TY - JOUR
T1 - Rapamycin selectively inhibits expression of an inducible keratin (K6a) in human keratinocytes and improves symptoms in pachyonychia congenita patients
AU - Hickerson, Robyn P.
AU - Leake, Devin
AU - Pho, Lana N.
AU - Leachman, Sancy A.
AU - Kaspar, Roger L.
N1 - Funding Information:
The authors are grateful for the participation of the PC patients, without whom this study would not have been possible and for the unfailing support of Mary Schwartz and the patient support organization, PC Project. The authors thank the International Pachyonychia Congenita Consortium (IPCC) and in particular W.H. Irwin McLean and Frances J.D. Smith for their critical reading and insightful suggestions, Manuel Flores and Heini Ilves for technical assistance and Tycho Speaker for assistance with data analysis. This work was supported by PC Project (SAL) and NIH grant R43AR056165 (RLK).
PY - 2009/11
Y1 - 2009/11
N2 - Background: The macrolide sirolimus (rapamycin) selectively blocks translation of mRNAs containing a terminal 5′ oligopyrimidine (TOP) tract by altering the activity of mammalian target of rapamycin (mTOR) and inhibiting downstream mTOR pathway components involved in TOP mRNA translation. The skin disorder pachyonychia congenita (PC) is caused by mutations in the inducible keratins (K) including K6a, K6b, K16 and K17. Published sequence data suggest the 5′ untranslated regions of K6a and K6b mRNAs contain 5′ TOP motifs and therefore may be sensitive to rapamycin treatment. Objective: Determine if mTOR inhibitors (rapamycin, temsirolimus or everolimus) are viable drug candidates for treatment of PC and other disorders caused by inappropriate expression of K6a and K6b. Methods: 5′ RACE analysis was used to map the transcriptional start sites for K5, K6a, K6b, K14, K16 and K17. The sensitivity of these keratins to mTOR inhibitors was determined by Western and qPCR analysis following treatment of a human HaCaT keratinocyte cell line with rapamycin, temsirolimus or everolimus. A small off-label study was undertaken using orally administered rapamycin in three PC patients and the effects were monitored by clinical examination, photography, a validated Dermatology Life Quality Index (DLQI) and a pain and activity diary. Results: Sequence comparison and 5′ RACE analysis of the 5′ untranslated regions of K6a and K6b revealed putative TOP regulatory elements. Treatment of a human HaCaT keratinocyte cell line with mTOR inhibitors (rapamycin, temsirolimus or everolimus) resulted in selective K6a repression. Furthermore, treatment of this HaCaT cell line with siRNAs targeting components of the mTOR pathway altered the levels of K6a expression. To test the ability of rapamycin to ameliorate PC symptoms, an off-label study was conducted. PC patient clinical responses to oral rapamycin showed a therapeutic response in callus character as well as subjective improvement. Of particular note, rapamycin greatly reduced the presence of painful cutaneous thromboses after reaching therapeutic serum levels. The well-known rapamycin side effects led to the early withdrawal of all of the patients from the study. Conclusion: Rapamycin selectively blocks K6a expression in human keratinocytes. The improvement of symptoms in PC patients following rapamycin treatment suggests rapamycin (or rapamycin analogs) may be a therapeutic option, particularly if topical formulations can be developed that avoid the side effects associated with systemic administration.
AB - Background: The macrolide sirolimus (rapamycin) selectively blocks translation of mRNAs containing a terminal 5′ oligopyrimidine (TOP) tract by altering the activity of mammalian target of rapamycin (mTOR) and inhibiting downstream mTOR pathway components involved in TOP mRNA translation. The skin disorder pachyonychia congenita (PC) is caused by mutations in the inducible keratins (K) including K6a, K6b, K16 and K17. Published sequence data suggest the 5′ untranslated regions of K6a and K6b mRNAs contain 5′ TOP motifs and therefore may be sensitive to rapamycin treatment. Objective: Determine if mTOR inhibitors (rapamycin, temsirolimus or everolimus) are viable drug candidates for treatment of PC and other disorders caused by inappropriate expression of K6a and K6b. Methods: 5′ RACE analysis was used to map the transcriptional start sites for K5, K6a, K6b, K14, K16 and K17. The sensitivity of these keratins to mTOR inhibitors was determined by Western and qPCR analysis following treatment of a human HaCaT keratinocyte cell line with rapamycin, temsirolimus or everolimus. A small off-label study was undertaken using orally administered rapamycin in three PC patients and the effects were monitored by clinical examination, photography, a validated Dermatology Life Quality Index (DLQI) and a pain and activity diary. Results: Sequence comparison and 5′ RACE analysis of the 5′ untranslated regions of K6a and K6b revealed putative TOP regulatory elements. Treatment of a human HaCaT keratinocyte cell line with mTOR inhibitors (rapamycin, temsirolimus or everolimus) resulted in selective K6a repression. Furthermore, treatment of this HaCaT cell line with siRNAs targeting components of the mTOR pathway altered the levels of K6a expression. To test the ability of rapamycin to ameliorate PC symptoms, an off-label study was conducted. PC patient clinical responses to oral rapamycin showed a therapeutic response in callus character as well as subjective improvement. Of particular note, rapamycin greatly reduced the presence of painful cutaneous thromboses after reaching therapeutic serum levels. The well-known rapamycin side effects led to the early withdrawal of all of the patients from the study. Conclusion: Rapamycin selectively blocks K6a expression in human keratinocytes. The improvement of symptoms in PC patients following rapamycin treatment suggests rapamycin (or rapamycin analogs) may be a therapeutic option, particularly if topical formulations can be developed that avoid the side effects associated with systemic administration.
KW - Sirolimus
KW - TOP mRNA
KW - Translational regulation
KW - mTOR inhibitor
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U2 - 10.1016/j.jdermsci.2009.07.008
DO - 10.1016/j.jdermsci.2009.07.008
M3 - Article
C2 - 19699613
AN - SCOPUS:70349987628
SN - 0923-1811
VL - 56
SP - 82
EP - 88
JO - Journal of Dermatological Science
JF - Journal of Dermatological Science
IS - 2
ER -