CJC-1295 (no DAC)

1. Studied for

   Sustained GH and IGF-I response in humans (DAC-conjugated form, NOT this compound)

   2 randomized, placebo-controlled, double-blind, ascending-level trials, running 28 and 49 days, were carried out in healthy adults aged 21 to 61. Participants received either the compound or placebo under the skin, as one of 4 ascending single amounts in the first study, and as 2 or 3 weekly or every-other-week amounts in the second. After a single exposure, average blood growth hormone rose 2- to 10-fold and stayed up for 6 days or more, and average blood IGF-I rose 1.5- to 3-fold for 9 to 11 days, both in proportion to the amount given. The estimated half-life of the compound was 5.8 to 8.1 days. After repeat exposures, average IGF-I stayed above starting levels for up to 28 days. No serious adverse reactions were reported. The scope fact that must never be separated from these numbers by layout: this work used the DAC-conjugated, albumin-binding CJC-1295, which is a different molecule from the no-DAC research peptide supplied here. No equivalent human trial of the no-DAC form was located. HUMAN human trial (2 randomized, placebo-controlled, double-blind ascending-level trials in healthy adults aged 21 to 61) Preclinical strong Convergent results from more than one independent laboratory, or a controlled human trial design. On this page 'strong' rates the quality of the study and never the relevance of its subject: both strong rows below were run on molecules other than the one supplied here, one on the albumin-binding DAC conjugate and one on native GHRH. Open for the full finding and the paper it came from.

   In the literature

   Sustained GH and IGF-I response in humans (DAC-conjugated form, NOT this compound)

   2 randomized, placebo-controlled, double-blind, ascending-level trials, running 28 and 49 days, were carried out in healthy adults aged 21 to 61. Participants received either the compound or placebo under the skin, as one of 4 ascending single amounts in the first study, and as 2 or 3 weekly or every-other-week amounts in the second. After a single exposure, average blood growth hormone rose 2- to 10-fold and stayed up for 6 days or more, and average blood IGF-I rose 1.5- to 3-fold for 9 to 11 days, both in proportion to the amount given. The estimated half-life of the compound was 5.8 to 8.1 days. After repeat exposures, average IGF-I stayed above starting levels for up to 28 days. No serious adverse reactions were reported. The scope fact that must never be separated from these numbers by layout: this work used the DAC-conjugated, albumin-binding CJC-1295, which is a different molecule from the no-DAC research peptide supplied here. No equivalent human trial of the no-DAC form was located.^{References 1}

   Modelhuman trial (2 randomized, placebo-controlled, double-blind ascending-level trials in healthy adults aged 21 to 61)

   Confidence: strong

   Convergent results from more than one independent laboratory, or a controlled human trial design. On this page 'strong' rates the quality of the study and never the relevance of its subject: both strong rows below were run on molecules other than the one supplied here, one on the albumin-binding DAC conjugate and one on native GHRH.

   Source

   Teichman SL, Neale A, Lawrence B, Gagnon C, Castaigne JP, Frohman LA. Prolonged stimulation of growth hormone (GH) and insulin-like growth factor I secretion by CJC-1295, a long-acting analog of GH-releasing hormone, in healthy adults. J Clin Endocrinol Metab. 2006;91(3):799-805. DOI: 10.1210/jc.2005-1536. PMID: 16352683

   PubMed 16352683

   How this citation was checked

   Re-resolved live in this pass. Crossref API on DOI 10.1210/jc.2005-1536 returned the exact title, all six authors in order (Teichman first, Frohman last), The Journal of Clinical Endocrinology and Metabolism, 2006, volume 91, issue 3, pages 799-805. PubMed efetch on PMID 16352683 returned identical bibliographic data and the structured abstract, from which we read verbatim: 'two randomized, placebo-controlled, double-blind, ascending dose trials with durations of 28 and 49 d'; 'Healthy subjects, ages 21-61 yr'; 'in one of four ascending single doses in the first study and in two or three weekly or biweekly doses in the second study'; 'dose-dependent increases in mean plasma GH concentrations by 2- to 10-fold for 6 d or more and in mean plasma IGF-I concentrations by 1.5- to 3-fold for 9-11 d'; 'The estimated half-life of CJC-1295 was 5.8-8.1 d'; 'After multiple CJC-1295 doses, mean IGF-I levels remained above baseline for up to 28 d'; 'No serious adverse reactions were reported'. The 'n=21' and 'phase I' descriptors from an earlier draft do not appear in the source and remain deleted. The abstract's conclusion names specific microgram-per-kilogram figures; those are deliberately omitted from this dossier.

2. Studied for

   Two independent GH-releasing pathways: GHRH plus a GH-releasing peptide

   In 18 normal men, a synthetic 6-amino-acid growth-hormone-releasing peptide (GHRP-6, sequence His-DTrp-Ala-Trp-DPhe-Lys-NH2) was given into a vein at 3 ascending levels and compared with the natural releasing hormone in its full-length form, GHRH-(1-44)-NH2. Average peak blood growth hormone was 1.2 micrograms per liter after placebo and rose across the 3 ascending levels to 7.6, then 16.5, then 68.7 micrograms per liter. When the 2 lower, deliberately submaximal levels of the peptide were given together with the natural releasing hormone, growth-hormone release was synergistic, meaning the combination produced more than the 2 agents added together. The authors read that as evidence the 2 agents work through independent mechanisms. This is the published mechanistic basis for the GHRH-analog-plus-GHRP research pairings listed in the co-marketed section below. Scope note: the human data used native GHRH-(1-44)-NH2, not this modified GRF(1-29) analog, and tested GHRP-6 specifically, so the synergy is documented at the receptor-class level and not for this exact pair of molecules. human study (18 normal men, intravenous ascending-level comparison) Preclinical strong Convergent results from more than one independent laboratory, or a controlled human trial design. On this page 'strong' rates the quality of the study and never the relevance of its subject: both strong rows below were run on molecules other than the one supplied here, one on the albumin-binding DAC conjugate and one on native GHRH. Open for the full finding and the paper it came from.

