Studies on the characterization of inhibin and inhibin-like factors have depended for the most pa... more Studies on the characterization of inhibin and inhibin-like factors have depended for the most part on the classical in vitro pituitary cell culture assay. A major drawback with this assay is the turnaround time which is in the order of two weeks and consequently slows down purification efforts. The 24 h bioassay for inhibin has been found to be sufficiently sensitive and also statistically valid. Unfortunately, based as it is on a secondary response, ambiguities arise in interpreting the results. By including a parallel assay in which the mice are primed with human menopausal gonadotropin rather than human chorionic gonadotropin, it was possible to device the coupled bioassay. This enables distinguishing inhibin-like factors acting to suppress pituitary follicle stimulating hormone output from those acting at the level of gonads. In this study the coupled assay for inhibin has been compared with the classical pituitary cell culture assay in order to assess its biological and statistical validity. The data validates the bioassay on both the above counts and when considered in conjunction with the short turnaround time suggests that this assay can be highly useful in studies on isolation of inhibin from various sources.
Journal of controlled release : official journal of the Controlled Release Society, Jan 17, 2015
Women urgently need a self-initiated, multipurpose prevention technology (MPT) that simultaneousl... more Women urgently need a self-initiated, multipurpose prevention technology (MPT) that simultaneously reduces their risk of acquiring HIV-1, HSV-2, and HPV (latter two associated with increased risk of HIV-1 acquisition) and prevent unintended pregnancy. Here, we describe a novel core-matrix intravaginal ring (IVR), the MZCL IVR, which effectively delivered the MZC combination microbicide and a contraceptive. The MZCL IVR contains four active pharmaceutical ingredients (APIs): MIV-150 (targets HIV), zinc acetate (ZA; targets HIV and HSV-2), carrageenan (CG; targets HPV and HSV-2), and levonorgestrel (LNG; targets unintended pregnancy). The elastomeric IVR body (matrix) was produced by hot melt extrusion of the non-water swellable elastomer, ethylene vinyl acetate (EVA-28), containing the hydrophobic small molecules, MIV-150 and LNG. The solid hydrophilic core, embedded within the IVR by compression, contained the small molecule ZA and a macromolecule CG. Hydrated ZA/CG from the core wa...
Multiple Sclerosis affects mainly women and consists in intermittent or chronic damages to the my... more Multiple Sclerosis affects mainly women and consists in intermittent or chronic damages to the myelin sheaths, focal inflammation, and axonal degeneration. Current therapies are limited to immunomodulators and antiinflammatory drugs, but there is no efficient treatment for stimulating the endogenous capacity of myelin repair. Progesterone and synthetic progestins have been shown in animal models of demyelination to attenuate myelin loss, reduce clinical symptoms severity, modulate inflammatory responses and partially reverse the age-dependent decline in remyelination. Moreover, progesterone has been demonstrated to promote myelin formation in organotypic cultures of cerebellar slices. In the present study, we show that progesterone and the synthetic 19-nor-progesterone derivative Nestorone® promote the repair of severe chronic demyelinating lesions induced by feeding cuprizone to female mice for up to 12 weeks. Progesterone and Nestorone increase the density of NG2(+) oligodendrocyt...
The Journal of Clinical Endocrinology & Metabolism, 2012
Context: Combinations of testosterone (T) and nestorone (NES; a nonandrogenic progestin) transder... more Context: Combinations of testosterone (T) and nestorone (NES; a nonandrogenic progestin) transdermal gels may suppress spermatogenesis and prove appealing to men for contraception. Objective: The objective of the study was to determine the effectiveness of T gel alone or combined with NES gel in suppressing spermatogenesis. Design and Setting: This was a randomized, double-blind, comparator clinical trial conducted at two academic medical centers. Participants: Ninety-nine healthy male volunteers participated in the study. Interventions: Volunteers were randomized to one of three treatment groups applying daily transdermal gels (group 1: T gel 10 g ϩ NES 0 mg/placebo gel; group 2: T gel 10 g ϩ NES gel 8 mg; group 3: T gel 10 g ϩ NES gel 12 mg). Main Outcome Variable: The main outcome variable of the study was the percentage of men whose sperm concentration was suppressed to 1 million/ml or less by 20-24 wk of treatment. Results: Efficacy data analyses were performed on 56 subjects who adhered to the protocol and completed at least 20 wk of treatment. The percentage of men whose sperm concentration was 1 million/ml or less was significantly higher for T ϩ NES 8 mg (89%, P Ͻ 0.0001) and T ϩ NES 12 mg (88%, P ϭ 0.0002) compared with T ϩ NES 0 mg group (23%). The median serum total and free T concentrations in all groups were maintained within the adult male range throughout the treatment period. Adverse effects were minimal in all groups. Conclusion: A combination of daily NES ϩ T gels suppressed sperm concentration to 1 million/ml or less in 88.5% of men, with minimal adverse effects, and may be further studied as a male transdermal hormonal contraceptive. (J Clin Endocrinol Metab 97: 3476-3486, 2012) N early 40% of pregnancies worldwide are unintended, which may result in undesirable outcomes including maternal mortality and morbidity, societal burden, and child abuse. Although many female contraceptive methods are available, discontinuation rates are high and hormonal methods may be contraindicated for many women (1). Male methods are limited to withdrawal, condom, and vasectomy. Condoms suffer from inconsistent use,
The Journal of Clinical Endocrinology & Metabolism, 2009
Context: Testosterone (T) plus progestin combinations are the most promising hormonal male contra... more Context: Testosterone (T) plus progestin combinations are the most promising hormonal male contraceptives. Nestorone (NES), a progestin without estrogenic or androgenic activity, when combined with T may be an excellent candidate for male contraception. Objective: Our objective was to determine the effect of transdermal NES gel alone or with T gel on gonadotropin suppression. Design and Setting: The randomized, unblinded clinical trial was conducted at two academic medical centers. Participants: A total of 140 healthy male volunteers participated. Interventions: One hundred subjects were randomized initially (20 per group) to apply NES gel 2 or 4 mg, T gel 10 g, or T gel 10 g plus NES gel 2 or 4 mg daily for 20 d. Because only about half of the subjects in T plus NES 4 mg group suppressed serum gonadotropins to 0.5 IU/liter or less (suboptimal suppression), two additional groups of 20 men were randomized to apply daily T gel 10 g plus NES gel 6 or 8 mg. Main Outcome Variable: Suppression of serum LH and FSH concentrations to 0.5 IU/liter or less after treatment was the main outcome variable. Results: A total of 119 subjects were compliant with gel applications with few study-related adverse events. NES alone reduced gonadotropins significantly but less than T gel alone. Combined T gel 10g plus NES gel 6 or 8 mg suppressed both serum gonadotropins to 0.5 IU/liter or less in significantly more men than either gel alone. Conclusion: Transdermal NES gel alone had gonadotropin suppression activity. Combined transdermal NES (6 or 8 mg) plus T gel demonstrated safe and effective suppression of gonadotropins, justifying a longer-term study of this combination for suppression of spermatogenesis.
