinary tract (LUT) function, which requires coordinated interaction of the autonomic and somatic nervous systems as well as the bladder muscle, has not yet been well defined. In a recent survey of childhood cancer survivors from treated in the Center for Cancer and Blood Disorders at Children’s Hospital Colorado, VCR exposure was linked to LUT dysfunction (LUTD). Differences were noted between male and female patients.

The study used juvenile mouse models to examine how VCR exposure affects LUT physiology and function. Led by a multi-disciplinary team of Children’s Colorado researchers, study authors included:

• Nicholas Cost, MD, co-director, Surgical Oncology Program and director, Pediatric Urology fellowship, Children’s Colorado 
• Duncan Wilcox, MD, surgeon-in-chief and pediatric urologist, Children’s Colorado
• Dexiang Gao, PhD, professor, Pediatrics-Endocrinology, University of Colorado School of Medicine 
• Anna P. Malykhina, PhD, professor, Surgery-Urology, University of Colorado School of Medicine 

Methods: data from mice who receive vincristine or saline 

Starting at 3.5 weeks old, mice in the VCR group were given 0.5 mg/kg vincristine sulfate injection USP. The control group was given the same volume of saline by intraperitoneal injections twice weekly for four weeks. Experiments were conducted at 5 weeks after the last administration cycle. 

Cystometry

A subset of mice had a catheter surgically implanted in the bladder for cystometry studies at 4 weeks after the last administration cycle:

• Observed for 6 voiding cycles of reproducible micturition patterns
• Continuous recording of urodynamic values
• Parameters analyzed:
  • Maximum intravesical pressure at urination
  • Functional bladder capacity
  • Voided volume
  • Non-void contractions (NVCs) - intravesical pressure rises greater than 1/3 of the average maximal voiding pressure in each animal without triggering urination

Other data

Additional data gathered included:

• Manual von Frey tests
• Histological analysis
• In vitro detrusor contractility measurements
• Quantitative real‑time polymerase chain reaction (qPCR)
• Western blotting
• Immunofluorescence labeling

Statistical analysis

Two-way ANOVA was used to analyze all data among four groups (each sex and treatment, in control and VCR groups).

• Outliers excluded from analysis in in vitro detrusor contractility measurements
• Probability value of p < 0.05 considered significant

Results:  effects of systemic vincristine exposure on LUT function

Systemic VCR does not affect bladder morphology in mice

Compared to the control group:

• Significant growth deficit in male and female mice who received a cumulative dose of 2 or 4 mg/kg of VCR
• Trend of increased bladder wet weight in male VCR mice 
• No apparent differences in bladder morphology in:
  •  Urothelial
  • Lamina propria, DSM layers
  • Distribution, proportional amount of collagen contents

Systemic VCR exposure induces sexual dimorphic changes in urodynamic parameters

All groups had about 100% of bladder voiding efficiency. Compared to the control group:

• 3 urodynamic parameters significantly increased in both sexes of VCR group:
  • Functional bladder capacity
  • Void volume
  • NVCs
• Notable increase in maximal intravesical pressure at urination in VCR group females
• Increased bladder compliance in VCR group males

VCR exposure induces mechanical hyposensitivity

Compared to control group:

• Significant decrease in paw withdrawal/flinching in both sexes of VCR group
• More severe mechanical hyposensitivity in VCR group females 

VCR-induced detrusor overactivity in male mice

• No changes in the contractile response to all stimuli tested (EFS, CCh, αβMeATP, KCl) between control and VCR groups, nor among males and females
• Significant increase in frequency and amplitude of spontaneous contractions in bladder strips of male VCR mice 
  • Suggests VCR induces male-specific detrusor overactivity

VCR induces sexually dimorphic changes in gene expression patterns in the bladders and Ls‑DRG

The study authors analyzed the molecular mechanisms contributing to VCR-induced changes in LUT function. They found VCR exposure caused sexual dimorphic changes. 

There were no differences in the expression level of PkcƐ detected in the bladder, Ls-DRG between groups.

Discussion and conclusion: Sex-specific changes from vincristine alter LUT function

Study findings suggest systemic VCR exposure during the juvenile period induces five key changes:

• Decreased urinary frequency along with increased functional bladder capacity
  • Similar clinical observations in prospective study of pediatric cancer survivors who received VCR and/or doxorubicin (DOX) therapy compared to controls
• Female-specific increase in intravesical pressure at urination 
  • VCR-induced downregulations of Htr3b likely compromise Htr3a/b channel activity in the bladder along with decrease in Cav1.2 in Ls-DRG, impairing bladder afferent firing, leads to decreased bladder sensation until filled to maximum capacity
  • Observed increased NVCs without detrusor overactivity in vitro may  be another sign the bladder was reaching maximum capacity
• Male-specific detrusor overactivity
  • Itga1 may be involved in VCR-induced neuroinflammation in bladder in male mice
  • VCR-induced inflammation responses enhance Trpa1 expression and signaling, results in detrusor overactivity in male mice
• Mechanical hypersensitivity
  • Suggests mice experience impairment in the afferent pathway
  • VCR-induced sensory dysfunction reduces bladder filling sensation, delays in recognizing bladder fullness and the urge to urinate
• Sex dimorphic changes in gene expression patterns in the bladder, Ls-DRG in mice
  • Systemic VCR exposure causes sensory neuropathy via sex-dimorphic mechanisms, leads to altered LUT function

VCR dosage and LUTD risk

The cumulative dose of 4 mg/kg VCR in mice was equivalent to 12 mg/m2 VCR in children. 

• Considered a low dose in childhood cancer treatments, yet mice showed signs of LUTD
• Higher doses of VCR, combined with DOX/other chemotherapy agents may lead to more side effects
  • Severe VIPN
  • Constipation
  • Syndrome of inappropriate secretion of antidiuretic hormone
• Suggests increased risk of LUTD in childhood cancer survivors treated with VCR and DOX

Study authors concluded their data revealed that systemic VCR exposure during the juvenile period affects genes and molecular pathways in the bladder and Ls-DRG in a sexual dimorphic pattern, leading to sex-specific side effects. VCR-induced changes could clinically present as gender-specific signs of LUTD. Thus, pediatric cancer patients treated with VCR may warrant a follow-up urological assessment.