   In the literature

   Two independent GH-releasing pathways: GHRH plus a GH-releasing peptide

   In 18 normal men, a synthetic 6-amino-acid growth-hormone-releasing peptide (GHRP-6, sequence His-DTrp-Ala-Trp-DPhe-Lys-NH2) was given into a vein at 3 ascending levels and compared with the natural releasing hormone in its full-length form, GHRH-(1-44)-NH2. Average peak blood growth hormone was 1.2 micrograms per liter after placebo and rose across the 3 ascending levels to 7.6, then 16.5, then 68.7 micrograms per liter. When the 2 lower, deliberately submaximal levels of the peptide were given together with the natural releasing hormone, growth-hormone release was synergistic, meaning the combination produced more than the 2 agents added together. The authors read that as evidence the 2 agents work through independent mechanisms. This is the published mechanistic basis for the GHRH-analog-plus-GHRP research pairings listed in the co-marketed section below. Scope note: the human data used native GHRH-(1-44)-NH2, not this modified GRF(1-29) analog, and tested GHRP-6 specifically, so the synergy is documented at the receptor-class level and not for this exact pair of molecules.^{References 2}

   Modelhuman study (18 normal men, intravenous ascending-level comparison)

   Confidence: strong

   Convergent results from more than one independent laboratory, or a controlled human trial design. On this page 'strong' rates the quality of the study and never the relevance of its subject: both strong rows below were run on molecules other than the one supplied here, one on the albumin-binding DAC conjugate and one on native GHRH.

   Source

   Bowers CY, Reynolds GA, Durham D, Barrera CM, Pezzoli SS, Thorner MO. Growth hormone (GH)-releasing peptide stimulates GH release in normal men and acts synergistically with GH-releasing hormone. J Clin Endocrinol Metab. 1990;70(4):975-982. PMID: 2108187

   PubMed 2108187

   How this citation was checked

   Re-resolved live in this pass. PubMed efetch on PMID 2108187 returned the exact title, all six authors in order (Bowers first, Thorner last), J Clin Endocrinol Metab, 1990, volume 70, issue 4, pages 975-982, plus the abstract. Read verbatim from it: 'The acute GH release stimulated by the synthetic hexapeptide, His-DTrp-Ala-Trp-DPhe-Lys-NH2 [GH releasing peptide (GHRP)], was determined in 18 normal men and compared with the effects of GH-releasing hormone, GHRH-(1-44)-NH2'; 'Mean (+/- SEM) peak serum GH levels after injection of placebo and 0.1, 0.3, and 1.0 microgram/kg GHRP were 1.2 +/- 0.3, 7.6 +/- 2.5, 16.5 +/- 4.1, and 68.7 +/- 15.5 micrograms/L, respectively'; 'The submaximal dosages of 0.1 and 0.3 microgram/kg GHRP plus 1 microgram/kg GHRH stimulated GH release synergistically'; and 'Since GHRP and GHRH together stimulate GH release synergistically, these results suggest that GHRP and GHRH act independently'. EDITS APPLIED IN THIS PASS: the microgram-per-kilogram dose figures have been removed and replaced with 'three ascending dose levels', while the peak growth-hormone outcome values are retained; and the GHRH form is now named as GHRH-(1-44)-NH2 rather than 'native GHRH'.

3. Studied for

   GHRH-receptor activation and acute GH release (the paper that named CJC-1295)

   Chemists built 3 modified versions of the human GHRH front-end fragment and attached each to human serum albumin, a carrier protein found in blood. In cultured rat pituitary cells, all 3 attached versions resisted the enzyme dipeptidylpeptidase-IV better than the unmodified fragment and still triggered growth hormone release. When the modified versions were given under the skin of normal male Sprague-Dawley rats, a burst of growth hormone appeared in the blood. The best of the 3, which the authors named CJC-1295, produced a 4-fold larger total growth-hormone response over 2 hours than the unmodified fragment, and the molecule itself was still detectable in plasma beyond 72 hours. Scope fact, which must never be separated from this result by layout: this paper DEFINES CJC-1295 as the tetrasubstituted hGRF(1-29) carrying an N-epsilon-3-maleimidopropionamide lysine at the C terminus, meaning the albumin-binding DAC form. Every compound tested here carried that tail. The no-DAC research peptide was not tested as a separate arm, and the comparator for the 4-fold figure was unmodified hGRF(1-29), not the no-DAC analog. IN VITRO RODENT in vitro (cultured rat anterior pituitary cells) plus rodent (normal male Sprague-Dawley rats, under the skin) Preclinical moderate A well-described result with several independent outcome measures, but from a single laboratory or a single study design. Open for the full finding and the paper it came from.

   In the literature

   GHRH-receptor activation and acute GH release (the paper that named CJC-1295)

   Chemists built 3 modified versions of the human GHRH front-end fragment and attached each to human serum albumin, a carrier protein found in blood. In cultured rat pituitary cells, all 3 attached versions resisted the enzyme dipeptidylpeptidase-IV better than the unmodified fragment and still triggered growth hormone release. When the modified versions were given under the skin of normal male Sprague-Dawley rats, a burst of growth hormone appeared in the blood. The best of the 3, which the authors named CJC-1295, produced a 4-fold larger total growth-hormone response over 2 hours than the unmodified fragment, and the molecule itself was still detectable in plasma beyond 72 hours. Scope fact, which must never be separated from this result by layout: this paper DEFINES CJC-1295 as the tetrasubstituted hGRF(1-29) carrying an N-epsilon-3-maleimidopropionamide lysine at the C terminus, meaning the albumin-binding DAC form. Every compound tested here carried that tail. The no-DAC research peptide was not tested as a separate arm, and the comparator for the 4-fold figure was unmodified hGRF(1-29), not the no-DAC analog.^{References 3}

   Modelin vitro (cultured rat anterior pituitary cells) plus rodent (normal male Sprague-Dawley rats, under the skin)

   Confidence: moderate

   A well-described result with several independent outcome measures, but from a single laboratory or a single study design.