The cytochrome P450 2A1 (CYP2A1) is a P450 enzyme that catalyzes the metabolism of testosterone. ... more The cytochrome P450 2A1 (CYP2A1) is a P450 enzyme that catalyzes the metabolism of testosterone. CYP2A1 has been reported to be present in rat testis. However, its developmental changes and function have not been well characterized. The purpose of this study was to measure the abundance of CYP2A1 (Cyp2a1) mRNA in the developing rat testis and Leydig cells and examine the effects of its product, 7α-hydroxytestosterone (7HT), on an important enzyme, 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) that interconverts active corticosterone and inactive 11-dehydrocorticosterone. As detected by real-time PCR, Cyp2a1 was found to be present exclusively in the Leydig cell. CYP2A1 activity in adult Leydig cells was 5-fold higher than those in progenitor or immature Leydig cells. 7HT competitively suppressed 11β-HSD1 oxidase and reductase activities in rat testis microsome with inhibitory constant of 1.2 and 2.9 μm, respectively. In intact Leydig cells, 7HT did not inhibit 11β-HSD1 reductas...
Enhancing the endogenous capacity of myelin repair is a major therapeutic challenge in demyelinat... more Enhancing the endogenous capacity of myelin repair is a major therapeutic challenge in demyelinating diseases such as multiple sclerosis. We found that progesterone and the synthetic 19-norprogesterone derivative 16-methylene-17␣-acetoxy-19-norpregn-4-ene-3,20-dione (Nestorone) promote the remyelination of axons by oligodendrocytes after lysolecithin-induced demyelination in organotypic cultures of cerebellar slices taken from postnatal rats or mice. The intracellular progesterone receptors (PR) mediate the proremyelinating actions of Nestorone, because they are not observed in slices from PR knockout mice. Notably, Nestorone was less efficient in heterozygous mice, expressing reduced levels of PR, suggesting PR haploinsufficiency in myelin repair. Using mice expressing the enhanced green fluorescent protein (EGFP) under the control of the proteolipid gene promoter, we showed that both progesterone and Nestorone strongly increased the reappearance of cells of the oligodendroglial lineage in the demyelinated slices. In contrast to Nestorone, the pregnane derivative medroxyprogesterone acetate had no effect. The increase in oligodendroglial cells by Nestorone resulted from enhanced NG2 ϩ and Olig2 ϩ oligodendrocyte progenitor cell (OPC) recruitment. In cocultures of lysolecithin-demyelinated cerebellar slices from wild-type mice apposed to brain stem slices of proteolipid gene promoter-EGFP mice, Nestorone stimulated the migration of OPC towards demyelinated axons. In this coculture paradigm, Nestorone indeed markedly increased the number of EGFP ϩ cells migrating into the demyelinated cerebellar slices. Our results show that Nestorone stimulates the recruitment and maturation of OPC, two steps which are limiting for efficient myelin repair. They may thus open new perspectives for the use of progestins, which selectively target PR, to promote the endogenous regeneration of myelin.