   Source

   Jette L, Leger R, Thibaudeau K, Benquet C, Robitaille M, Pellerin I, Paradis V, van Wyk P, Pham K, Bridon DP. Human growth hormone-releasing factor (hGRF)1-29-albumin bioconjugates activate the GRF receptor on the anterior pituitary in rats: identification of CJC-1295 as a long-lasting GRF analog. Endocrinology. 2005;146(7):3052-3058. DOI: 10.1210/en.2004-1286. PMID: 15817669

   PubMed 15817669

   How this citation was checked

   Re-resolved live in this pass. Crossref API on DOI 10.1210/en.2004-1286 returned the exact title, all ten authors in order (Jette first, Bridon last), Endocrinology, 2005, volume 146, issue 7, pages 3052-3058. PubMed efetch on PMID 15817669 returned identical bibliographic data plus the full abstract, from which we read verbatim: 'All three human serum albumin conjugates showed enhanced in vitro stability against dipeptidylpeptidase-IV and were bioactive in a GH secretion assay in cultured rat anterior pituitary cells'; 'administered sc to normal male Sprague Dawley rats'; 'The best compound, CJC-1295, showed a 4-fold increase in GH area under the curve over a 2-h period compared with hGRF(1-29)'; 'it was found to be present in plasma beyond 72 h'; and 'CJC-1295, a tetrasubstituted form of hGRF(1-29) with an added N epsilon-3-maleimidopropionamide derivative of lysine at the C terminus'. We note for accuracy that the separate 'beyond 24 h' figure in that abstract belongs to a Western blot observation, which this dossier does not cite.

4. Studied for

   Whether continuous GHRH-receptor stimulation flattens natural GH pulsatility (DAC form)

   Growth hormone is normally released in bursts through the night, and researchers wanted to know whether a long-acting version of the signal would flatten that rhythm into a constant drip. Healthy men aged 20 to 40 had blood drawn every 20 minutes across an overnight 12-hour window, once before and once a week after a single exposure at one of 2 ascending levels. Overall growth-hormone output went up, and the rhythm survived: the number and size of the secretory bursts were unchanged, while the baseline trough level between bursts rose markedly (7.5-fold), average growth hormone rose 46 percent and IGF-I rose 45 percent. No significant difference was seen between the 2 levels. Scope note: the compound studied was the DAC-conjugated form, so this is cited as pharmacology of the GHRH receptor, not as data on the no-DAC research peptide. human study (healthy men aged 20 to 40, overnight frequent-sampling protocol) Preclinical moderate A well-described result with several independent outcome measures, but from a single laboratory or a single study design. Open for the full finding and the paper it came from.

   In the literature

   Whether continuous GHRH-receptor stimulation flattens natural GH pulsatility (DAC form)

   Growth hormone is normally released in bursts through the night, and researchers wanted to know whether a long-acting version of the signal would flatten that rhythm into a constant drip. Healthy men aged 20 to 40 had blood drawn every 20 minutes across an overnight 12-hour window, once before and once a week after a single exposure at one of 2 ascending levels. Overall growth-hormone output went up, and the rhythm survived: the number and size of the secretory bursts were unchanged, while the baseline trough level between bursts rose markedly (7.5-fold), average growth hormone rose 46 percent and IGF-I rose 45 percent. No significant difference was seen between the 2 levels. Scope note: the compound studied was the DAC-conjugated form, so this is cited as pharmacology of the GHRH receptor, not as data on the no-DAC research peptide.^{References 4}

   Modelhuman study (healthy men aged 20 to 40, overnight frequent-sampling protocol)

   Confidence: moderate

   A well-described result with several independent outcome measures, but from a single laboratory or a single study design.

   Source

   Ionescu M, Frohman LA. Pulsatile secretion of growth hormone (GH) persists during continuous stimulation by CJC-1295, a long-acting GH-releasing hormone analog. J Clin Endocrinol Metab. 2006;91(12):4792-4797. DOI: 10.1210/jc.2006-1702. PMID: 17018654

   PubMed 17018654

   How this citation was checked

   Re-resolved live in this pass, with extra scrutiny because an earlier draft carried a DOI that does not resolve. Crossref API on DOI 10.1210/jc.2006-1702 returned the exact title, both authors (Ionescu, Frohman), The Journal of Clinical Endocrinology and Metabolism, 2006, volume 91, issue 12, pages 4792-4797. PubMed efetch on PMID 17018654 returned identical bibliographic data and the abstract, confirming verbatim 'GH pulsatility was assessed by 20-min blood sampling during an overnight 12-h period in healthy 20- to 40-yr-old men before and 1 wk after injection'; 'GH secretion was increased after CJC-1295 administration with preserved pulsatility. The frequency and magnitude of GH secretory pulses were unaltered. However, basal (trough) GH levels were markedly increased (7.5-fold; P < 0.0001)'; 'mean GH levels, 46%; P < 0.01'; 'IGF-I levels (45%; P < 0.001)'; and 'No significant differences were observed between the responses to the two drug doses'. The abstract's microgram-per-kilogram figures are deliberately omitted from this dossier and replaced with 'one of two ascending dose levels'. The discarded DOI 10.1210/jc.2006-1290 is not used anywhere.