Myelin regeneration is a major therapeutic goal in demyelinating diseases, and the failure to rem... more Myelin regeneration is a major therapeutic goal in demyelinating diseases, and the failure to remyelinate rapidly has profound consequences for the health of axons and for brain function. However, there is no efficient treatment for stimulating myelin repair, and current therapies are limited to anti-inflammatory agents. Males are less likely to develop multiple sclerosis than females, but often have a more severe disease course and reach disability milestones at an earlier age than females, and these observations have spurred interest in the potential protective effects of androgens. Here, we demonstrate that testosterone treatment efficiently stimulates the formation of new myelin and reverses myelin damage in chronic demyelinated brain lesions, resulting from the long-term administration of cuprizone, which is toxic for oligodendrocytes. In addition to the strong effect of testosterone on myelin repair, the number of activated astrocytes and microglial cells returned to low control levels, indicating a reduction of neuroinflammatory responses. We also identify the neural androgen receptor as a novel therapeutic target for myelin recovery. After the acute demyelination of cerebellar slices in organotypic culture, the remyelinating actions of testosterone could be mimicked by 5a-dihydrotestosterone, a metabolite that is not converted to oestrogens, and blocked by the androgen receptor antagonist flutamide. Testosterone treatment also failed to promote remyelination after chronic cuprizoneinduced demyelination in mice with a non-functional androgen receptor. Importantly, testosterone did not stimulate the formation of new myelin sheaths after specific knockout of the androgen receptor in neurons and macroglial cells. Thus, the neural brain androgen receptor is required for the remyelination effect of testosterone, whereas the presence of the receptor in microglia and in peripheral tissues is not sufficient to enhance remyelination. The potent synthetic testosterone analogue 7a-methyl-19-nortestosterone, which has been developed for long-term male contraception and androgen replacement therapy in hypogonadal males and does not stimulate prostate growth, also efficiently promoted myelin repair. These data establish the
Purpose-Overt male hypogonadism induces not only osteoporosis but also unfavorable changes in bod... more Purpose-Overt male hypogonadism induces not only osteoporosis but also unfavorable changes in body composition, which can be prevented by testosterone (T) replacement. In this preclinical study, the potential of synthetic androgen 7α-methyl-19-nortestosterone (MENT) as alternative treatment for male hypogonadism was evaluated in comparison with T. Methods-11-month-old male rats were orchidectomized (orch) and left untreated for 2-months. Subsequently, the effects of 4-months MENT (12 µg/day) and T (72µg/day) treatment on bone, muscle and fat were analyzed by microcomputed tomography, dual-energy X-ray absorptiometry, dynamic bone histomorphometry and muscle fiber typing. Results-At the onset of treatment orch rats were clearly hypogonadal. This was evidenced by significant reductions of androgen-sensitive organ weight, lean mass, cortical thickness and trabecular bone volume compared with sham-operated aged-matched controls (sham). MENT and T restored weight of androgen-sensitive organs to a similar extent, with a superior anabolic action of MENT on levator ani muscle. Both androgens not only fully rescued hypogonadal loss of lean mass, but also restored muscle fiber type composition and trabecular bone volume. Cortical bone loss was similarly prevented by MENT and T, but without full recovery to sham. Both androgens stimulated periosteal bone formation, but with a stronger effect of T. In contrast, MENT more strongly suppressed endocortical bone formation and bone turnover rate and reduced fat mass and serum leptin to a greater extent than T. Conclusion-MENT and T are both effective replacement therapies to stimulate bone and muscle in hypogonadal rats, with stronger lipolytic action of MENT.
Antimicrobial agents and chemotherapy, Jan 6, 2016
The global dissemination amidst increasing incidence of carbapenem-resistant, Gram-negative organ... more The global dissemination amidst increasing incidence of carbapenem-resistant, Gram-negative organisms has resulted in acute public health concerns. Here, we present a retrospective multicenter study on molecular characterization of metallo-beta-lactamase (MBL) producing clinical E. coli isolates recovered from extraintestinal infections in two hospitals in Pune, India. We screened a large sample size of 510 E. coli isolates for MBL production wherein we profiled their molecular determinants, antimicrobial resistance phenotypes, functional virulence properties, genomic features and transmission dynamics. Approximately ∼8% of these isolates were MBL producers, the majority of which were NDM-1 (69%) type followed subsequently by NDM-5 (19%), NDM-4 (5.5%) and NDM-7 (5.5%). MBL producers were resistant to all antibiotics tested, except for colistin, fosfomycin and chloramphenicol which were effective up to a varying extent. Plasmids were found to be an effective means of dissemination of...
Some methods of contraception involve the use of drugs that affect the secretion of hormones esse... more Some methods of contraception involve the use of drugs that affect the secretion of hormones essential for repro-duction. Combined oral contraceptives are used as inhi-bitors of pituitary gonadotrophins secretion, ovarian hormone secretion and ovulation. The oestrogens and ...
Plasma cell leukemia (PCL) is a rare and aggressive plasma cell neoplasm. In PCL, clonal plasma c... more Plasma cell leukemia (PCL) is a rare and aggressive plasma cell neoplasm. In PCL, clonal plasma cells comprise ≥20% of the peripheral blood (PB) leukocytes and/or the absolute clonal PB plasma cell count is ≥2×10(9)/L. Primary PCL (PPCL) originates de novo, whereas, secondary PCL (SPCL) evolves from pre-existing multiple myeloma. Clinicohematological features, immunophenotypic profile, and survival of PCL patients were analyzed retrospectively. Between January 2007 and December 2014, ten PPCL and four SPCL patients were investigated (8 PPCLs and 3 SPCLs had complete clinical data). All were North Indians, sharing common geography and ethnicity. Our cohort showed less frequent renal failure, more frequent hepatomegaly, and non-secretory type disease. In contrast to western literature, flow cytometric immunophenotyping of our cohort revealed altered expression of CD138 (67%), CD56 (33%), and CD20 (0%). With novel therapeutic agents, these PPCL patients had a median overall survival of 15 months. We highlight that our PPCL patients from North India had distinct clinicohematological and immunophenotypic profiles. The significance of our findings must be tested in a larger patient cohort and must be supported by molecular and cytogenetic investigations to unmask possible significant effects on pathogenesis.
The action of testosterone (T) on the sex accessory organs, such as ventral prostate (VP) and sem... more The action of testosterone (T) on the sex accessory organs, such as ventral prostate (VP) and seminal vesicles (SV) is amplified by its 5α-reduction to dihydrotestosterone (DHT). This does not happen in the case of muscle (levator ani, LA) which contains little or no 5α-reductase activity. It has been suggested that the regulation of gonadotropins by T may also be mediated by its 5α-reduced metabolites. We investigated this question by utilizing two types of androgens: (1) T and 17α-methyl-testosterone (17MT), whose potency increases following 5α-reduction; and (2) 19-nortestosterone (NT) and 17α-methyl-19-nortestosterone (17MNT) whose potency decreases following 5α-reduction. Castrated rats were used to investigate the ability of these androgens to stimulate VP, and SV (androgenic action) and LA growth (anabolic action) and to suppress the post-castration rise in LH levels. In addition, modification of these actions by a 5α-reductase inhibitor (5α-RI) was studied. Compared to T, NT was approximately 5 times less potent in stimulating VP and SV. By contrast, it was twice as potent as T in stimulating LA growth. Similarly, 17MNT was 5 times less androgenic but twice as anabolic as 17MT. The antigonadotropic potency of both the 19-nor compounds was 2–3 times greater than that of their respective 19-methylated parent compounds. The similarity in their anabolic and antigonadotropic potency suggested that 5α-reduction is not a factor in their antigoanabolic and antigonadotropic potency suggested that 5α-reduction is not a factor in their antigonadotropic action. This was confirmed by the use of the 5α-RI. Treatment of rats receiving the androgens with 5α-RI showed that it decreases the androgenic activity of T and 17MT while it increases the androgenic activity of NT and 17 MNT. In all cases the anabolic activity and the antigonadotropic potency remained unchanged. It is concluded that the regulation of pituitary gonadotropin secretion by T does not depend upon its 5α-reduction to DHT.
Some methods of contraception involve the use of drugs that affect the secretion of hormones esse... more Some methods of contraception involve the use of drugs that affect the secretion of hormones essential for repro-duction. Combined oral contraceptives are used as inhi-bitors of pituitary gonadotrophins secretion, ovarian hormone secretion and ovulation. The oestrogens and ...
This study reports the preclinical evaluation of the bone and muscle protective potential of the ... more This study reports the preclinical evaluation of the bone and muscle protective potential of the synthetic androgen 7α-methyl-19-nortestosterone (MENT™), as assessed in the aged orchidectomized rat model. Aged (13-month-old) orchidectomized Wistar rats were treated with different doses of MENT (4, 12 or 36 μg/day) subcutaneously for 16 weeks via mini-osmotic pumps. Analysis of the effects of androgen deficiency versus MENT replacement was performed using quantitative computed tomography (pQCT), dual energy X-ray absorptiometry (DEXA) and biochemical markers of bone turnover. At the end of the study period, prostate weight in orchidectomized rats treated with low- (4 μg/day) or mid-dose (12 μg/day) MENT remained significantly lower compared to the sham-operated animals (−47% and −25%, respectively). High-dose MENT (36 μg/day), on the other hand, induced prostate hypertrophy (+21% versus sham). Low-, mid- and high-dose MENT were found to be effective in suppressing the acceleration of bone remodeling following orchidectomy, as assessed by osteocalcin and deoxypyridinoline. In addition, low-, mid- and high-dose were able to prevent the orchidectomy-induced bone loss, as evaluated by DEXA at the femur and total-body and by pQCT at the femur. Compared to sham-operated animals, the low- and mid-dose MENT groups showed no decline in lean body mass and no muscle atrophy (as measured by m. quadriceps weight) at 16 weeks, whereas high-dose MENT was associated with a significant decline in lean body mass (−8.5% versus sham) and quadriceps weight (−10.6%). We conclude that, in the aged orchidectomized rat model, low- and mid-doses of the synthetic androgen MENT have bone and muscle protective effects and do not induce prostate hypertrophy. The bone protective action of high-dose MENT, however, occurs at the expense of muscle wasting and prostate hypertrophy. Our findings support the need for human studies to explore the potential of MENT as an option for androgen replacement in aging men.
Hairy cell leukaemia (HCL) is a B cell neoplasm which constitutes around 2 per cent of all the ly... more Hairy cell leukaemia (HCL) is a B cell neoplasm which constitutes around 2 per cent of all the lymphoid leukaemias. It has a characteristic morphology and immunophenotypic profile. It is important to distinguish HCL from other B cell lymphoproliferative disorders due to availability of different chemotherapeutic agents. This study presents clinical, haematological and immunophenotypic profile of patients with HCL seen over a period of four years in a tertiary care hospital in north India. Twenty one cases of hairy cell leukaemia were analyzed for their clinical details, haemogram, bone marrow examination and immunophenotypic findings. Age of the patients ranged from 28-76 yr with male predominance. Weakness and fever were commonest presentations. Splenomegaly, hepatomegaly, lymphadenopathy were seen in decreasing order of frequency. Anaemia was noted in all 21 patients, leukopenia in 15 and thrombocytopenia in 19 cases. Fourteen patients were pancytopenic. Bone marrow examination sh...
Determine the lowest effective of three Nestorone (NES)/estradiol (E2) transdermal gel doses to e... more Determine the lowest effective of three Nestorone (NES)/estradiol (E2) transdermal gel doses to ensure ovulation suppression in 90-95% of cycles. This was a randomized, open-label, three-treatment period crossover study to evaluate the effects of NES/E2 transdermal gel on ovulation inhibition, suppression of follicular growth and pharmacokinetic parameters. The doses were: low (1.5mg NES/0.5mg E2), medium (3.0mg NES/1.0mg E2) and high (4.5mg NES/1.5mg E2). Participants applied gel daily to a fixed area on the abdomen for 21 consecutive days. They were interviewed regarding their experiences using the gel. Eighteen participants were randomized; 16 completed the study. Median NES Cmax values for low, medium and high dose at day 21 were: 318.6 pmol/L, 783.0 pmol/L and 1063.8 pmol/L respectively. Median maximum follicular diameter was higher with the lowest dose with 16.2 mm versus 10.0 and 10.4 mm with the medium and high doses respectively. Among adherent participants, ovulation was i...