5. Studied for

   GHRH-receptor dependence tested in a GHRH-deficient animal model (DAC form)

   Mice bred without the gene for growth-hormone-releasing hormone grow poorly, which makes them a clean test of whether a molecule works through the GHRH receptor. Starting at 1 week old, 3 groups of these mice received the same amount of compound for 5 weeks, given at 24-, 48- or 72-hour intervals. The mice on the 24-hour schedule ended up with normal body weight and length. The 48- and 72-hour groups grew heavier and longer than the placebo group but did not fully normalize. The exposed animals showed an increase in total pituitary RNA and in growth-hormone messenger RNA, which the authors read as multiplication of the pituitary cells that make growth hormone, supported by immunohistochemistry images. Because the effect requires an intact pituitary downstream, this supports action through the GHRH receptor. Scope note: the compound studied was the DAC-conjugated form. RODENT rodent (GHRH knockout mice, from 1 week of age for 5 weeks) Preclinical moderate A well-described result with several independent outcome measures, but from a single laboratory or a single study design. Open for the full finding and the paper it came from.

   In the literature

   GHRH-receptor dependence tested in a GHRH-deficient animal model (DAC form)

   Mice bred without the gene for growth-hormone-releasing hormone grow poorly, which makes them a clean test of whether a molecule works through the GHRH receptor. Starting at 1 week old, 3 groups of these mice received the same amount of compound for 5 weeks, given at 24-, 48- or 72-hour intervals. The mice on the 24-hour schedule ended up with normal body weight and length. The 48- and 72-hour groups grew heavier and longer than the placebo group but did not fully normalize. The exposed animals showed an increase in total pituitary RNA and in growth-hormone messenger RNA, which the authors read as multiplication of the pituitary cells that make growth hormone, supported by immunohistochemistry images. Because the effect requires an intact pituitary downstream, this supports action through the GHRH receptor. Scope note: the compound studied was the DAC-conjugated form.^{References 5}

   Modelrodent (GHRH knockout mice, from 1 week of age for 5 weeks)

   Confidence: moderate

   A well-described result with several independent outcome measures, but from a single laboratory or a single study design.

   Source

   Alba M, Fintini D, Sagazio A, Lawrence B, Castaigne JP, Frohman LA, Salvatori R. Once-daily administration of CJC-1295, a long-acting growth hormone-releasing hormone (GHRH) analog, normalizes growth in the GHRH knockout mouse. Am J Physiol Endocrinol Metab. 2006;291(6):E1290-E1294. DOI: 10.1152/ajpendo.00201.2006. PMID: 16822960

   PubMed 16822960

   How this citation was checked

   Re-resolved live in this pass. Crossref API on DOI 10.1152/ajpendo.00201.2006 returned the exact title, all seven authors in order (Alba first, Salvatori last), American Journal of Physiology-Endocrinology and Metabolism, 2006, volume 291, issue 6, pages E1290-E1294. PubMed efetch on PMID 16822960 returned identical bibliographic data and the abstract, confirming verbatim 'Three groups of 1-wk-old GHRHKO mice were treated for 5 wk with 2 microg of CJC-1295 at intervals of 24, 48, and 72 h'; 'GHRHKO animals receiving daily doses of CJC-1295 exhibited normal body weight and length'; 'Mice treated every 48 and 72 h reached higher body weight and length than placebo-treated animals, without full growth normalization'; 'Relative lean mass and subcutaneous fat mass were normal in all treated groups'; and 'CJC-1295 caused an increase in total pituitary RNA and GH mRNA, suggesting that proliferation of somatotroph cells had occurred, as confirmed by immunohistochemistry images'. CORRECTION APPLIED IN THIS PASS: an earlier draft said 'increased pituitary weight and GH mRNA'. The paper does not report pituitary weight; it reports total pituitary RNA. The sentence has been corrected to match the source. The microgram dose figure in the abstract is deliberately omitted from this dossier.

6. Studied for

   The unmodified parent fragment GHRH(1-29) in older men: GH measures rose, IGF-I did not

   11 healthy, non-obese, walking-independent men aged 64 to 76, selected because their baseline IGF-I levels were low, received the unmodified GHRH 1-29 fragment under the skin nightly at home for 6 weeks, with blood sampled every 20 minutes overnight before and after. Average nighttime growth-hormone release, the area under the growth-hormone peak, and the peak height all rose significantly, while the number of bursts per night did not change. The downstream markers did not move: the study reported no change in IGF-I, IGF binding protein-3 or growth-hormone binding protein. This null result is included on purpose. It shows that pushing up nighttime growth-hormone measures with a GHRH fragment did not produce a measurable IGF-I change in this trial. Scope note: the compound studied was the unmodified 1-29 fragment (sermorelin), not the 4-substitution research analog. HUMAN human trial (11 healthy men aged 64 to 76, 6 weeks of nightly self-administration under the skin) Preclinical moderate A well-described result with several independent outcome measures, but from a single laboratory or a single study design. Open for the full finding and the paper it came from.

   In the literature

   The unmodified parent fragment GHRH(1-29) in older men: GH measures rose, IGF-I did not

   11 healthy, non-obese, walking-independent men aged 64 to 76, selected because their baseline IGF-I levels were low, received the unmodified GHRH 1-29 fragment under the skin nightly at home for 6 weeks, with blood sampled every 20 minutes overnight before and after. Average nighttime growth-hormone release, the area under the growth-hormone peak, and the peak height all rose significantly, while the number of bursts per night did not change. The downstream markers did not move: the study reported no change in IGF-I, IGF binding protein-3 or growth-hormone binding protein. This null result is included on purpose. It shows that pushing up nighttime growth-hormone measures with a GHRH fragment did not produce a measurable IGF-I change in this trial. Scope note: the compound studied was the unmodified 1-29 fragment (sermorelin), not the 4-substitution research analog.^{References 6}

   Modelhuman trial (11 healthy men aged 64 to 76, 6 weeks of nightly self-administration under the skin)

   Confidence: moderate

   A well-described result with several independent outcome measures, but from a single laboratory or a single study design.

   Source

   Vittone J, Blackman MR, Busby-Whitehead J, Tsiao C, Stewart KJ, Tobin J, Stevens T, Bellantoni MF, Rogers MA, Baumann G, Roth J, Harman SM, Spencer RG. Effects of single nightly injections of growth hormone-releasing hormone (GHRH 1-29) in healthy elderly men. Metabolism. 1997;46(1):89-96. PMID: 9005976

   PubMed 9005976

   How this citation was checked

   Re-resolved live in this pass. PubMed efetch on PMID 9005976 returned the exact title, the complete 13-author list in order (Vittone first, Spencer last), Metabolism, 1997, volume 46, issue 1, pages 89-96, plus the abstract. Read verbatim from it: 'Eleven healthy, ambulatory, non-obese men aged 64 to 76 years with low baseline IGF-I levels were treated at home as outpatients by nightly subcutaneous self-injections'; 'for 6 weeks'; 'We measured GH levels in blood samples obtained every 20 minutes from 8:00 PM to 8:00 AM'; and 'GHRH treatment increased mean nocturnal GH release (P < .02), the area under the GH peak ([AUPGH] P < .006), and GH peak amplitude (P < .05), with no change in GH pulse frequency or in levels of IGF-I, IGFBP-3, or GHBP'. This record carries no DOI and none is asserted. TWO EDITS APPLIED IN THIS PASS: the enrollment criterion 'with low baseline IGF-I levels' has been restored, because it is material context for the null IGF-I result this entry exists to carry; and the milligram dose figure has been removed. The trial's strength and muscle-function measures remain deliberately excluded from this dossier.

7. Studied for

   The reverse dependency: endogenous GHRH is required for a maximal GHRP-6 response

   9 healthy men aged 20 to 30 were studied twice, receiving either saline or a blocker built to sit on the GHRH receptor without switching it on, followed 20 minutes later by an intravenous bolus of GHRP-6, with blood sampled every 10 minutes. With the blocker on board, most of the growth-hormone response to GHRP-6 disappeared: the maximal rise above starting level fell from 33.8 plus or minus 4.8 to 6.2 plus or minus 1.8 micrograms per liter, and the total response measured as area under the curve fell from 1701 plus or minus 278 to 376 plus or minus 113 micrograms per minute per liter. The authors concluded that the body's own GHRH is necessary for most of the growth-hormone response to GHRP-6 in humans, meaning the 2 systems lean on each other rather than simply running in parallel. Note on magnitude: the paper says most of the response was eliminated, not all of it, and this dossier does not describe the response as abolished. human study (9 healthy men, a GHRH antagonist followed by a GHRP-6 bolus) Preclinical moderate A well-described result with several independent outcome measures, but from a single laboratory or a single study design. Open for the full finding and the paper it came from.

   In the literature

   The reverse dependency: endogenous GHRH is required for a maximal GHRP-6 response

   9 healthy men aged 20 to 30 were studied twice, receiving either saline or a blocker built to sit on the GHRH receptor without switching it on, followed 20 minutes later by an intravenous bolus of GHRP-6, with blood sampled every 10 minutes. With the blocker on board, most of the growth-hormone response to GHRP-6 disappeared: the maximal rise above starting level fell from 33.8 plus or minus 4.8 to 6.2 plus or minus 1.8 micrograms per liter, and the total response measured as area under the curve fell from 1701 plus or minus 278 to 376 plus or minus 113 micrograms per minute per liter. The authors concluded that the body's own GHRH is necessary for most of the growth-hormone response to GHRP-6 in humans, meaning the 2 systems lean on each other rather than simply running in parallel. Note on magnitude: the paper says most of the response was eliminated, not all of it, and this dossier does not describe the response as abolished.^{References 7}

   Modelhuman study (9 healthy men, a GHRH antagonist followed by a GHRP-6 bolus)

   Confidence: moderate

   A well-described result with several independent outcome measures, but from a single laboratory or a single study design.

   Source

   Pandya N, DeMott-Friberg R, Bowers CY, Barkan AL, Jaffe CA. Growth hormone (GH)-releasing peptide-6 requires endogenous hypothalamic GH-releasing hormone for maximal GH stimulation. J Clin Endocrinol Metab. 1998;83(4):1186-1189. PMID: 9543138

   PubMed 9543138

   How this citation was checked

   Re-resolved live in this pass. PubMed efetch on PMID 9543138 returned the exact title, the author list Pandya N, DeMott-Friberg R, Bowers CY, Barkan AL, Jaffe CA, J Clin Endocrinol Metab, 1998, volume 83, issue 4, pages 1186-1189, plus the abstract. Read verbatim from it: 'Nine healthy men between the ages of 20 and 30 yr were studied on two occasions'; 'Blood was sampled every 10 min from 0800-1100 h'; 'GHRH-Ant eliminated most of the GH response to GHRP-6 [maximal increase over the baseline GH concentration, 33.8 +/- 4.8 vs. 6.2 +/- 1.8 microg/L (mean +/- SEM; P < 0.0001); area under the curve, 1701 +/- 278 vs. 376 +/- 113 microg/min x L (P < 0.001)]'; and 'These data show that endogenous GHRH is necessary for most of the GH response to GHRP-6 in humans'. An earlier draft attributed this PMID to 'Pombo M, et al.'; that name appears nowhere on the author list and the attribution stays corrected. The microgram-per-kilogram figures for the antagonist and the bolus have been removed in this pass.

8. Studied for

   Analytical chemistry: identifying GHRH analogs in human plasma

   Anti-doping chemists built and validated a method to pull GHRH-type molecules out of human plasma using an antibody, then identify them by high-resolution tandem mass spectrometry. The method targeted 4 compounds, named in the paper as sermorelin, CJC-1293, CJC-1295 and tesamorelin, plus 2 breakdown products of sermorelin and CJC-1293. Reported method performance was a lower limit of detection below 50 picograms per milliliter, imprecision under 20 percent, and analyte recovery of 19 to 37 percent. The stability and breakdown of the compounds were examined using both test-tube and live-animal approaches, with blood collected from rats after intravenous administration; all intact substances were still detectable at least 4 hours later, though no expected breakdown product was confirmed in the rodent samples. This is method-development work, not a biological finding. RODENT analytical method development and validation (human plasma), plus a rodent (rat) breakdown arm Preclinical moderate A well-described result with several independent outcome measures, but from a single laboratory or a single study design. Open for the full finding and the paper it came from.