Studies on the characterization of inhibin and inhibin-like factors have depended for the most pa... more Studies on the characterization of inhibin and inhibin-like factors have depended for the most part on the classical in vitro pituitary cell culture assay. A major drawback with this assay is the turnaround time which is in the order of two weeks and consequently slows down purification efforts. The 24 h bioassay for inhibin has been found to be sufficiently sensitive and also statistically valid. Unfortunately, based as it is on a secondary response, ambiguities arise in interpreting the results. By including a parallel assay in which the mice are primed with human menopausal gonadotropin rather than human chorionic gonadotropin, it was possible to device the coupled bioassay. This enables distinguishing inhibin-like factors acting to suppress pituitary follicle stimulating hormone output from those acting at the level of gonads. In this study the coupled assay for inhibin has been compared with the classical pituitary cell culture assay in order to assess its biological and statistical validity. The data validates the bioassay on both the above counts and when considered in conjunction with the short turnaround time suggests that this assay can be highly useful in studies on isolation of inhibin from various sources.
Journal of controlled release : official journal of the Controlled Release Society, Jan 17, 2015
Women urgently need a self-initiated, multipurpose prevention technology (MPT) that simultaneousl... more Women urgently need a self-initiated, multipurpose prevention technology (MPT) that simultaneously reduces their risk of acquiring HIV-1, HSV-2, and HPV (latter two associated with increased risk of HIV-1 acquisition) and prevent unintended pregnancy. Here, we describe a novel core-matrix intravaginal ring (IVR), the MZCL IVR, which effectively delivered the MZC combination microbicide and a contraceptive. The MZCL IVR contains four active pharmaceutical ingredients (APIs): MIV-150 (targets HIV), zinc acetate (ZA; targets HIV and HSV-2), carrageenan (CG; targets HPV and HSV-2), and levonorgestrel (LNG; targets unintended pregnancy). The elastomeric IVR body (matrix) was produced by hot melt extrusion of the non-water swellable elastomer, ethylene vinyl acetate (EVA-28), containing the hydrophobic small molecules, MIV-150 and LNG. The solid hydrophilic core, embedded within the IVR by compression, contained the small molecule ZA and a macromolecule CG. Hydrated ZA/CG from the core wa...
Multiple Sclerosis affects mainly women and consists in intermittent or chronic damages to the my... more Multiple Sclerosis affects mainly women and consists in intermittent or chronic damages to the myelin sheaths, focal inflammation, and axonal degeneration. Current therapies are limited to immunomodulators and antiinflammatory drugs, but there is no efficient treatment for stimulating the endogenous capacity of myelin repair. Progesterone and synthetic progestins have been shown in animal models of demyelination to attenuate myelin loss, reduce clinical symptoms severity, modulate inflammatory responses and partially reverse the age-dependent decline in remyelination. Moreover, progesterone has been demonstrated to promote myelin formation in organotypic cultures of cerebellar slices. In the present study, we show that progesterone and the synthetic 19-nor-progesterone derivative Nestorone® promote the repair of severe chronic demyelinating lesions induced by feeding cuprizone to female mice for up to 12 weeks. Progesterone and Nestorone increase the density of NG2(+) oligodendrocyt...
The Journal of Clinical Endocrinology & Metabolism, 2012
Context: Combinations of testosterone (T) and nestorone (NES; a nonandrogenic progestin) transder... more Context: Combinations of testosterone (T) and nestorone (NES; a nonandrogenic progestin) transdermal gels may suppress spermatogenesis and prove appealing to men for contraception. Objective: The objective of the study was to determine the effectiveness of T gel alone or combined with NES gel in suppressing spermatogenesis. Design and Setting: This was a randomized, double-blind, comparator clinical trial conducted at two academic medical centers. Participants: Ninety-nine healthy male volunteers participated in the study. Interventions: Volunteers were randomized to one of three treatment groups applying daily transdermal gels (group 1: T gel 10 g ϩ NES 0 mg/placebo gel; group 2: T gel 10 g ϩ NES gel 8 mg; group 3: T gel 10 g ϩ NES gel 12 mg). Main Outcome Variable: The main outcome variable of the study was the percentage of men whose sperm concentration was suppressed to 1 million/ml or less by 20-24 wk of treatment. Results: Efficacy data analyses were performed on 56 subjects who adhered to the protocol and completed at least 20 wk of treatment. The percentage of men whose sperm concentration was 1 million/ml or less was significantly higher for T ϩ NES 8 mg (89%, P Ͻ 0.0001) and T ϩ NES 12 mg (88%, P ϭ 0.0002) compared with T ϩ NES 0 mg group (23%). The median serum total and free T concentrations in all groups were maintained within the adult male range throughout the treatment period. Adverse effects were minimal in all groups. Conclusion: A combination of daily NES ϩ T gels suppressed sperm concentration to 1 million/ml or less in 88.5% of men, with minimal adverse effects, and may be further studied as a male transdermal hormonal contraceptive. (J Clin Endocrinol Metab 97: 3476-3486, 2012) N early 40% of pregnancies worldwide are unintended, which may result in undesirable outcomes including maternal mortality and morbidity, societal burden, and child abuse. Although many female contraceptive methods are available, discontinuation rates are high and hormonal methods may be contraindicated for many women (1). Male methods are limited to withdrawal, condom, and vasectomy. Condoms suffer from inconsistent use,
The Journal of Clinical Endocrinology & Metabolism, 2009
Context: Testosterone (T) plus progestin combinations are the most promising hormonal male contra... more Context: Testosterone (T) plus progestin combinations are the most promising hormonal male contraceptives. Nestorone (NES), a progestin without estrogenic or androgenic activity, when combined with T may be an excellent candidate for male contraception. Objective: Our objective was to determine the effect of transdermal NES gel alone or with T gel on gonadotropin suppression. Design and Setting: The randomized, unblinded clinical trial was conducted at two academic medical centers. Participants: A total of 140 healthy male volunteers participated. Interventions: One hundred subjects were randomized initially (20 per group) to apply NES gel 2 or 4 mg, T gel 10 g, or T gel 10 g plus NES gel 2 or 4 mg daily for 20 d. Because only about half of the subjects in T plus NES 4 mg group suppressed serum gonadotropins to 0.5 IU/liter or less (suboptimal suppression), two additional groups of 20 men were randomized to apply daily T gel 10 g plus NES gel 6 or 8 mg. Main Outcome Variable: Suppression of serum LH and FSH concentrations to 0.5 IU/liter or less after treatment was the main outcome variable. Results: A total of 119 subjects were compliant with gel applications with few study-related adverse events. NES alone reduced gonadotropins significantly but less than T gel alone. Combined T gel 10g plus NES gel 6 or 8 mg suppressed both serum gonadotropins to 0.5 IU/liter or less in significantly more men than either gel alone. Conclusion: Transdermal NES gel alone had gonadotropin suppression activity. Combined transdermal NES (6 or 8 mg) plus T gel demonstrated safe and effective suppression of gonadotropins, justifying a longer-term study of this combination for suppression of spermatogenesis.