   In the literature

   Analytical chemistry: identifying GHRH analogs in human plasma

   Anti-doping chemists built and validated a method to pull GHRH-type molecules out of human plasma using an antibody, then identify them by high-resolution tandem mass spectrometry. The method targeted 4 compounds, named in the paper as sermorelin, CJC-1293, CJC-1295 and tesamorelin, plus 2 breakdown products of sermorelin and CJC-1293. Reported method performance was a lower limit of detection below 50 picograms per milliliter, imprecision under 20 percent, and analyte recovery of 19 to 37 percent. The stability and breakdown of the compounds were examined using both test-tube and live-animal approaches, with blood collected from rats after intravenous administration; all intact substances were still detectable at least 4 hours later, though no expected breakdown product was confirmed in the rodent samples. This is method-development work, not a biological finding.^{References 8}

   Modelanalytical method development and validation (human plasma), plus a rodent (rat) breakdown arm

   Confidence: moderate

   A well-described result with several independent outcome measures, but from a single laboratory or a single study design.

   Source

   Knoop A, Thomas A, Fichant E, Delahaut P, Schanzer W, Thevis M. Qualitative identification of growth hormone-releasing hormones in human plasma by means of immunoaffinity purification and LC-HRMS/MS. Anal Bioanal Chem. 2016;408(12):3145-3153. DOI: 10.1007/s00216-016-9377-3. PMID: 26879649

   PubMed 26879649

   How this citation was checked

   Re-resolved live in this pass. Crossref API on DOI 10.1007/s00216-016-9377-3 returned the exact title, all six authors in order (Knoop first, Thevis last), Analytical and Bioanalytical Chemistry, 2016, volume 408, issue 12, pages 3145-3153. PubMed efetch on PMID 26879649 returned identical bibliographic data and the abstract, confirming verbatim 'The target analytes included Geref (Sermorelin), CJC-1293, CJC-1295, and Egrifta (Tesamorelin) as well as two metabolites of Geref and CJC-1293'; 'recovery (19-37%), lower limit of detection (<50 pg/mL), imprecision (<20%)'; and 'EDTA blood samples were collected from rats 2, 4, and 8 h after intravenous administration of GHRH (one compound per test animal). All intact substances were detected for at least 4 h but no anticipated metabolite was confirmed in laboratory rodents' samples'. PRECISION EDIT APPLIED IN THIS PASS: the earlier draft's loose 'four GHRH analogs and their metabolites' is now stated as the paper states it, two metabolites of sermorelin and CJC-1293, and the rat arm is disclosed in the model field.

9. Studied for

   In vitro metabolism of GHRH analogs, and independent confirmation that the DAC and no-DAC forms are distinct analytes

   An anti-doping laboratory studied how 4 of the larger synthetic GHRH-type molecules break down, working in fortified human urine. The 4 are listed by the authors as sermorelin, tesamorelin, CJC-1295, and CJC-1295 with drug affinity complex. 19 major breakdown products were identified, then synthesized and characterized in house so they could be used as reference standards, and a liquid chromatography tandem mass spectrometry method was built around them, reaching detection limits generally at or below the 1 nanogram per milliliter performance limit required by the World Anti-Doping Agency. Beyond the analytical result, this paper is cited here for a labeling reason: an independent laboratory handles CJC-1295 and CJC-1295 with DAC as 2 separate chemical entities needing separate breakdown-product panels, which corroborates the naming distinction this dossier draws throughout. It is also the only entry on this page whose work is done on the no-DAC molecule as a distinct chemical entity, and it is analytical chemistry rather than biology. IN VITRO in vitro (fortified human urine) metabolism experiments plus analytical method development Preclinical moderate A well-described result with several independent outcome measures, but from a single laboratory or a single study design. Open for the full finding and the paper it came from.

   In the literature

   In vitro metabolism of GHRH analogs, and independent confirmation that the DAC and no-DAC forms are distinct analytes

   An anti-doping laboratory studied how 4 of the larger synthetic GHRH-type molecules break down, working in fortified human urine. The 4 are listed by the authors as sermorelin, tesamorelin, CJC-1295, and CJC-1295 with drug affinity complex. 19 major breakdown products were identified, then synthesized and characterized in house so they could be used as reference standards, and a liquid chromatography tandem mass spectrometry method was built around them, reaching detection limits generally at or below the 1 nanogram per milliliter performance limit required by the World Anti-Doping Agency. Beyond the analytical result, this paper is cited here for a labeling reason: an independent laboratory handles CJC-1295 and CJC-1295 with DAC as 2 separate chemical entities needing separate breakdown-product panels, which corroborates the naming distinction this dossier draws throughout. It is also the only entry on this page whose work is done on the no-DAC molecule as a distinct chemical entity, and it is analytical chemistry rather than biology.^{References 9}

   Modelin vitro (fortified human urine) metabolism experiments plus analytical method development

   Confidence: moderate

   A well-described result with several independent outcome measures, but from a single laboratory or a single study design.

   Source

   Memdouh S, Gavrilovic I, Ng K, Cowan D, Abbate V. Advances in the detection of growth hormone releasing hormone synthetic analogs. Drug Test Anal. 2021;13(11-12):1871-1887. DOI: 10.1002/dta.3183. PMID: 34665524

   PubMed 34665524

   How this citation was checked

   Re-resolved live in this pass. Crossref API on DOI 10.1002/dta.3183 returned the exact title, all five authors in order (Memdouh first, Abbate last), Drug Testing and Analysis, 2021, volume 13, issue 11-12, pages 1871-1887. PubMed efetch on PMID 34665524 returned identical bibliographic data, and because the PubMed abstract truncates mid-sentence we additionally pulled the EuropePMC core record for the same PMID and read the complete abstract, which states verbatim 'This study investigates the in vitro metabolism and detection of four of the larger GHRH synthetic analogs (sermorelin, tesamorelin, CJC-1295, and CJC-1295 with drug affinity complex) in fortified urine'; 'Nineteen major in vitro metabolites were identified, selected for synthesis, purified, and characterized in house'; and 'Limits of detection of the target peptides were generally 1 ng/ml (WADA required performance limit) or less'. PRECISION EDIT APPLIED IN THIS PASS: the model field previously read 'human matrix'; the paper specifies fortified urine, and the model field now says so.

10. Studied for

    Equine anti-doping detection methods for CJC-1295 (the albumin-binding DAC form)

    2 companion papers from the same group built the screening and confirmation methods used to catch the compound named CJC-1295 in racehorses. The screen is an immuno polymerase chain reaction assay that detected the compound bound to plasma protein down to 0.8 picograms per milliliter, with a practical screening threshold set at 50 picograms per milliliter because horses make their own GHRH. The confirmation method uses antibody capture, enzymatic digestion and LC-MS/MS, and identified the compound in horse plasma down to 180 picograms per milliliter. Included because analytical detectability is a verifiability fact about the molecule, not a claim about any biological effect. Scope note: both papers describe their target as a peptide carrying a reactive maleimidopropionic acid group that covalently links it to plasma proteins, one of them explicitly calling it a 30-amino-acid peptide. That is the albumin-binding DAC form, not the 29-residue no-DAC research peptide. analytical method development and validation (horse plasma, including samples from thoroughbred racehorses) Preclinical moderate A well-described result with several independent outcome measures, but from a single laboratory or a single study design. Open for the full finding and the paper it came from.

    In the literature

    Equine anti-doping detection methods for CJC-1295 (the albumin-binding DAC form)

    2 companion papers from the same group built the screening and confirmation methods used to catch the compound named CJC-1295 in racehorses. The screen is an immuno polymerase chain reaction assay that detected the compound bound to plasma protein down to 0.8 picograms per milliliter, with a practical screening threshold set at 50 picograms per milliliter because horses make their own GHRH. The confirmation method uses antibody capture, enzymatic digestion and LC-MS/MS, and identified the compound in horse plasma down to 180 picograms per milliliter. Included because analytical detectability is a verifiability fact about the molecule, not a claim about any biological effect. Scope note: both papers describe their target as a peptide carrying a reactive maleimidopropionic acid group that covalently links it to plasma proteins, one of them explicitly calling it a 30-amino-acid peptide. That is the albumin-binding DAC form, not the 29-residue no-DAC research peptide.^{References 10, 11}

    Modelanalytical method development and validation (horse plasma, including samples from thoroughbred racehorses)

    Confidence: moderate

    A well-described result with several independent outcome measures, but from a single laboratory or a single study design.

    Source

    Timms M, Ganio K, Steel R. A method for confirming CJC-1295 abuse in equine plasma samples by LC-MS/MS. Drug Test Anal. 2019;11(8):1248-1257. DOI: 10.1002/dta.2599. PMID: 30938069. Companion screen: Timms M, Ganio K, Forbes G, Bailey S, Steel R. An immuno polymerase chain reaction screen for the detection of CJC-1295 and other growth-hormone-releasing hormone analogs in equine plasma. Drug Test Anal. 2019;11(6):804-812. DOI: 10.1002/dta.2554. PMID: 30489688

    PubMed 30938069 PubMed 30489688

    How this citation was checked

    Both re-resolved live in this pass. Crossref API on DOI 10.1002/dta.2599 returned the exact title, authors Timms M, Ganio K, Steel R, Drug Testing and Analysis, 2019, volume 11, issue 8, pages 1248-1257. Crossref API on DOI 10.1002/dta.2554 returned the exact title, authors Timms M, Ganio K, Forbes G, Bailey S, Steel R, same journal, volume 11, issue 6, pages 804-812. PubMed efetch on PMIDs 30938069 and 30489688 returned matching bibliographic data for both plus their abstracts. Read verbatim: from the confirmation paper, 'It incorporates a functional maleimido group at the C-terminus that allows it to covalently bind plasma proteins such as serum albumin' and 'CJC-1295 was identified down to concentrations as low as 180 pg/mL in 1 mL of equine plasma'; from the screen paper, 'CJC-1295 is a 30 amino acid peptide-based drug', 'an immuno-polymerase chain reaction (I-PCR) assay that is capable of detecting the CJC-1295-protein conjugate at concentrations down to 0.8 pg/mL', 'Detection of endogenous equine GHRH necessitated a screening threshold for CJC-1295 in equine plasma of 50 pg/mL', and 'confirmed in equine blood samples after administration in thoroughbred race horses'. EDIT APPLIED IN THIS PASS: the DAC scope note is new. The source text makes clear the analytical target is the maleimido, albumin-conjugating form, which the earlier draft did not disclose in this entry.

11. Studied for

    Why the position-2 substitution exists: DPP-IV is the enzyme that cleaves GHRH

    Researchers wanted to know what actually destroys the body's own growth-hormone-releasing hormone once it enters the bloodstream. Incubating the hormone in human plasma, they found it is snipped at the bond between residues 2 and 3, producing a shortened fragment. That snipping was blocked by diprotin A, a competitive inhibitor of the enzyme dipeptidyl peptidase IV, which identifies DPP-IV as the enzyme responsible. Crucially for this research peptide, the paper also reported that swapping in a mirror-image (D) amino acid at either position 1 or position 2 prevented the cleavage entirely. A separate, slower trypsin-like plasma activity was described as a secondary degradation route. This is the published biochemical reason the D-Ala sits at position 2 in this molecule. IN VITRO in vitro (incubations of human plasma) Preclinical preliminary A single study, an early mechanistic observation, or a result whose interpretation is the authors' own inference. Open for the full finding and the paper it came from.