The cytochrome P450 2A1 (CYP2A1) is a P450 enzyme that catalyzes the metabolism of testosterone. ... more The cytochrome P450 2A1 (CYP2A1) is a P450 enzyme that catalyzes the metabolism of testosterone. CYP2A1 has been reported to be present in rat testis. However, its developmental changes and function have not been well characterized. The purpose of this study was to measure the abundance of CYP2A1 (Cyp2a1) mRNA in the developing rat testis and Leydig cells and examine the effects of its product, 7α-hydroxytestosterone (7HT), on an important enzyme, 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) that interconverts active corticosterone and inactive 11-dehydrocorticosterone. As detected by real-time PCR, Cyp2a1 was found to be present exclusively in the Leydig cell. CYP2A1 activity in adult Leydig cells was 5-fold higher than those in progenitor or immature Leydig cells. 7HT competitively suppressed 11β-HSD1 oxidase and reductase activities in rat testis microsome with inhibitory constant of 1.2 and 2.9 μm, respectively. In intact Leydig cells, 7HT did not inhibit 11β-HSD1 reductas...
Enhancing the endogenous capacity of myelin repair is a major therapeutic challenge in demyelinat... more Enhancing the endogenous capacity of myelin repair is a major therapeutic challenge in demyelinating diseases such as multiple sclerosis. We found that progesterone and the synthetic 19-norprogesterone derivative 16-methylene-17␣-acetoxy-19-norpregn-4-ene-3,20-dione (Nestorone) promote the remyelination of axons by oligodendrocytes after lysolecithin-induced demyelination in organotypic cultures of cerebellar slices taken from postnatal rats or mice. The intracellular progesterone receptors (PR) mediate the proremyelinating actions of Nestorone, because they are not observed in slices from PR knockout mice. Notably, Nestorone was less efficient in heterozygous mice, expressing reduced levels of PR, suggesting PR haploinsufficiency in myelin repair. Using mice expressing the enhanced green fluorescent protein (EGFP) under the control of the proteolipid gene promoter, we showed that both progesterone and Nestorone strongly increased the reappearance of cells of the oligodendroglial lineage in the demyelinated slices. In contrast to Nestorone, the pregnane derivative medroxyprogesterone acetate had no effect. The increase in oligodendroglial cells by Nestorone resulted from enhanced NG2 ϩ and Olig2 ϩ oligodendrocyte progenitor cell (OPC) recruitment. In cocultures of lysolecithin-demyelinated cerebellar slices from wild-type mice apposed to brain stem slices of proteolipid gene promoter-EGFP mice, Nestorone stimulated the migration of OPC towards demyelinated axons. In this coculture paradigm, Nestorone indeed markedly increased the number of EGFP ϩ cells migrating into the demyelinated cerebellar slices. Our results show that Nestorone stimulates the recruitment and maturation of OPC, two steps which are limiting for efficient myelin repair. They may thus open new perspectives for the use of progestins, which selectively target PR, to promote the endogenous regeneration of myelin.
Myelin regeneration is a major therapeutic goal in demyelinating diseases, and the failure to rem... more Myelin regeneration is a major therapeutic goal in demyelinating diseases, and the failure to remyelinate rapidly has profound consequences for the health of axons and for brain function. However, there is no efficient treatment for stimulating myelin repair, and current therapies are limited to anti-inflammatory agents. Males are less likely to develop multiple sclerosis than females, but often have a more severe disease course and reach disability milestones at an earlier age than females, and these observations have spurred interest in the potential protective effects of androgens. Here, we demonstrate that testosterone treatment efficiently stimulates the formation of new myelin and reverses myelin damage in chronic demyelinated brain lesions, resulting from the long-term administration of cuprizone, which is toxic for oligodendrocytes. In addition to the strong effect of testosterone on myelin repair, the number of activated astrocytes and microglial cells returned to low control levels, indicating a reduction of neuroinflammatory responses. We also identify the neural androgen receptor as a novel therapeutic target for myelin recovery. After the acute demyelination of cerebellar slices in organotypic culture, the remyelinating actions of testosterone could be mimicked by 5a-dihydrotestosterone, a metabolite that is not converted to oestrogens, and blocked by the androgen receptor antagonist flutamide. Testosterone treatment also failed to promote remyelination after chronic cuprizoneinduced demyelination in mice with a non-functional androgen receptor. Importantly, testosterone did not stimulate the formation of new myelin sheaths after specific knockout of the androgen receptor in neurons and macroglial cells. Thus, the neural brain androgen receptor is required for the remyelination effect of testosterone, whereas the presence of the receptor in microglia and in peripheral tissues is not sufficient to enhance remyelination. The potent synthetic testosterone analogue 7a-methyl-19-nortestosterone, which has been developed for long-term male contraception and androgen replacement therapy in hypogonadal males and does not stimulate prostate growth, also efficiently promoted myelin repair. These data establish the
Purpose-Overt male hypogonadism induces not only osteoporosis but also unfavorable changes in bod... more Purpose-Overt male hypogonadism induces not only osteoporosis but also unfavorable changes in body composition, which can be prevented by testosterone (T) replacement. In this preclinical study, the potential of synthetic androgen 7α-methyl-19-nortestosterone (MENT) as alternative treatment for male hypogonadism was evaluated in comparison with T. Methods-11-month-old male rats were orchidectomized (orch) and left untreated for 2-months. Subsequently, the effects of 4-months MENT (12 µg/day) and T (72µg/day) treatment on bone, muscle and fat were analyzed by microcomputed tomography, dual-energy X-ray absorptiometry, dynamic bone histomorphometry and muscle fiber typing. Results-At the onset of treatment orch rats were clearly hypogonadal. This was evidenced by significant reductions of androgen-sensitive organ weight, lean mass, cortical thickness and trabecular bone volume compared with sham-operated aged-matched controls (sham). MENT and T restored weight of androgen-sensitive organs to a similar extent, with a superior anabolic action of MENT on levator ani muscle. Both androgens not only fully rescued hypogonadal loss of lean mass, but also restored muscle fiber type composition and trabecular bone volume. Cortical bone loss was similarly prevented by MENT and T, but without full recovery to sham. Both androgens stimulated periosteal bone formation, but with a stronger effect of T. In contrast, MENT more strongly suppressed endocortical bone formation and bone turnover rate and reduced fat mass and serum leptin to a greater extent than T. Conclusion-MENT and T are both effective replacement therapies to stimulate bone and muscle in hypogonadal rats, with stronger lipolytic action of MENT.
Antimicrobial agents and chemotherapy, Jan 6, 2016
The global dissemination amidst increasing incidence of carbapenem-resistant, Gram-negative organ... more The global dissemination amidst increasing incidence of carbapenem-resistant, Gram-negative organisms has resulted in acute public health concerns. Here, we present a retrospective multicenter study on molecular characterization of metallo-beta-lactamase (MBL) producing clinical E. coli isolates recovered from extraintestinal infections in two hospitals in Pune, India. We screened a large sample size of 510 E. coli isolates for MBL production wherein we profiled their molecular determinants, antimicrobial resistance phenotypes, functional virulence properties, genomic features and transmission dynamics. Approximately ∼8% of these isolates were MBL producers, the majority of which were NDM-1 (69%) type followed subsequently by NDM-5 (19%), NDM-4 (5.5%) and NDM-7 (5.5%). MBL producers were resistant to all antibiotics tested, except for colistin, fosfomycin and chloramphenicol which were effective up to a varying extent. Plasmids were found to be an effective means of dissemination of...
Some methods of contraception involve the use of drugs that affect the secretion of hormones esse... more Some methods of contraception involve the use of drugs that affect the secretion of hormones essential for repro-duction. Combined oral contraceptives are used as inhi-bitors of pituitary gonadotrophins secretion, ovarian hormone secretion and ovulation. The oestrogens and ...
Plasma cell leukemia (PCL) is a rare and aggressive plasma cell neoplasm. In PCL, clonal plasma c... more Plasma cell leukemia (PCL) is a rare and aggressive plasma cell neoplasm. In PCL, clonal plasma cells comprise ≥20% of the peripheral blood (PB) leukocytes and/or the absolute clonal PB plasma cell count is ≥2×10(9)/L. Primary PCL (PPCL) originates de novo, whereas, secondary PCL (SPCL) evolves from pre-existing multiple myeloma. Clinicohematological features, immunophenotypic profile, and survival of PCL patients were analyzed retrospectively. Between January 2007 and December 2014, ten PPCL and four SPCL patients were investigated (8 PPCLs and 3 SPCLs had complete clinical data). All were North Indians, sharing common geography and ethnicity. Our cohort showed less frequent renal failure, more frequent hepatomegaly, and non-secretory type disease. In contrast to western literature, flow cytometric immunophenotyping of our cohort revealed altered expression of CD138 (67%), CD56 (33%), and CD20 (0%). With novel therapeutic agents, these PPCL patients had a median overall survival of 15 months. We highlight that our PPCL patients from North India had distinct clinicohematological and immunophenotypic profiles. The significance of our findings must be tested in a larger patient cohort and must be supported by molecular and cytogenetic investigations to unmask possible significant effects on pathogenesis.