    In the literature

    Why the position-2 substitution exists: DPP-IV is the enzyme that cleaves GHRH

    Researchers wanted to know what actually destroys the body's own growth-hormone-releasing hormone once it enters the bloodstream. Incubating the hormone in human plasma, they found it is snipped at the bond between residues 2 and 3, producing a shortened fragment. That snipping was blocked by diprotin A, a competitive inhibitor of the enzyme dipeptidyl peptidase IV, which identifies DPP-IV as the enzyme responsible. Crucially for this research peptide, the paper also reported that swapping in a mirror-image (D) amino acid at either position 1 or position 2 prevented the cleavage entirely. A separate, slower trypsin-like plasma activity was described as a secondary degradation route. This is the published biochemical reason the D-Ala sits at position 2 in this molecule.^{References 12}

    Modelin vitro (incubations of human plasma)

    Confidence: preliminary

    A single study, an early mechanistic observation, or a result whose interpretation is the authors' own inference.

    Source

    Frohman LA, Downs TR, Heimer EP, Felix AM. Dipeptidylpeptidase IV and trypsin-like enzymatic degradation of human growth hormone-releasing hormone in plasma. J Clin Invest. 1989;83(5):1533-1540. DOI: 10.1172/JCI114049. PMID: 2565342

    PubMed 2565342

    How this citation was checked

    Re-resolved live in this pass. Crossref API on DOI 10.1172/JCI114049 returned the exact title, authors Frohman LA, Downs TR, Heimer EP, Felix AM, Journal of Clinical Investigation, 1989, volume 83, issue 5, pages 1533-1540. PubMed efetch on PMID 2565342 returned identical bibliographic data and the abstract, confirming verbatim 'Conversion to GRH(3-44)-NH2 was blocked by diprotin A, a DPP type IV (DPP IV) competitive inhibitor', 'D-Amino acid substitution at either position 1 or 2 also prevented hydrolysis, characteristic of DPP IV', and 'GRH degradation in plasma occurs primarily by DPP IV, and to a lesser extent by trypsin-like enzyme(s)'. Because the abstract does not name the plasma species, we downloaded the free full-text PDF from jci.org and confirmed the matrix in the Methods and Results text, which states the peptides were 'added to human plasma' and describes incubating 'shortened forms of GRH with human plasma in vitro'.

12. Studied for

    Quantified in vitro plasma stability of the unmodified parent fragment and substituted analogs

    The unmodified 1-29 fragment was incubated in pig plasma at body temperature, 98.6 F (37 C), and measured over time by HPLC. Half of it was gone in 13 minutes, and the main breakdown product was the fragment missing its first 2 amino acids, which pins the break to the bond between residues 2 and 3. Swapping glycine for alanine at position 15 stretched that to 17 minutes, a small gain. The paper also reported that removing the free amino group at the front end, or putting a mirror-image (D) amino acid at the second-to-last position from the front, completely prevented the break at the 2-3 bond in the straight-chain analogs. This independently corroborates, in a second species and by a different method, the cleavage site characterized in human plasma by Frohman 1989. Important: these are enzyme-driven stability numbers in a biological fluid. They say nothing about how long the material lasts in a vial. IN VITRO in vitro (incubations of pig plasma at 98.6 F / 37 C) Preclinical preliminary A single study, an early mechanistic observation, or a result whose interpretation is the authors' own inference. Open for the full finding and the paper it came from.

    In the literature

    Quantified in vitro plasma stability of the unmodified parent fragment and substituted analogs

    The unmodified 1-29 fragment was incubated in pig plasma at body temperature, 98.6 F (37 C), and measured over time by HPLC. Half of it was gone in 13 minutes, and the main breakdown product was the fragment missing its first 2 amino acids, which pins the break to the bond between residues 2 and 3. Swapping glycine for alanine at position 15 stretched that to 17 minutes, a small gain. The paper also reported that removing the free amino group at the front end, or putting a mirror-image (D) amino acid at the second-to-last position from the front, completely prevented the break at the 2-3 bond in the straight-chain analogs. This independently corroborates, in a second species and by a different method, the cleavage site characterized in human plasma by Frohman 1989. Important: these are enzyme-driven stability numbers in a biological fluid. They say nothing about how long the material lasts in a vial.^{References 13}

    Modelin vitro (incubations of pig plasma at 98.6 F / 37 C)

    Confidence: preliminary

    A single study, an early mechanistic observation, or a result whose interpretation is the authors' own inference.

    Source

    Su CM, Jensen LR, Heimer EP, Felix AM, Pan YC, Mowles TF. In vitro stability of growth hormone releasing factor (GRF) analogs in porcine plasma. Horm Metab Res. 1991;23(1):15-21. DOI: 10.1055/s-2007-1003601. PMID: 1826667

    PubMed 1826667

    How this citation was checked

    Re-resolved live in this pass. Crossref API on DOI 10.1055/s-2007-1003601 returned the exact title, the six authors in order (Su first, Mowles last), Hormone and Metabolic Research, 1991, volume 23, issue 1, pages 15-21. PubMed efetch on PMID 1826667 returned identical bibliographic data and the abstract, confirming verbatim the 37 degrees C porcine plasma incubation with HPLC analysis, 'GRF(1-29)-NH2 was rapidly broken down in the plasma with a degradation rate of t1/2 = 13 min', 'The primary degradation product was identified as GRF(3-29)-NH2', 'Substitution of Gly15 by Ala15 slightly prolonged the plasma half-life (t1/2 = 17 min)', and 'Absence of the free amino group at the N-terminus and/or substitution of a D-amino acid residue at the penultimate position completely prevented cleavage between the 2 and 3 position in the structural linear GRF analogs'. The prior draft's looser paraphrase 'D-Ala2 substitution conferred additional stability' has been replaced with the paper's own wording, because in this abstract the explicit D-Ala2 additivity statement is made about the CYCLIC analogs, not the linear ones.

2 strong / 8 moderate / 2 preliminaryRated by how many independent labs and how many kinds of experiment found the same thing.