The action of testosterone (T) on the sex accessory organs, such as ventral prostate (VP) and sem... more The action of testosterone (T) on the sex accessory organs, such as ventral prostate (VP) and seminal vesicles (SV) is amplified by its 5α-reduction to dihydrotestosterone (DHT). This does not happen in the case of muscle (levator ani, LA) which contains little or no 5α-reductase activity. It has been suggested that the regulation of gonadotropins by T may also be mediated by its 5α-reduced metabolites. We investigated this question by utilizing two types of androgens: (1) T and 17α-methyl-testosterone (17MT), whose potency increases following 5α-reduction; and (2) 19-nortestosterone (NT) and 17α-methyl-19-nortestosterone (17MNT) whose potency decreases following 5α-reduction. Castrated rats were used to investigate the ability of these androgens to stimulate VP, and SV (androgenic action) and LA growth (anabolic action) and to suppress the post-castration rise in LH levels. In addition, modification of these actions by a 5α-reductase inhibitor (5α-RI) was studied. Compared to T, NT was approximately 5 times less potent in stimulating VP and SV. By contrast, it was twice as potent as T in stimulating LA growth. Similarly, 17MNT was 5 times less androgenic but twice as anabolic as 17MT. The antigonadotropic potency of both the 19-nor compounds was 2–3 times greater than that of their respective 19-methylated parent compounds. The similarity in their anabolic and antigonadotropic potency suggested that 5α-reduction is not a factor in their antigoanabolic and antigonadotropic potency suggested that 5α-reduction is not a factor in their antigonadotropic action. This was confirmed by the use of the 5α-RI. Treatment of rats receiving the androgens with 5α-RI showed that it decreases the androgenic activity of T and 17MT while it increases the androgenic activity of NT and 17 MNT. In all cases the anabolic activity and the antigonadotropic potency remained unchanged. It is concluded that the regulation of pituitary gonadotropin secretion by T does not depend upon its 5α-reduction to DHT.
Some methods of contraception involve the use of drugs that affect the secretion of hormones esse... more Some methods of contraception involve the use of drugs that affect the secretion of hormones essential for repro-duction. Combined oral contraceptives are used as inhi-bitors of pituitary gonadotrophins secretion, ovarian hormone secretion and ovulation. The oestrogens and ...
This study reports the preclinical evaluation of the bone and muscle protective potential of the ... more This study reports the preclinical evaluation of the bone and muscle protective potential of the synthetic androgen 7α-methyl-19-nortestosterone (MENT™), as assessed in the aged orchidectomized rat model. Aged (13-month-old) orchidectomized Wistar rats were treated with different doses of MENT (4, 12 or 36 μg/day) subcutaneously for 16 weeks via mini-osmotic pumps. Analysis of the effects of androgen deficiency versus MENT replacement was performed using quantitative computed tomography (pQCT), dual energy X-ray absorptiometry (DEXA) and biochemical markers of bone turnover. At the end of the study period, prostate weight in orchidectomized rats treated with low- (4 μg/day) or mid-dose (12 μg/day) MENT remained significantly lower compared to the sham-operated animals (−47% and −25%, respectively). High-dose MENT (36 μg/day), on the other hand, induced prostate hypertrophy (+21% versus sham). Low-, mid- and high-dose MENT were found to be effective in suppressing the acceleration of bone remodeling following orchidectomy, as assessed by osteocalcin and deoxypyridinoline. In addition, low-, mid- and high-dose were able to prevent the orchidectomy-induced bone loss, as evaluated by DEXA at the femur and total-body and by pQCT at the femur. Compared to sham-operated animals, the low- and mid-dose MENT groups showed no decline in lean body mass and no muscle atrophy (as measured by m. quadriceps weight) at 16 weeks, whereas high-dose MENT was associated with a significant decline in lean body mass (−8.5% versus sham) and quadriceps weight (−10.6%). We conclude that, in the aged orchidectomized rat model, low- and mid-doses of the synthetic androgen MENT have bone and muscle protective effects and do not induce prostate hypertrophy. The bone protective action of high-dose MENT, however, occurs at the expense of muscle wasting and prostate hypertrophy. Our findings support the need for human studies to explore the potential of MENT as an option for androgen replacement in aging men.
Hairy cell leukaemia (HCL) is a B cell neoplasm which constitutes around 2 per cent of all the ly... more Hairy cell leukaemia (HCL) is a B cell neoplasm which constitutes around 2 per cent of all the lymphoid leukaemias. It has a characteristic morphology and immunophenotypic profile. It is important to distinguish HCL from other B cell lymphoproliferative disorders due to availability of different chemotherapeutic agents. This study presents clinical, haematological and immunophenotypic profile of patients with HCL seen over a period of four years in a tertiary care hospital in north India. Twenty one cases of hairy cell leukaemia were analyzed for their clinical details, haemogram, bone marrow examination and immunophenotypic findings. Age of the patients ranged from 28-76 yr with male predominance. Weakness and fever were commonest presentations. Splenomegaly, hepatomegaly, lymphadenopathy were seen in decreasing order of frequency. Anaemia was noted in all 21 patients, leukopenia in 15 and thrombocytopenia in 19 cases. Fourteen patients were pancytopenic. Bone marrow examination sh...
Determine the lowest effective of three Nestorone (NES)/estradiol (E2) transdermal gel doses to e... more Determine the lowest effective of three Nestorone (NES)/estradiol (E2) transdermal gel doses to ensure ovulation suppression in 90-95% of cycles. This was a randomized, open-label, three-treatment period crossover study to evaluate the effects of NES/E2 transdermal gel on ovulation inhibition, suppression of follicular growth and pharmacokinetic parameters. The doses were: low (1.5mg NES/0.5mg E2), medium (3.0mg NES/1.0mg E2) and high (4.5mg NES/1.5mg E2). Participants applied gel daily to a fixed area on the abdomen for 21 consecutive days. They were interviewed regarding their experiences using the gel. Eighteen participants were randomized; 16 completed the study. Median NES Cmax values for low, medium and high dose at day 21 were: 318.6 pmol/L, 783.0 pmol/L and 1063.8 pmol/L respectively. Median maximum follicular diameter was higher with the lowest dose with 16.2 mm versus 10.0 and 10.4 mm with the medium and high doses respectively. Among adherent participants, ovulation was i...
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Papers by Narender Kumar